SC INBRE Science Symposium Magazine 2023

Exhibit Hall

Molecular Analytics and Informatics Core (supported by SC INBRE)

Jeremy Barth, Medical University of South Carolina

Molecular Analytics and Informatics Core (formerly Proteogenomics) is an SC INBRE-supported core located at MUSC. The core offers 1) instrumentation for various molecular biological applications; 2) guidance and informatics support for sequencing projects.

Functional Genomics Core (the Microarray Core Facility, located within FGC, is supported by SC INBRE)

Michael Shtutman, Emily (Hao) Ji, Deigo Altomare, University of South Carolina

The FGC Facility offers state-of-the-art resources and solutions for conducting genomics, transcriptomics, epigenomics and functional genomics projects.

FGC works with researchers to determine project goals and design custom solutions. FGC assists at all stages of the projects, from support in grant development to generation of publication-quality data.

Nuclear Magnetic Resonance Facility located at the Henry N. Tisdale Research Center

Md Rahman, Brandon Yarbrough, Claflin University

Claflin University's NMR facility, in addition to its 700 MHz NMR, also has an automated nuclear magnetic resonance (NMR) sample changer to do NMR-based metabolomics research. The field of metabolomics focuses on changes to the small molecules that make up metabolism upon some stressful event or condition. NMR serves as an excellent detector of these small compounds; and statistical analysis

is applied to many NMR sample replicates to increase the robustness of the metabolomics technique. Although a predominantly undergraduate university and HBCU, Claflin University has the third highest field NMR laboratory in the state of South Carolina, just behind the Medical University of South Carolina and the Hollings Marine Laboratory.

Clemson University Genomics and Bioinformatics Facility

Dr. Rooksana Noorai, Dr. Elio Ortiz, Ms. Kaitlyn Williams, Clemson

University

The Clemson University Genomics and Bioinformatics Facility (CUGBF) helps investigators use the latest genomics technologies in their research. The heart of our facility is Illumina’s NextSeq 550 NGS platform, which offers high throughput sequencing used in many applications such as whole-genome, transcriptome, and targeted resequencing. CUGBF offers services to support the use of the NextSeq 550 from experimental design through bioinformatic deliverables. In addition to offering a full-service genomics lab and bioinformatics team, we pride ourselves in offering training to students to work alongside our staff members to learn genomic and bioinformatic techniques.

South Carolina Translational Research Improving Musculoskeletal Health (SC TRIMH) COBRE Scientific Cores Thomas Gallien,

Clemson University

The SC TRIMH Center, a National Institutes of Health Center of Biomedical Research Excellence (COBRE) funded by NIH/ NIGMS (P20GM121342), is dedicated to enhancing and expanding the Biomedical Research capacity at Clemson University and the state of South Carolina to promote outstanding multidisciplinary, collaborative, and translational research in musculoskeletal and related diseases.

SC TRIMH welcomes the addition of new participants who wish to join the center to assist us in creating a critical mass and community of investigators, at Clemson and across the state, that will accelerate the rate of discoveries in musculoskeletal and related health.

Each week, SC INBRE sends out a newsletter

full of helpful information to assist your success in your career including workshops, webinars, grants deadlines, jobs opportunities and so much more. You can access the weekly newsletter from our home page - or better yet, subscribe from the archives page located under the Resources tab on our website!

Symposium Agenda

8 am Registrant Check-In and Poster set up Beverages and limited breakfast items, Lounge Area (204)

8:30 am Opening Remarks and Introduction of DRP Program, Russell House Theater

Dr. Edie Goldsmith, USC SOMC, SC INBRE Program Director

Session 1: Developmental Research Project Program (DRP), Theater

8:45 am Dr. Meghan Breen, Furman University

9 am Daniel Stovall, Winthrop University

9:15 am Subramanya Pandruvada, Medical University of South Carolina

9:30 am Jordan Gilmore, Clemson University

Session 2: Bioinformatics Pilot Project Program (BIPP), Theater

9:45 am Introduction of BIPP Program

Dr. Holly LaVoie, USC SOMC, SC INBRE Program Coordinator

9:50 am Dr. Walden Ai, Benedict College

10 am Dr. Mohamad Azhar, USC School of Medicine Columbia

10:10 am Dr. April DeLaurier, USC Aiken

10:20 am Break — Beverage Service in Lounge Area (204)

Session 3: Three Minute Madness Flash Talks, Theater

10:45 am Introduction, Dr. Edie Goldsmith

10:50 am BI-02 : Laken Fulmber, Presbyterian College

BI-03: Jeremiah Jackson, Presbyterian College

BI-04: Elena Renshaw, University of South Carolina

CB-14: Shemyia Smith, USC Upstate

MCB-06: Ridha Fatima, USC School of Medicine Columbia

MCB-09: Megan Wilson, Presbyterian College

MCB-26: Matthew Defreitas, Coastal Carolina University

RET-01: Lakshmi Sunitha Arava, Fairfield Central High School

Poster Sessions, Exhibit Hall and Lunch

11:30 am Poster Session, Ballroom and Classrooms (odd # presenters)

Exhibit Hall, Second Floor Lobby

12:30 pm Networking Lunch, Ballroom (pick up in Lounge area, 202)

1:30 pm Poster Session, Ballroom and Classrooms (even # presenters)

Exhibit Hall, Second Floor Lobby

2:30 pm Break

Session 4: Student-Initiated Research Program (SIRP) Theater

2:45 pm Introduction of SIRP Program, Dr. Holly LaVoie

2:50 pm William Hobbs, University of South Carolina

3 pm Darby Porter, University of South Carolina

3:10 pm Duncan Nowling, Medical University of South Carolina

3:20 pm Graham Warner, Medical University of South Carolina

3:30 pm Breanna Pederson, USC School of Medicine Columbia

3:45 pm Closing Remarks and Awards Presentation, Theater

5

6

9

45

“

is to see what everybody else has seen, and to think what nobody else has thought.

Research

”

Albert Szent-Gyorgyi

Venue Maps and Location

From the Director

Welcome to our 14th Annual SC INBRE Science Symposium. We are so excited to be back meeting in-person! We are very fortunate to have a great team who was able to pivot our Symposium to a virtual format last two years so we did not have to miss presenting our Symposium. As nice as it was to be able to see everyone online, it is not the same as being able to be together face-to-face. Thank you for being here today.

The purpose of our annual Science Symposium is to bring together those who have been funded by SC INBRE to share their research of our grants programs recipients. Presenting during the Symposium are representatives of most of our 14 member and four outreach/alumni institutions. As always, we also welcome some of the teachers who participated in the Research Experiences for Teachers (RET). These middle and high school STEM teachers were directed in carrying out unique, individualized research projects by faculty mentors matched from the teacher’s own local area. See page 45 to learn more about this program (which is currently taking applications for this summer).

This year is going to be an exciting year in the life of SC INBRE! In addition to our recently-released annual Call for Proposals for our three funding opportunities, we will be opening a Call for Institutional Membership this Spring/Summer as we head into submitting our renewal for INBRE Cycle V next year. Our annual Academic Leadership and Career Development Workshop will return this June. Additionally, we will be the host of the biennial Southeast Regional IDeA Conference this September 15-17. Watch our newsletters and website as more information comes available on these.

SC INBRE would not exist without the hard work and contributions of many people across the state: the Institutional PIs at our network institutions, our Core Facilities Directors and staff, and our faculty and students. Thank you to the families of the students for their support of their student’s research. We’d like to especially recognize the members of our External Advisory Committee, Drs. Laura Furge, George Littleton, and Nancy Mills (EAC Chair) and our newest EAC member, Dr. Shafiq Khan from Clark Atlanta University, for attending our Symposium today and giving their valuable feedback to our program. We would also like to thank Dr. Merry Lindsey who stepped down from our EAC earlier this year for her years of service to our Program. Additionally, we thank the members of our Evaluation Team for their invaluable service to SC INBRE and encourage you to please fill out the survey when you are contacted by them. Your responses will make our program stronger and allow us to continue to present value-added events in the future.

And last, but not least, a big thank you to the members of the SC INBRE Program Office. Their tireless dedication, not only this Symposium, but also to the day-to-day operations of SC INBRE has no parallel: Dr. Holly LaVoie, John Clarkson, and Cyndy Buckhaults

Thank you for attending today and your support of SC INBRE!

Edie C. Goldsmith, Ph.D. SC INBRE Program Director

Find and follow us on social. Post and tweet today using #scinbre and/ or #IamSCINBRE

SC INBRE Office USC School of Medicine 6439 Garners Ferry Rd. Bldg. 1, Rm. B53 Columbia, SC 29209 http://scinbre.org

SC INBRE PROGRAM OFFICE Dr. Edie Goldsmith Program Director edie.goldsmith@uscmed.sc.edu

Dr. Holly LaVoie Program Coordinator holly.lavoie@uscmed.sc.edu

John G. Clarkson, IV, MPH Program Manager john.clarkson@uscmed.sc.edu

Cyndy Buckhaults Communications Manager cyndy.buckhaults@uscmed.sc.edu

Presentation and Flash Talk Abstracts

*Abstract embargoed

Flash Talk presenter

Underlined = presenter(s) Bold = mentor(s)

BC Benedict College

Clem Clemson Univeristy

CCU Coastal Carolina University

CofC College of Charleston

CC Columbia College

Conv Converse University

FMU Francis Marion University

FU Furman University

MUSC Medical University of South Carolina

PC Presbyterian College

SA Scholars Academy High School at CCU

USC University of South Carolina

USCA USC Aiken

USC UP USC UPSTATE

SOMC USC School of Medicine

Columbia

SOMG USC School of Medicine

Greenville

WU Winthrop University

PODIUM PRESENTERS

2021 SC INBRE GRANT RECIPIENTS

DEVELOPMENTAL RESEARCH PROJECT PROGRAM (DRP)

*DRP-01 (CB), 8:30 am: Mutation of predicted Pdr1 phosphosites restores fluconazole sensitivity in Candida glabrata, Meghan Breen, Jane McCallum, Abby Stapleton, and Elizabeth Chandler, FU

DRP-02 (MCB), 8:45 am (page 9):

The Role and Regulation of RYBP in Glioblastoma Multiforme, Daniel B. Stovall, Ronald W. Bucher, Alex Lee, Lauren Patterson, Mason Linker, Brayden Fults, Daniela Torricos, Abigail Derick, WU

DRP-03 (MCB), 9 am (page 10):

Modeling macrophage responses to periodontal infections in vitro, Nico Farrar, Waleed Al-Kakhan, Vanessa Sikora, Subramanya Pandruvada, MUSC

DRP-04 (BE), 9:15 am (page 1011): Point-of-Care Colorimetric Biosensor for Detection of Pseudomonas aeruginosa Signaling Molecule 3OC12HSL in Wound Exudate from Negative Pressure

Wound Therapy, Skylar Leslie, David Karig, Jordon Gilmore, Clem

BIOINFORMATICS PILOT PROJECT PROGRAM (BIPP)

BIPP-01 (MCB), 9:45 am (page 11): Linking trained immunity with tumor dormancy by KLF4-mediated cell fusion, Arianna Bastian1, Joshua Garbutt1, Mitzi Nagarkatti2, Daping Fan3, Walden Ai1,3 , 1BC, 2SOMC, 3SOMC

*BIPP-02 (BI), 9:55 am: Identification of Differentially Expressed Genes Involved in Thoracic Aortopathy, Mengistu Gebere, Mrinmay Chakrabarti, John Johnson, Mohamad Azhar, SOMC

*BIPP-03 (MCB), 10:05 am: Establishing targets of hdac1 function in zebrafish using RNAseq, DeLaurier, A1; DeLorenzo, L2; Mohammed, AD3; Mishoe Hernandez, L1; Willoner, TJ1; Jones, AA1; Powder KE2; Kubinak, JL3, 1USCA, 2Clem, 3SOMC

STUDENT-INITIATED RESEARCH PROJECT PROGRAM (SIRP)

SIRP-01 (BI), 2:50 pm (page 12): Analyzing Cell Slide Images Using Machine Learning Techniques, William Hobbs, Yan Tong, USC

*SIRP-02 (NEU), 3 pm: The effects of SRI-32743 on the human dopamine transporter, K. Darby Porter1, Ana C. Jimenez-Torres1, Sarah E. Davis1, J. Avery Hastie1, Charles A. Adeniran2, Chang-Guo Zhan2, Jun Zhu1 , 1USC, 2U of KY

SIRP-03 (NEU), 3:10 pm (page 12-13): Connectomes in Predictive Modeling of AD Cognitive Performance Scores, Duncan Nowling, Graham Warner, Jane Joseph, MUSC

SIRP-04 (NEU), 3:15 pm (page 13): Similarities in resting-state functional brain networks in AUD and aMCI, Graham Warner, Jane Joseph, MUSC

SIRP-05 (BE), 3:25 pm (page 14): Relating Protein Biomarkers to Vascular Calcification in Femoral Endarterectomy Patients,

Breanna Pederson1, Brian Haimerl2, Shuvangee Dhar2, Daniel G. Clair3, Kathryn Fong4, John F. Eberth5, Susan Lessner1,2 , 1USC, 2SOMC, 3Vanderbilt, 4Prisma Health-Midlands, 5Drexel

POSTER PRESENTERS

BIOENGINEERING/ BIOMEDICAL ENGINEERING (BE)

BE-01 (page 15): Characterization of Sn Sputtering, Wil Armstrong, Rachel Vondergeest, Chad Rodekohr, PC

BE-02 (page 15): Soft Robotic Prosthetics Utilizing Granular Materials, Marigordon R. Varner, Eli T. Owens, PC

*BE-03: Differences in Triple Negative Breast Cancer Tumors from African American and Caucasian Patients, Z. Dinkel, M.G. Galloway, K. Joiner, E. Withers, H. Dunn, Clem

*BE-04: Protocol to Test the Effects of Chemotherapy Regimens on Skeletal Muscle Mitochondrial Function in Breast Cancer Patients Measured by Near Infrared Spectroscopy, Chloe Caudell1, Shannon Smith1, Yaseen Echekki2, Randy Hutchison2, Jennifer Trilk1 , 1SOMG, 2FU

BE-05 (page 16): Point-of-Care Colorimetric Biosensor for Detection of Pseudomonas aeruginosa Signaling Molecule 3OC12HSL in Wound Exudate from Negative Pressure Wound Therapy, Skylar Leslie, David Karig, Jordon Gilmore, Clem

BIOINFORMATICS (BI)

BI-01 (page 16): An Agent-Based Modeling Simulation of DNA SelfAssembly and Design Strategies for Dipyramidal Graphs, Ryleigh Henderson, Abbigale Outlaw, Jessica Sorrells, Conv

BI-02 (page 17): Clinical Significance of Human NPC2 Mutations, Laken Fulmer, Margo Petukh, PC

BI-03 (page 18): Effects of Dietary Iron on the Taxonomic Composition and Function of the Zebrafish Gut

Microbiome, Megan Whisonant, Jeremiah Jackson, Stuart Gordon, PC

BI-04 (page 18): Using nanopore sequencing to analyze protist diversity and identify Pseudonitzschia, Elena Renshaw1, Aramis Lawson2, Zachary Padgett2, Megan Cevasco2 , 1USC, 2CCU

CHEMISTRY/ BIOCHEMISTRY (CB)

CB-01 (page 19): Synthesis of zinc glycoconjugated phthalocyanines as a potential phototherapeutic, Katrina McCarter, Jessica Silva-Conejo, Tessa Greene, Joshua Ruppel, USC UP

CB-02 (page 19-20): The Role of SARS-CoV Non-structural protein 1 in Regulating RNA stability, Kevin Lee Xiong, Kaitlin Marie Bridges, Anita Nag, USC UP

CB-03 (page 20): Visible Light Promoted Alkylation of Imines using a Photocatalyst, Ryan A. Wernsman, Marcus I. Schlueter, James M. Hanna, Jr., WU

CB-04 (page 21): The Synthesis of Diarylpyridines as Inhibitors of Amyloid Beta Aggregation for Alzheimer’s Disease, Casey Kopyc, Mary Stegall-Smith, James M. Hanna Jr., Robin K. Lammi, WU

CB-05 (page 21-22): Characterizing azole drugs in the fungal pathogen Cryptococcus neoformans, Sakina Naqvi, Jordyn Wilson, Srikripa Chandrasekaran, FU

CB-06 (page 22): Effects of Antioxidants on Fluconazole Resistance in Cryptococcus neoformans, Neha Bhatnagar, Srikripa Chandrasekaran, FU

CB-07 (page 23): Interaction

Between the Viral RNA Leader Sequence and nsp1 in SARS Coronavirus, Kaitlin Marie Caughman, Jonathan Luke Cromer, Anita Nag, USC UP

CB-08 (page 23): Pd Catalyzed Cross-Coupling of Alkylbisboronic Esters, Ellie Kraichely, Timothy J. Barker, CofC

*CB-09: Mutation of predicted Pdr1 phosphosites restores fluconazole sensitivity in Candida glabrata, Meghan Breen, Jane McCallum, Abby Stapleton, Elizabeth Chandler, FU

CB-10 (page 24): Identification of Small Regulatory RNA UspS Associated with the Universal Stress Protein in Lactobacillus

Species, Zarah M. Fowler, Sasha S. Bronovitskiy, Finn K. Rose, Gabriela C. Pérez Alvarado, Brian M. Lee, CCU

*CB-11: Structural and functional characterization of EToV endoribonuclease nsp12, Jaci Fleming, Patrick O'Reilly, Meredith Frazier, CofC

CB-12 (page 25): Studying the Transposition Mechanism of the mJing Miniature Inverted Repeat Transposable Element in Yeast, Megan Collins, C. Nathan Hancock, USCA

CB-13 (page 25): Isolation and Analysis of Extracellular Vesicles from Lactic Acid Bacteria, Isabel Myers, Brooke Busby, Klea Hoxha, Gabriela C. Pérez Alvarado, Brian M. Lee, CCU

CB-14 (page 26): Peanut allergen reduction using phytic acid functionalized magnetic chitosan nanoparticles, Johanna Bravo, Qurian Davis, Shemyia Smith, Anselm Omoike, USC UP

CB-15 (page 26): Detecting Sequences Pseudo-nitzchia Species Using HRM Primers, Zachary Padgett1, Aramis Lawson1, Elaina Renshaw2, Megan Cevasco1 , 1CCU, 2USC

CB-16 (page 27): Identification and Analysis of the Regulatory RNA TRMS in the Probiotic Bacteria Streptococcus Thermophilus, Finn K. Rose1,2, Zarah M. Fowler2,3, Sasha S. Bronovitskiy2, Gabriela C. Pérez Alvarado2, Brian M. Lee2 , 1Scholars Academy, 2,3CCU

CB-17 (page 27): Synthesis of 1-phenylpent-4-en-2-ol, Alex Martin, Timothy Barker, CofC

CB-18 (page 28): Nucleic Acid Aptamer Au/Ag Nanoparticle Conjugates as Trojan-Horse drug Delivery Vehicles in the Fight Against Bacterial Infections, Kierra McCall, Morgan Hunter, Joshua Quarles, Ashley Wood, Allen Livingston, Victoria Frost, Timea Gerczei Fernandez, WU

MOLECULAR/CELL BIOLOGY (MCB)

MCB-01 (page 28): Histone proteomic profiling of EMTtransformed MCF10A breast cells reveal dynamics changes in epigenetic modifications, Charlotte McGuinness1, Megan A. Wilson1, Maria Ouzounova2, Hasan Korkaya3, Austin Y. Shull1 , 1PC, 2U of Lyon, France, 3MCG, GA

*MCB-02: Left ventricular function in MMP14-overexpressing mice during pregnancy and post-partum, Jessica Simpson, Frank Spinale, Holly LaVoie, SOMC

MCB-03 (page 29): Determining if the Bases Adjacent to the mPing Element Impact Transposition, M. Elizabeth Colón-LaBorde, C. Nathan Hancock, USCA

MCB-04 (page 29-30): The RhoG guanine nucleotide exchange factor SGEF controls junctional localization of desmosomal proteins, Hannah C. Lee1, Agustin Rabino2, Rafael Garcia-Mata2, Adi D. Dubash1 , 1FU, 2U of Toledo, OH

MCB-05 (page 30): The desmosomal cadherin Desmoglein-2 controls extracellular matrix gene expression via Src and NF-kB signaling, Josh M. Powers, Maya M. Mrus, Beatriz T. Mateo, Adi D. Dubash, FU

*MCB-06: Matrix Metalloproteinase 14 overexpression impacts extracellular matrix-related mRNAs in pregnant and postpartum mouse left ventricles, Ridha Fatima, Holly LaVoie, SOMC

MCB-07 (page 31): Profiling the response of individual gut microbes to common nutritional supplements used in the neonatal intensive care unit (NICU), Megan E. Waller, Caroline J. Eichhorn, Taylor D. Ticer, Janiece S. Glover, John E. Baatz, Carol L. Wagner, Katherine E. Chetta, Melinda A. Engevik, MUSC

*MCB-08: Ovarian Phenotypes and Proteomics in Mice Overexpressing a Human MMP14 Transgene, Lauren James, Holly LaVoie, SOMC

MCB-09 (page 32): IL32 expression contributes to a cell stress response and ECM-remodeling expression signature in breast cancer stem cells, Megan A. Wilson1, Elayne M. Benson1, Emma V. Gray1, Paris L. Rizzo1, Paige Cairns1, Maria Ouzounova2, Hasan Korkaya3, Austin Y. Shull1 , 1PC, 2U of Lyon, France, 3MCG

*MCB-10: Cyst reduction in vivo in a fruit fly (Drosophila) model of polycystic kidney disease by new leads, Jay DeLoriea1, Edenborough Hibionada1, Michael Monroe1, Christian Linen1,2, Eliya Karoutchy1,3, Amber Wilson1, Anh Minh Thao Nguyen4, William D. Lubell4, Chiara Gamberi1 , 1CCU, 2Acad for the Arts Sci & Tech, 3Scholars Acad, 4U de Montréal, Canada

*MCB-11: Microenvironmental cues drive macrophage polarization

program in simulated periodontal disease, Vanessa Sikora1, Waleed Alkakhan2, Nico Farrar2, Subramanya Pandruvada2 , 1BC, 2MUSC

MCB-12 (page 32): Cytotoxicity in Human Breast Cancer MCF-7 Cells following Exposure to Polycyclic Aromatic Hydrocarbons, Estrogen, and Tamoxifen, Steve Samson1, Selby Artis1, Samir Raychoudhury2 , 1Ridge View HS, 2BC

*MCB-13: Investigation of Intestinal Disease Using Caenorhabditis Elegans, MaKenna DeYoung, Summer Johnson, Mikayla Spangler, Rhiannon Follweiler, Scott Tanner, USC UP

MCB-14 (page 33): Testing HIVdependency using firefly luciferase reporter plasmids, Kaitlynn Cook, William H. Jackson, USCA

MCB-15 (page 34): Investigating the Role of RNA Polymerase V in mPing Excision Site Repair, Kaili Renken, C. Nathan Hancock, USCA

MCB-16 (page 34): Inducing Cell Death using a HIV-Dependent Expression System, Andrew P. Gregory, William H. Jackson, USCA

*MCB-17: The Developmental Biology of Bicaudal C, Edenborough Hibionada1, Jay DeLoriea1, Christian Linen1,2, Chiara Gamberi1 , 1CCU, 2Acad of Arts Science & Technology, 3Scholars Academy

*MCB-18: An investigation of the effects of miR-718 on epigenetic regulators, Cameron Honeycutt, Danielle Schmidt, Kimberly Shorter, USC UP

MCB-19 (page 35): The Effect of Cellular Differentiation on AdenoAssociated Viral Vector Genome Status, Farouk Chatila, Jennifer Lyles, FMU

*MCB-20: Probing the potential role of the gut microbiota in Polycystic Kidney Disease using the established Drosophila melanogaster BicC model, Jay DeLoriea1, Gerrit A. Stuivenberg2, Jeremy P. Burton2, Chiara Gamberi1 , 1CCU, 2Western U, London, Canada

MCB-21 (page 35): Investigations of Nucleotide Modifications in Winthrop’s Bacteriophage Collection, Gabrielle Walker, Victoria Frost, WU

*MCB-22: The Role of Calcium in Regulating Proper Chromatin Decondensation During B Lymphocyte Activation, Ruthie

Richardson, Xenia Weislamle, Jack Field, Gregory Kubista, Madison Hurst, Cassidy Rahn, Sierra McDonald, Alynna Knaub, Jason S. Rawlings, FU

MCB-23 (page 36): Generating a Gene Library of Bacteriophage Phayonce, Kyla Thomas, Matthew Defreitas, Jacqueline Gould, Daniel C. Williams, CCU

MCB-24 (page 36): The Effect of Fibronectin on Epithelial-Normal and Cancer-Cells using Electrical Impedance Sensing, Brooke Daniels, Jessica Holder, Maddy Behravan, Conv

MCB-25 (page 37): Investigating Protein Interactions of Mycobacteriophage Cain Genes 2, 9, and 55, Dallas Nivens, Laela Walker, Victoria Frost, WU

MCB-26 (page 37): Systematic Characterization of Mycobacteriophage Gene Function on Bacterial Cell Growth, Matthew Defreitas, Jackie Gould, Kyla Thomas, Daniel C. Williams, CCU

MCB-27 (page 38): Determining the Cytotoxic Effects of Phayonce Genes on Growth of Mycobacterium smegmatis, Jacqueline Gould, Matthew Defreitas, Kyla Thomas, Daniel C. Williams, CCU

MCB-28 (page 38): Characterization and Analysis of Mycobacteriophage Cain’s Genes, Laela Walker, Jessica Morgan, Victoria Frost, WU

*MCB-29: Optimization of Nucleic Acid Isolation in Eublaberus posticus, Kaitlin Brown, Neval Erturk, Christopher Varnon, Mark Bohler, Conv

MCB-30 (page 39): Effects of Estrogen, Anti-estrogen, and Polycyclic Aromatic Hydrocarbons (PAHs) on Human Breast Cancer Cells, Christian Ward, Samir Raychoudhury, BC

NEUROSCIENCE (NEU)

NEU-01 (page 39): The Analgesic Effects of Arnica montana Extracts on Post-Operative Pain, Reagan B Turner, Deltrice T Holmes, Theodore J Price, Wei Lei, PC

NEU-02 (page 40): The Impact of Taste Experience during Early Development on Later Taste and Neural Processing, Kaliana Reifsnyder, Veronica L. Flores, FU

*NEU-03: Differential Expression of the Proto-Oncogene pim-2 in the Brain Response to Injury in Fish, Zoe E. Gaskin, Shane A. Embury, David M. Hollis, FU

NEU-04 (page 41): The Impact of Innocuous Taste Experience on Long-Term Taste Learning and Memory Persistence, Dallas Shuman, Annika Patterson, Parker Pack, Veronica Lee Flores, FU

NEU-05 (page 41): The Effects of Adenosine on Temporal Perception, Kelsy White, Anna Joe Powell, Nathaly Camargo, Kelsie Glass, Richard Keen, Neval Erturk, Conv

NEU-06 (page 42): Regulation of food intake: leptin and serotonin interactions, Daniel C. Giles, Nicholas D. Maxwell, DeWitt M. Coleman, Victoria M. Glass, James R. Fadel, Claudia A. Grillo, SOMC, WJB Dorn VA Med Ctr

NEU-07 (page 42): Conformational Analysis of μ-Opioid Receptor Agonists, Leah Juechter, Brenna Outten, Lauren Jones, George Shields, FU

*NEU-08: Exploratory Investigations of Spatial Learning and Behavior in Orange Head Cockroaches (Eublaberus posticus), Amber Cox, Abeeha Sajid, Entum Knickerbocker, Christopher Varnon, Conv

NEU-09 (page 43): Blocking the Expression of PTSD in a Female Rodent Model, Alex O’Neal, Kirsten Folger, Jason Hayden, Onarae Rice, FU

PUBLIC HEALTH (PH)

PH-01 (page 43-44): Overexpression of ABC Transporters by Reflux Medication Induces Drug Resistance and Promotes Cancer Growth in Esophageal Tumor Cells, Mayah Lee, Christopher Farrell, PC

PH-02 (page 44): Modeling COVID-19 in Spartanburg County, SC with a Susceptible-VaccinatedInfected-Recovered-Deceased (SVIRD) Model, D. Chloe Griffin1 , Amanda Mangum2 , 1Brown U, 2Conv

RESEARCH EXPERIENCES FOR TEACHERS (RET)

RET-01 (page 46): GLV effects on herbivory by Tobacco Hornworms & Beet Armyworms on Tomato Plants, Lakshmi Sunitha Arava1, Harshita

Negi2, Johannes Stratmann2 , 1Fairfield Central High School, 2USC

RET-02 (page 47): Evaluation of the Aegilops tauschii chromosome introgression lines in common wheat for reduced-gluten content, Habibunnisa Begum1, Zachary T. Jones2, Sachin Rustgi2 , 1Marion HS, 2Clem

RET-03 (page 47): Exploring the Light-Induced Transformation of Photoswitchable Compound in a High School Chemistry Classroom, Asia Shaik1, Gina Wilson2, Natalia Shustova2, 1Laurens District 55 HS, 2USC

Podium Presenter Abstracts

DRP-02 (MCB) | 8:45 am ET

The Role and Regulation of RYBP in Glioblastoma Multiforme

Daniel B. Stovall, Ronald W. Bucher, Alex Lee, Lauren Patterson, Mason Linker, Brayden Fults, Daniela Torricos and Abigail Derick Department of Biology, Winthrop University, Rock Hill, SC

introduction. Glioblastoma multiforme (GBM) is an aggressive, incurable cancer of the central nervous system. GBM tumors display abnormal gene expression profiles that contribute to their malignant features. Polycomb (Pc) proteins are important regulators of gene expression that mediate normal cell differentiation and identity. Multiple Pc proteins are de-regulated in cancers and play roles in chemo- and radio-resistance in GBM, specifically. The Pc protein Ring1- and YY1-binding protein (RYBP) exerts antitumor effects in multiple cancers, but its molecular functions are context-dependent and the precise role of RYBP in GBM remains unclear.

hypothesis. We hypothesized that forced expression of RYBP in GBM cell lines would activate apoptosis while decreasing cell invasion, migration, and proliferation, and that microRNAs (miRNAs) contribute to aberrant epigenetic silencing of RYBP in GBM.

methods and results. We transduced U-118 or T98 GBM cell lines with lentivirus expressing RYBP or a GFP control and established stable cell lines. RYBPexpressing U-118 and T98 cells showed decreased migration in wound-healing assays and decreased invasion in Matrigel-coated Boyden chamber assays when compared to control cells. SDS-PAGE and Western blots were performed to measure changes in epithelial-to-mesenchymal transition (EMT) and apoptotic protein markers among transduced cells, and WST-1 assays were conducted to study the changes in proliferation. We also transiently transfected U-118 and T98 GBM cells and U-87 glioma cells with synthetic miRNA inhibitors. Inhibition of miR-9-5p and miR-128-3p led to a marked increase in RYBP protein levels across cell types.

conclusion. Overall, our findings suggest RYBP exerts anti-tumor effects in GBM and that miRNAs contribute to RYBP silencing in GBM. Further delineation of the pathways leading to RYBP silencing and the networks activated by restored RYBP levels may illuminate cellular programs that can be therapeutically exploited in GBM treatment.

citation/acknowledgements. This work was funded by SC INBRE DRP 5P20GM103499-21.

Meet DRP Recipient Daniel Stovall (FY2021-2023)

Institution/

Department: Winthrop University, Biology

Project Title: Role and Regulation of Ring1 and YY1 binding protein in Glioblastoma Multiforme

Outside the Lab/ Classroom:

When I am not in the lab or classroom, I enjoy spending my time reading, traveling, and sketching.

How SC INBRE funding will help accomplish project goals: Funding from SC INBRE has allowed us to involve so many undergraduate biology majors at Winthrop University in biomedical research! Funds are also used to purchase the necessary supplies and reagents needed, without which this work would not be possible.

Learn more about Dr. Stovall at https://bit.ly/ scinbre-spotlights

Modeling macrophage responses to periodontal infections in vitro

Nico Farrar, Waleed Al-Kakhan, Vanessa Sikora and Subramanya Pandruvada Department of Oral Health Sciences, College of Dental Medicine, Medical University of South Carolina, Charleston, SC

background. Periodontal disease (PD) is a chronic inflammatory disorder characterized by the destruction of connective tissue, tooth loss, and systemic infections. Numerous factors derived from opportunistic oral pathogens are responsible for suppressing host defense and enhanced microbial colonization. Individuals with PD present a complex array of inflammatory cues and cellular infiltrates. While macrophages comprise 5−30% infiltrate of periodontal lesions, how macrophage skewing contributes to PD is unclear. We have shown previously that Porphyromonas gingivalis infection induces M1-macrophage formation, whereas opsonized-P. gingivalis challenge was able to switch to an M2-like phenotype demonstrating macrophage plasticity. However, the modulation mechanism during the process is unclear. Since macrophage polarization dynamics are hard to demonstrate in vivo, a reliable in vitro system is needed to understand them in PD.

goal of study. Our research aims to understand how macrophage skewing contributes to periodontal disease.

methods and results. Peripheral blood mononuclear cells (PBMCs) and leukemic cell line THP-1 cells were used to generate macrophages. Briefly, THP-1 cells were primed with phorbol 12-myristate (PMA) and cocultured with human primary gingival fibroblasts (GFs) and sequentially exposed to various factors at different times: CCL2, LPS, IFN-g, TNF-a, IL-4, IL-10, and TGF-b. Similarly, PBMC-derived monocytes were also exposed to the same inflammatory program without PMA activation. Samples were collected at predetermined points for RNA profiling using NanoString. Subsequently, candidate genes were validated using RT-PCR and chemokine arrays at different stages of polarization. Our novel gingival fibroblast-macrophage co-culture model successfully portrayed the transition from M0-to-M1-to-M2. In addition to classic macrophage markers, gene expression analysis confirmed differential expression of specific M1 (ICOS) and M2 genes (IL9). Chemokine profiling displayed the differential protein expression (M1: CXCL7; M2: CCL17) corresponding to macrophage benchmarks validating the co-culture model.

conclusions. We have developed a new model that summarizes macrophage response when exposed to a battery of cues mimicking macrophage polarization/depolarization observed in vivo citation/acknowledgements. This work was supported by SC INBRE, University of South Carolina/ NIGMS 2P20GM103499-20 (Developmental Research Project Program).

DRP-04

(BE) | 9:15 am ET

Point-of-Care Colorimetric Biosensor for Detection of Pseudomonas aeruginosa Signaling Molecule 3OC12HSL in Wound Exudate from Negative Pressure Wound Therapy

Skylar Leslie, David Karig, and Jordon Gilmore Department of Bioengineering, Clemson University, Clemson, SC introduction. Pseudomonas aeruginosa is an opportunistic pathogen, recently highlighted by the World Health Organization (WHO) as one of the infamous ESKAPE pathogens, known for their ability to evade common antibiotic treatments through various forms of resistance (1-3). Pseudomonas prevalence (32,000 infections in hospitalized patients, 2,700 deaths in 2017 (4)) presents a severe threat, particularly to patients with compromised wound healing (i.e., diabetics) (1). Contributing to resistance, P. aeruginosa coordinates biofilm formation through quorum sensing (QS), in which diffusible molecules are secreted and sensed as a means of detecting local population density (2). This work focuses on the detection of P. aeruginosa QS signaling molecules to inform

infection diagnosis and treatment. Specifically, we detect N-(3-Oxydodecanoyl)-L-homoserine lactone (3OC12HSL) from the Las QS pathway, which coordinates virulence. 3OC12HSL in part activates Las quorum sensing-controlled promoters, that in turn activate other P. aeruginosa QS pathways such as the Rhl, IQS, and PQS pathways.

goal of study. We employ, in a nonpathogenic Escherichia coli host, a synthetic plasmid that codes for expression of LasR from a constitutive promoter and expression of a colorimetric (blue) reporter, amilCP, from a quorum sensing-controlled promoter.

methods and results. DH5a E. coli were transformed and then cultured in LB medium with kanamycin. Conical tubes (n=12) were prepared as follows: 3mL with 10uM of exogeneous 3OC12HSL added to tube 1; 2mL with 1 mL serially diluted aliquots added to tubes 2-12. Tubes were plated in 24-well plates, placed in a 37°C plate reader, and read at 588nm, 600nm, 700nm, and GFP for 24 hours. In the presence of 3OC12HSL, DH5a+amilCP E. coli cells turn blue, providing a simple visual readout proportional to the target concentration, without additional equipment required.

conclusions. This biosensor successfully detects 3OC12HSL (LOD approximately 0.0565nM), producing visible blue color within 24 hours. Next steps include incorporation of the biosensor into negative pressure wound therapy devices.

References:

1. 2019. Pseudomonas aeruginosa Infection | HAI | CDC. https://www.cdc.gov/hai/ organisms/pseudomonas.html. Retrieved 30 November 2022.

2. Thi MTT, Wibowo D, Rehm BHA. 2020. Pseudomonas aeruginosa Biofilms. Int J Mol Sci 21:8671.

3. Talebi Bezmin Abadi, A., Rizvanov, A.A., Haertlé, T. et al. World Health Organization Report: Current Crisis of Antibiotic Resistance. BioNanoSci. 9, 778–788 (2019). https://doi. org/10.1007/s12668-019-00658-4

4. CDC. 2022. The biggest antibiotic-resistant threats in the U.S. Centers for Disease Control and Prevention. https://www.cdc.gov/ drugresistance/biggest-threats.html. Retrieved 30 November 2022.

citation/acknowledgements.

This work was supported by SC INBRE grant #P20GM103499-20 (Developmental Research Project Program) awarded to JG.

Linking trained immunity with tumor dormancy by KLF4-mediated cell fusion

Arianna Bastian1, Joshua Garbutt1, Mitzi Nagarkatti2, Daping Fan3, Walden Ai1,3

1Department

of Biology, Chemistry and Environmental Health

Science, Benedict College, Columbia, SC, 2Department of Pathology, Microbiology and Immunology, University of South Carolina School of Medicine, Columbia, SC, 3Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC

introduction/background. Immune memory classically occurs in the adaptive arm of our immune system. However, recent studies suggest a key role of memory of the innate immune system in inflammation-linked processes. Altered secondary response in the innate system is called trained immunity. It has protective effects on cancer. However, short-lived innate cells may not explain their long-lasting effect of their anti-tumor activities. On the other hand, longlived tissue stem cells can acquire memory. Whether and how cancer stem cells acquire similar memory in the early stage of cancer leading to later protective effects such as tumor dormancy is not known.

hypothesis/goal of study. We hypothesize that monocyte expressing KLF4 controls the cell fusion between breast cancer cells and monocytes. The goal of this study is to examine whether cell fusion between breast cancer cells and monocytes is responsible for the late stage of breast cancer dormancy and recurrence.

methods and results. We cultured different breast cancer cells (MCF-7, MDA-MB-231, and MDA-MB-468 cells) with different monocytes (THP-1, U-937, and primary monocytes) and examined cell fusion by flow cytometry. Gene expression was tested by Western blotting analysis and qRT-PCR. We found that only triple negative breast cancer cells including MDA-MB-231 and MDA-MB-468 cells but not estrogen receptor positive MCF-7 cells fused with monocytes. In this process, gene expression of KLF4 as well as markers of dormant cancer stem cells was elevated. In addition, we have generated KLF4-deficient THP-1 and U-937 cells using CRISPR-CAS9 system and will test cell fusion of these cells with MDA-MB-231 cells. Gene profiling will be performed using control and KLF4-deficient monocytes and fusion cells.

conclusions. We expect to establish that KLF4 in monocytes determines cell fusion between triple negative breast cancer cells with monocytes leading to the generation of dormant cancer stem cells in breast cancer.

citation/acknowledgements. This work was supported by NIH SC INBRE (Bioinformatics Pilot Project Program) and SC3GM136639 (NIGMS) awards to WA.

Analyzing Cell Slide Images Using Machine Learning Techniques

William Hobbs and Yan Tong Department of Computer Science and Engineering, University of South Carolina, Columbia, SC

Lung sputum slides can contain many different types of cells, but two common ones are macrophages and neutrophiles. The University of New Mexico studies lung sputum as an indicator of population health due to smoke, whether from nicotine addiction or contaminated air in certain areas. Prolonged exposure to contaminated air particles has been seen to result in black carbon particles in macrophages. Macrophages, which engulf and capture black carbon particles, can be studied to give epidemiologists an idea of population air health, when collected in a lab. However, the manual segmentation and measurement of these images is time consuming and intensive. An experimental pipeline was built that (through manual inputs) can take in a batch of cell images, augment them, and train a network to use for semantic segmentation via Matlab and Python scripts. The semantic segmentation implementation uses a neural network to “cut” out the cell images present in the data set but does not do this to an acceptable degree of accuracy. Python scripts and Matlab scripts, using public tutorials for Python, Matlab, OpenCV, and the Python Image Library, were built to expand the dataset six-fold.

citation/ acknowledgements. This work funded by grant # 10011476.

Connectomes in Predictive Modeling of AD Cognitive Performance Scores

Duncan Nowling, Graham Warner, Jane Joseph Department of Neuroscience, Medical University of South Carolina, Charleston, SC

introduction/background. Alzheimer’s Disease (AD) has no cure, but earlier diagnosis may improve treatment. Subjective Cognitive Decline (SCD) is a possible preclinical stage before Mild Cognitive Impairment (MCI), AD’s prodromal stage. SCD’s pathophysiology remains poorly defined. Graph theory (GT) measures can further understanding by showing how communication changes across the whole network. This study proposed to create GT-based models predictive of cognitive scores across healthy and MCI subject groups that show an over or under prediction in SCD.

hypothesis/goal of study. Our primary hypothesis is that connectome based predictive models of cognitive scores identified using healthy control and MCI connectome and cognitive performance data will remain accurate when transferred to held out data, but over or underpredict SCD scores.

methods and results. GT measures of betweenness centrality (BTW), eigenvector centrality (EVC), and clustering coefficient (CC) were calculated from the resting state fMRI scans of 122 healthy, 101 MCI, and 35 SCD participants. We built models using each of the measures as predictors of cognitive scores for Montreal Cognitive Assessment (MoCA), Logic Delayed (LogDel), Trails A (TraA), and Trails B (TraB) in 85 healthy and 71 MCI scans. These models were transferred to 35 SCD and 30% held out healthy and MCI datasets. Models remained accurate post-transfer, with no model’s prediction gap being significantly different from 0. No differences were found in predicted scores or gap scores between healthy and SCD data.

conclusion. Our findings indicate SCD is likely part of the AD continuum. However, rather than being over or underpredicted based on the HC/MCI continuum, SCD predicted and gap scores did not significantly differ from healthy data. This indicates that at the whole network level, connectome organization does not differ between HC and SCD. Future work should focus on narrowing predictors to specific nodes of interest, as differences between healthy and SCD may be found at smaller regional levels, while simplifying model interpretation.

citation/acknowledgements. This work was supported by funding through the SC INBRE Student Initiated Research Project (SIRP) award and the NIH/NIA grant R01

AG55132. Some data used in the preparation of this study were obtained from the Alzheimer’s Disease Neuroimaging Initiative (ADNI) database (adni.loni.usc.edu). The ADNI was launched in 2003 as a public-private partnership, led by Principal Investigator Michael W. Weiner, MD. The primary goal of ADNI has been to test whether serial magnetic resonance imaging (MRI), positron emission tomography (PET), other biological markers, and clinical and neuropsychological assessment can be combined to measure the progression of mild cognitive impairment (MCI) and early Alzheimer’s disease (AD). For up-todate information, see www.adni-info.org.

The Alzheimer’s Disease Neuroimaging Initiative (ADNI) is a longitudinal multicenter study designed to develop clinical, imaging, genetic, and biochemical biomarkers for the early detection and tracking of Alzheimer’s disease (AD). Since its launch more than a decade ago, the landmark public-private partnership has made major contributions to AD research, enabling the sharing of data between researchers around the world.

ADNI unites researchers with study data as they work to define the progression of AD. ADNI researchers collect, validate and utilize data, including MRI and PET images, genetics, cognitive tests, CSF and blood biomarkers as predictors of the disease. Study resources and data from the North American ADNI study are available through their website, including Alzheimer’s disease patients, mild cognitive impairment subjects, and elderly controls.

Use QR code to learn more about ADNI.

Similarities in

resting-state functional brain networks in AUD and aMCI

Graham Warner and Jane Joseph Department of Neuroscience, Medical University of South Carolina, Charleston, SC

introduction/background. Alcohol use disorder (AUD) and both Alzheimer’s Disease (AD) and its prodromal stage, amnestic mild cognitive impairment (aMCI) are known to cause alteration in resting-state functional brain networks. AUD is also known to be a risk factor for AD though the mechanism by which AUD increases the risk of AD remains unknown. It is possible that AUD causes similar brain network dysfunctions to AD and aMCI and that these dysfunctions may be what causes AUD to increase the risk of AD.

goal of study. The goal of this study is to determine whether AUD and aMCI show similar alterations in resting state brain networks, compared to canonical networks that emerge in healthy control (HC) subjects.

methods and results. Resting-state fMRI data from 110 aMCI, 22 AUD, and 84 HC subjects were used for this analysis. FSL’s MELODIC tool was used to temporally concatenate and decompose each group’s resting-state fMRI data independently. The resulting independent components for each group were then manually labeled as part of the default mode network, episodic memory encoding network, fronto-parietal network, or not part of any of the three networks. The components matching each network for each group were then spatially merged resulting in three network masks for each of the three groups. Spatial crosscorrelations were performed between corresponding network maps in each of the three groups.

conclusions. The HC and AUD populations showed the highest correlations in each of the three networks while the HC and amnestic aMCI groups consistently showed the lowest correlations. These results indicate that aMCI involves greater network alteration than AUD. The relatively low correlations between AUD and aMCI networks suggest that the two disorders are associated with dissimilar alterations in functional brain networks.

citatation/acknowledgements. This work funded by SC INBRE grant # 5P20GM103499-21.

Relating Protein Biomarkers to Vascular Calcification in Femoral Endarterectomy Patients

Breanna Pederson1,

Brian

Haimerl2,

Shuvangee Dhar2, Daniel G. Clair3, Kathryn Fong4, John F.

Eberth5,

Susan Lessner1,2 1University of South Carolina, Biomedical Engineering Program, Columbia, SC, 2University of South Carolina School of Medicine, Columbia, SC, 3Vanderbilt University Medical Center, Department of Vascular Surgery, Nashville, TN, 4Department of Surgery, Prisma Health-Midlands, Columbia, SC, 5Drexel University, School of Biomedical Engineering, Science and Health Systems, Philadelphia, PA

introduction. Peripheral arterial disease (PAD), one of the leading factors for nontraumatic limb amputation1, involves a pathological narrowing of arteries supplying blood to the extremities. Symptoms range from intermittent claudication to chronic limb-threatening ischemia. Patients experience reductions in ambulatory ability, functional capacity, and quality of life1. Femoral endarterectomy (FEA), a common surgical intervention for PAD, involves removing a stenotic plaque from the femoral artery3.

goal of study. To correlate plasma biomarker proteins with vascular calcification in PAD patients undergoing FEA.

methods and results. Six target proteins — myostatin, osteoprotegerin(OPG), osteocalcin, follistatin-related gene(FLRG), fetuin A, and thrombospondin-1(TSP-1) — were quantified using enzyme-linked immunosorbent assays in plasma obtained from 29 FEA patients and 6 male controls. Lower extremity calcification was quantified in vivo using non-contrast CT scans. Principal component analysis was used to analyze the relationship between biomarkers and calcification volume5. Average plasma concentrations of FLRG and osteocalcin were lower in controls than in FEA patients (5.1±2.0 vs.12.2±8.9 ng/mL and 1.4±1.4 vs. 14.8±18.1 ng/ mL, respectively; p<0.001). Myostatin and fetuin A were higher in controls than in patients (4.5±0.9 vs. 3.1±1.8 mg/mL and 1.0±0.3 vs. 0.4 ±0.2 mg/ mL, respectively; p<0.05). OPG was lower in controls than patients (38.1±4.1 vs. 48.4±8.2 pg/mL; p<0.001). TSP-1 was not significantly different between groups. Fetuin A and OPG were significantly correlated with calcification volume by linear regression. The first principal component was positively correlated to increased osteoprotegerin and FLRG and decreased myostatin and TSP-1.

conclusions. OPG, FLRG, and osteocalcin were significantly elevated in patients while fetuin A and myostatin were significantly reduced. Fetuin A and OPG were significantly correlated with calcification. This biomarker data has the potential for improved diagnosis and risk stratification of PAD patients via simple bloodwork analysis. In future, patient demographics and clinical data will be included to determine which variables best predict outcomes. acknowledgements. We would like to thank our collaborators at Prisma Health Midlands and our funding from SC INBRE Student-Initiated Research Projects Program 10011728 and NIH R01 HL145064. Literature Cited: [1] P.P. Goodney et al. J Vasc Surg. 2009; 57:1471-1480.e3. [2] N.C. Dolar et al. Jama. 2001; 286:1599-1606. [3] M. Christopher. Physiol Behav. 2016; 176:100–106. [4] S.M. Vartanian et al. Circ Res. 2015; 116:1614-1628. [5] I.T. Jollife et al. Philos Trans R Soc A Math Phys Eng Sci. 2016; 374.

Meet SIRP

Recipient Brenna Pederson (FY2022-2023)

Institution/Department:

USC School of Medicine

Columbia, Cell Biology & Anatomy (graduate student)

Project Title: Predicting Outcomes of Femoral Endarterectomies from Protein levels

Project Information:

Peripheral Arterial Disease is a common disease associated with aging in which the blood flow to the lower limbs is reduced, often affecting mobility. A common treatment is a femoral endarterectomy surgery, in which a plaque is removed from the femoral artery to help increase blood flow. In this study we investigated the concentration of six proteins in the blood of patients undergoing this procedure and compared them to healthy individuals. We also quantified the amount of calcium in the patient’s arteries as well as in the plaque which was removed. We are using this information to try to predict the outcomes of PAD in patients as well as to identify the relationship of the proteins to calcium amount.

Outside the Lab/Classroom: Outside of the classroom I really enjoy traveling, especially to historical places, and highropes courses, which are like obstacle courses elevated off the ground.

Learn more about Brenna at https://bit.ly/ scinbre-spotlights

Characterization of Sn Sputtering

Wil Armstrong, Rachel Vondergeest, Chad Rodekohr Presbyterian College, Clinton, SC

In the realm of modern electronics, spontaneous Sn Whisker propagation is a growing concern. The purpose of this research was to characterize a sputter deposition system and study the effects of background Ar pressure on the stress within thin Sn films created by this system. Several modifications have been made to this sputter deposition system in the pursuit of developing a system capable of reaching the necessary voltage for sputtering Sn at low pressures. With these modifications, Sn sputtering has become a reality, but an effective method of maintaining plasma in a high vacuum is difficult at best. To better understand the abilities of this new sputter system and the necessary balance of pressure and voltage, the system was tested at various pressures to find the necessary starting voltage to produce plasma. Using the SEM, we hope to study the growth of Sn whiskers produced by the induced stress within the thin Sn films. The goal is to understand Sn whiskers in order to mitigate device failure caused by whisker growth in thin films and possibly use them in innovative technologies such as MEMS (Microelectromechanical systems). citation/ acknowledgements. This work was funded by institutional funds from Presbyterian College and SC INBRE.

Soft Robotic Prosthetics Utilizing Granular Materials

Marigordon R. Varner and Eli T. Owens Presbyterian College, Clinton, SC

introduction. As technological advancements continue to be incorporated into prosthetics, there is a gap that is created by the quality/cost ratio associated with using technologically advanced prosthetics. Many open-sourced prosthetics are relatively inexpensive prosthetics but sacrifice the ability to grip unique shapes. Prior work in the field has successfully used compressed air and a small granular material to build a gripper that can hold various objects. The use of compressed air makes these devices relatively slow and limits the applicability of this design.

hypothesis. Our goal is to explore the use of magnetic granular materials and an external magnetic field to induce the jamming of the material. The granular material is inside flexible membranes wherein it can go from a more relaxed state to the more rigid state depending on the state of the magnetic field.

methods and results. The flexible membrane also allows the object to be held while the solenoid is on, and then released when the solenoid is off. Iron filings work well as our granular material as they use solenoids to conform to the unique shape.

conclusions. This, when combined with the myoelectric effect, the electrical signals sent through the muscles, allows the magnetic field to be turned off and on. This design furthers our pursuit of creating a low-cost but high functioning upper prosthetic.

citation/acknowledgements. This project was supported by grant P20GM103499-20 (SC INBRE).

“Granular materials are unique because they can act like any of the three states of matter,” Marigordon said. “For example, sand can act like a solid when we walk on the beach, a liquid when it flows within an hourglass, and a gas as it is blown during a sandstorm.”

Specifically, Marigordon and Dr. Owens are studying the jamming transition using sound to measure the impedance of granular material and characterize how far the granular material is from the jamming transition. Read more at https://bit.ly/scisymp23-varner

Marigordon Varner and physics professor Dr. Eli Owens, Presbyterian College, are researching granular materials like sand, grains and soil.

Point-of-Care Colorimetric Biosensor for Detection of Pseudomonas aeruginosa Signaling Molecule 3OC12HSL in Wound Exudate from Negative Pressure Wound Therapy

Skylar Leslie, David Karig, and Jordon Gilmore Department of Bioengineering, Clemson University, Clemson, SC introduction. Pseudomonas aeruginosa is an opportunistic pathogen commonly found across various environments and hosts (1, 2). P. aeruginosa prevalence (32,000 infections in hospitalized patients, 2,700 deaths in 2017 (3)) presents a severe threat with multiple drug resistances (MDR) (1). Contributing to MDR, P. aeruginosa coordinates biofilm formation through quorum sensing (QS), in which diffusible molecules are secreted and sensed as a means of detecting local population density (2). This work focuses on the detection of P. aeruginosa QS signaling molecules to inform infection diagnosis and treatment. Specifically, we detect N-(3Oxydodecanoyl)-L-homoserine lactone (3OC12HSL) from the Las QS pathway, which coordinates virulence. 3OC12HSL binds the protein LasR, forming a complex that activates Las QS-controlled promoters. We employ a synthetic plasmid that codes for expression of LasR from a constitutive promoter and expression of a colorimetric (blue) reporter, amilCP, from a QS-controlled promoter. The biosensor host is n onpathogenic Escherichia coli (E. coli) offering a sensitive, specific, and low-cost detection method with visual readouts interpretable without additional equipment.

hypothesis. The proposed biosensor will facilitate visible detection and relative quantification of 3OC12HSL in culture within 24 hours.

methods and results. DH5a E. coli were transformed and cultured in LB medium with kanamycin. Conical tubes (n=12) were prepared as follows: 3mL with 10uM of exogeneous 3OC12HSL added to tube 1; 2mL with 1 mL serially diluted aliquots added to tubes 2-12. Tubes were plated in 24-well plates, placed in a 37°C plate reader, and read at 588nm, 600nm, and 700nm for 24 hours. In the presence of 3OC12HSL, DH5a+amilCP E. coli cells turn blue, providing a simple visual readout proportional to the target concentration, without additional equipment.

conclusions. This biosensor successfully detects 3OC12HSL (LOD approximately 0.0565nM), producing visible blue color within 24 hours. Next steps include incorporation of the biosensor into negative pressure wound therapy devices.

citation/acknowledgements. This work funded by an SC INBRE DRP grant awarded to J.G.

References: 1. 2019. Pseudomonas aeruginosa Infection | HAI | CDC. https://www.cdc.gov/hai/organisms/pseudomonas.html. Retrieved 30 November 2022. 2. Thi MTT, Wibowo D, Rehm BHA. 2020. Pseudomonas aeruginosa Biofilms. Int J Mol Sci 21:8671. 3. CDC. 2022. The biggest antibiotic-resistant threats in the U.S. Centers for Disease Control and Prevention. https://www.cdc.gov/drugresistance/biggest-threats.html. Retrieved 30 November 2022.

An AgentBased Modeling Simulation of DNA SelfAssembly and Design Strategies

for Dipyramidal Graphs

Ryleigh Henderson, Abbigale Outlaw, and Jessica Sorrells Department of Mathematics, Converse University, Spartanburg, SC

introduction. DNA selfassembly is a type of molecular self-assembly. DNA selfassembly is a rapidly advancing field as DNA nanostructures created via self-assembly are useful in diagnosing and treating various illnesses, biomolecular computing, drug delivery, and biosensors. Due to the utility of DNA self-assembly, laboratories seek efficient methods to construct DNA complexes. In this case, efficiency translates into using the minimum number of different component molecules. To determine the minimum number, discrete graphs are used to model the DNA nanostructures and the self-assembly process. This work investigates the problem for the dipyramidal graph family within the framework of the flexible-tile self-assembly model. Dipyramidal graphs can be visualized as the skeletons of polyhedra which consist of two pyramids conjoined at the base. goal

of study.

This work aimed to determine, under two different sets of constraints, the minimum number of tile and bond-edge types (i.e. component molecule types) necessary to

construct dipyramidal graphs. This work also aimed to produce a program to model the self-assembly process in the agent-based modeling environment NetLogo. NetLogo, a programming language and integrated development environment (IDE), creates a visual output that is easier for diverse audiences to understand.

methods and results. Within the flexible-tile graph theoretical model we employ various mathematical techniques, such as valency sequences of graphs, linear algebra, and graph isomorphism. We determine that the minimum numbers of tiles and bond-edge types for dipyramidal graphs in the least restrictive scenario are two and one, respectively. Minimum numbers of tile and bondedge types for odd sizes of dipyramidal graphs in the more restrictive scenario are also two and one, respectively.

conclusions. Using a graph theoretical model, minimum numbers of theoretical component molecule types were determined for dipyramidal DNA nanostructures. A program for a simple agent-based model of DNA self-assembly was programmed in NetLogo.

citation/ acknowledgements.

This project is supported by a grant from the South Carolina INBRE funded by the National Institutes of Health National Institute of General Medical Sciences (P20GM103499).

Clinical Significance of Human

NPC2 Mutations

Laken Fulmer and Margo Petukh Biology Department, Presbyterian College

introduction/background. Cholesterol (CLR), vital to eukaryotic cell membranes, rigidifies fluid membranes thus reducing passive permeability and increasing mechanical durability of lipid bilayers. CLR enables post-Golgi protein sorting by regulating lipid bilayer thickness. Synthesis, influx, and efflux of CLR levels are regulated by specialized cellular systems. NPC intracellular cholesterol transporter 2 (NPC2) is a soluble 151 aa protein in late endosomes/lysosomes that binds unesterified CLR and transfers it to NPC1 for transport to other cellular compartments. Some NPC2 mutations cause Niemann-Pick disease type C (NPC)- a heritable disorder characterized by accumulation of lipids in endosomes and lysosomes with delayed induction of CLR homeostatic reactions. NPC symptoms include: ataxia, vertical supranuclear gaze palsy, dystonia, liver disease, and interstitial lung disease. Affected individuals often experience intellectual decline, and approximately one-third experience seizures. NPC symptoms frequently manifest during childhood, but they can begin at any stage including adulthood. Out of 152 documented missense mutations in NPC2, 23 have uncertain pathogenicity.

hypothesis/goal of study. This study evaluates missense mutations of NPC2 with unknown clinical significance to predict their pathogenicity concerning NPC. Residues crucial to protein function show high evolutionary conservation, so we expect evolutionary analysis, structural analysis, and data mining to provide adequate data about each residue’s involvement in functionality to distinguish between benign and pathogenic mutations.

methods and results. Data mining provided demographic information, providing insight into how mutations affect specific populations. It also addressed other questions related to NPC2 function, cellular location, optimum environmental conditions, binding partners, 3D structure availability, and residues critical for NPC2 structure/function. Evolutionary analysis allowed evaluation of residue conservation. NPC2 residues affecting protein stability, “hot-spots'', were evaluated with alanine-scanning.

conclusions. We propose nine (A19D, E20K, S24W, V30M, V57I, K71R, D91N, N98H, T170I) likely pathogenic mutations commonly detected at advanced life stages for further analysis.

citation/acknowledgements. This project is supported by a grant from SC INBRE (P20GM103499).

Effects of Dietary Iron on the Taxonomic Composition and Function of the Zebrafish

Gut Microbiome

Megan Whisonant, Jeremiah Jackson and Stuart Gordon

Department

of Biology, Presbyterian College, Clinton, SC

Microbial communities have a myriad of effects on their hosts, which can be exemplified by dietary changes and their effects on the human gut. Iron, an essential component of heme and iron-sulfur proteins, plays a central role in many biological activities, including oxygen transport and cellular respiration. In particular, the iron homeostasis system is one of the best characterized due to iron's causative relationship with iron-deficiency anemia — of which, diagnosed individuals harbor pressure to elucidate the findings on this topic of research. In the present study, using Zebrafish (Danio rerio) as a model organism, we sought to confirm that increases in dietary iron would cause changes in taxonomic composition and alter the metabolic function of the gut microbiome. The findings indicated that dietary iron appeared to significantly alter the taxonomic composition with notable increases in the phyla Firmicutes and Proteobacteria. Functional potential analysis suggests that iron enriched physiological functions related to aerobic respiration. These results will be further explored through metabolomic analysis of primary metabolites and lipids.

citation/acknowledgements.

This project is supported by SC INBRE grant (P20GM103499).

Elena Renshaw1, Aramis Lawson2, Zachary Padgett2, and Megan Cevasco2

1Department of Biological Sciences, University of South Carolina, Columbia, SC, 2Department of Biology, Coastal Carolina University, Conway, SC

introduction/background. Harmful algal blooms (HABs) pose a risk to human and environmental health, especially in coastal areas. A particular taxon of interest when considering shellfish harvesting grounds is Pseudo-nitzschia, as some species produce domoic acid, a neurotoxin that can bioaccumulate in shellfish and cause amnesiac shellfish poisoning in humans.

hypothesis/goal of study. This study aimed to explore the use of ribosomal primers of varying specificity and nanopore sequencing as an assay of ecological diversity. It was hypothesized that species-specific and protist-specific primers would detect Psuedo-nitzschia DNA, and the broader Eukaryotic primer sets would be less likely to pick up the sequence if it was present.

methods and results. Water samples were collected from Oyster Landing in Huntington Beach State Park on five different days in June-July 2022. DNA was extracted from the filtered samples and then amplified, cleaned, and quantified. The samples were prepared using Oxford Nanopore’s protocol for ligation, cleaning, and library preparation. The MinION nanopore sequencer was used on the prepared DNA library, and results were analyzed using NCBI’s Nucleotide Blast. Five primer sets of varying specificity were used: two Psuedo-nitzschia, one diatom, and two Eukaryotic. All the primer sets detected Pseudo-nitzschia except for the diatom-specific primers. The diatom-specific primers detected the largest variety of diatoms with the least offtarget amplification but did not pick up Psuedo-nitzschia. Both Eukaryotic primer sets only detected one species of Pseudonitzschia, P. delicatissima, which is a domoic acid producer.

conclusions. The Pseudo-nitzschia specific primers were the most effective in detecting several species of the diatom, while the broader primers provided a more comprehensive analysis of ecological diversity. This could inform future research on the best method of detecting the presence of Pseudo-nitzschia and other HAB taxa in shellfish harvesting waters, which could be used to mitigate health impacts of HABs.

citation/acknowledgements. This project was supported by grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health.

Using nanopore sequencing to analyze protist diversity and identify Pseudo-nitzschia

Synthesis of zinc glycoconjugated phthalocyanines as a potential phototherapeutic

Katrina McCarter, Jessica Silva-Conejo, Tessa Greene, Joshua V. Ruppel University of South Carolina Upstate, Spartanburg, SC

introduction. Photodynamic Therapy (PDT) uses photosensitizers that are activated by light to produce singlet oxygen that can cause cell damage and potential cell death. Phthalocyanines are promising photosensitizers due to their absorption at clinically relevant wavelengths, meaning that they absorb at a wavelength that can penetrate deeper into tissues. We will present our efforts towards the synthesis of a zinc glycoconjugated phthalocyanine photosensitizer displaying different symmetries such as the so called A4 and A3B phthalocyanines. goal of study. One challenge with using Phthalocyanines is that they have very low solubility in biological media. One potential approach to increase solubility is through glycoconjugation. Glycoconjugation can also be used as a strategy to increase selectivity toward targeted tissues. Glycoconjugated phthalocyanines can be synthesized using a copper catalyzed azide-alkyne cycloaddition reaction, otherwise known as a “click reaction.” Moreover, zinc glycoconjugated phthalocyanines are of interest due to their efficiency in generating singlet oxygen.

methods and results. Through a multiple step synthesis, phthalocyanines can be made through phthalonitrile derivatives. Alkynylated phthalonitrile derivatives can be made readily produced from nitro phthalonitrile through a substitution reaction. These alkynylated derivates can undergo a ‘click reaction’ in the presence of azido carbohydrates to create glycoconjugated derivates. These glycoconjugated phthalonitrile derivates can be subjected to a base promoted cyclization process to create the desired glycoconjugated phthalocyanine products. We have shown that this synthetic method can produce the glycoconjugated phthalocyanines, although challenges in purification and subsequent characterization persist.

conclusion. While the synthesis of the phthalocyanine glycoconjugates has been completed and NMR evidence indicates their presence, purification of the final product and complete characterization remains a challenge. Ongoing efforts are directed to the development of a reliable and reproducible purification protocol for the desired zinc glycoconjugated phthalocyanines.

citation/acknowledgements. This work was supported by funding through SC INBRE/NIH (P20GM103499-20) and an NIH R15 award (1R15GM148916-01).

The Role of SARS-CoV Non-structural protein 1 in Regulating RNA stability

Kevin Lee Xiong, Kaitlin Marie Bridges, Anita Nag Department of Natural Sciences and Engineering, USC Upstate, Spartanburg, SC

introduction. Photodynamic Therapy (PDT) uses photosensitizers that are activated by light to produce singlet oxygen that can cause cell damage and potential cell death. Phthalocyanines are promising photosensitizers due to their absorption at clinically relevant wavelengths, meaning that they absorb at a wavelength that can penetrate deeper into tissues. We will present our efforts towards the synthesis of a zinc glycoconjugated phthalocyanine photosensitizer displaying different symmetries such as the so called A4 and A3B phthalocyanines.

goal of study. One challenge with using Phthalocyanines is that they have very low solubility in biological media. One potential approach to increase solubility is through glycoconjugation. Glycoconjugation can also be used as a strategy to increase selectivity toward targeted tissues. Glycoconjugated phthalocyanines can be synthesized using a copper catalyzed azide-alkyne cycloaddition reaction, otherwise known as a “click reaction.” Moreover, zinc glycoconjugated phthalocyanines are of interest due to their efficiency in generating singlet oxygen.

NAG FY22 (3RD YR) DRP RECIPIENT | CB-02

methods and results.

Through a multiple step synthesis, phthalocyanines can be made through phthalonitrile derivatives. Alkynylated phthalonitrile derivatives can be made readily produced from nitro phthalonitrile through a substitution reaction. These alkynylated derivates can undergo a ‘click reaction’ in the presence of azido carbohydrates to create glycoconjugated derivates. These glycoconjugated phthalonitrile derivates can be subjected to a base promoted cyclization process to create the desired glycoconjugated phthalocyanine products. We have shown that this synthetic method can produce the glycoconjugated phthalocyanines, although challenges in purification and subsequent characterization persist.

conclusions. While the synthesis of the phthalocyanine glycoconjugates has been completed and NMR evidence indicates their presence, purification of the final product and complete characterization remains a challenge. Ongoing efforts are directed to the development of a reliable and reproducible purification protocol for the desired zinc glycoconjugated phthalocyanines.

citation/ acknowledgements.

This work was supported by funding through SC INBRE/NIH (P20GM103499-20) and an NIH R15 award (1R15GM148916-01).

Visible Light Promoted Alkylation of Imines using a Photocatalyst

Ryan A. Wernsman, Marcus I. Schlueter, and James M. Hanna, Jr. Department of Chemistry, Physics, Geology and the Environment, Winthrop University, Rock Hill, SC

introduction/background. During the last several years, visible-light photoredox catalysis (VLPC) has been developed into a practical method to achieve a wide variety of synthetic transformations. The approach often involves a transition metal complex of ruthenium or iridium, which, upon absorption of visible light, can participate in a series of single-electron-transfer (SET) events with organic substrates, leading to productive chemistry. However, these transition metal catalysts can be quite expensive, which led our group to investigate the use of less expensive organic photocatalysts in our studies of the alkylation of aryl imines with potassium organotrifluoroborates, the product of which (an alphaarylamine), is a substructure found in about 10% of the top-selling pharmaceuticals in the U.S.

hypothesis/goal of study. Previous optimization studies in our group established that the organic photocatalyst 9-mesityl-10-methylacridinium tetrafluoroborate (Mes-Acr-Me) in dichloromethane solvent, using 450 nm LEDs under an inert atmosphere gave the best yields in the alkylation protocol. In this study, the reactions of various potassium organotrifluoroborates with several aromatic aldimines were examined under these conditions to establish the scope and limitations of this protocol.

methods and results. The reactions of various potassium organotrifluoroborates with several aromatic aldimines were examined under the above conditions (Mes-Acr-Me, 450 nm LEDs, dichloromethane solvent, Ar atmosphere), and the protocol appears to be applicable to broad range of substituted imines, including those substituted with both electron donating and electron withdrawing groups.

conclusions. Visible light photocatalysis using an organic photocatalyst is a viable method for the alkylation of aromatic aldimines with potassium organotrifluoroborates, leading to alphaarylamines in moderate to good yields.

citation/acknowledgements. We would like to the the donors of the American Chemical Society Petroleum Research Fund (58270-UR1). Additional support was provided by an SC INBRE grant from the National Institute for General Medical Sciences (P20GM103499). We would also like to thank the Winthrop Department of Chemistry, Physics, and Geology.

The Synthesis of Diarylpyridines as Inhibitors of Amyloid Beta Aggregation for Alzheimer’s Disease

Casey Kopyc, Mary Stegall-Smith, James M. Hanna Jr., and Robin K. Lammi Department of Chemistry, Physics, Geology & the Environment, Winthrop University, Rock Hill, SC

introduction/background. Amyloid-beta peptide (A-beta) self-assembles into neurotoxic, betastructured aggregates, which are the primary component of the extracellular senile plaques characteristic of Alzheimer’s disease. A variety of small molecules have been shown to inhibit the aggregation process; typically, these contain aromatic groups and one or more hydrogenbond donors. Previous studies in our group have demonstrated that terphenyltetrols exhibit some degree of efficacy as Aβ aggregation inhibitors. For example, o-terphenyl-3,3″,4,4″-tetrol (OTT) is a moderately effective inhibitor of A-beta aggregation (IC50 = 2.7±0.3X). Recent modeling studies suggest that binding of small molecules to A-beta may occur via several types of intermolecular interactions, including both hydrogen bonding and pi-pi interactions (i.e., pi-stacking). In addition, other studies indicate that pi-interactions between benzene and electron-deficient heterocyclic aromatic rings are stronger than similar benzene-benzene interactions.

hypothesis/goal of study. Based on these observations, we hypothesized that incorporation of a pyridine unit as the central linker in the above-described tetrahydroxyteraryl scaffold may lead to increased inhibition of A-beta aggregation.

methods and results. We therefore synthesized several bis(dihydroxyphenyl)pyridines via Suzuki coupling of 3,4-dimethoxybenzene-boronic acid with an appropriate dibromopyridine, followed by demethylation in refluxing aqueous HBr. Evaluation of these compounds was carried out using a Congo red spectral shift assay to assess efficacy of A-beta aggregation inhibition.

conclusions. Preliminary results indicated that both 2,3-bis(3,4-dihydroxyphenyl)pyridine and 2,4-bis(3,4dihydroxyphenyl)pyridine exhibited some inhibition of A-beta aggregation.

citation/acknowledgements. Support

was provided by an SC INBRE grant from the National Institute for General Medical Sciences (P20GM103499). We would also like to thank the Winthrop Department of Chemistry, Physics, and Geology.

Characterizing azole drugs in the fungal pathogen Cryptococcus neoformans

Sakina Naqvi, Jordyn Wilson, and Srikripa Chandrasekaran Department of Biology, Furman University, Greenville, SC

introduction/background.

Understanding mechanisms of anti-fungal drugs in the pathogenic fungus, Cryptococcus neoformans, is crucial for improving anticryptococcal therapy. We have previously determined that presence of anti-oxidants in growth medium can reverse the fungal inhibition caused by the azole drug, fluconazole. However, treatment with the azole drug, fluconazole (32 ug/ml), presents the problem of resistance of C. neoformans to fluconazole treatment.

hypothesis/goal of study.

We screened several azole drugs, including voriconazole (1.75 ug/ml), propiconazole (6.25 ug/ml), cyproconazole (2 ug/ml), miconazole (5 ug/ml), isavuconazole (0.5 ug/ml), tebuconazole (12.5 ug/ml), myclobutanil (2 ug/ml), and ketoconazole (1.75 ug/ml) to determine the following: (1) ability to inhibit growth of H99 strain of C. neoformans, (2) MIC of the azole drugs in H99 strain of C. neoformans, (3) ability to induce Reactive Oxygen Species (ROS) in H99 strain of C. neoformans, and (4) ability of the anti-oxidant vitamin C (10mM) to reverse growth of H99 in the presence of each of the drugs.

methods and results. In this study, spot assays and disc diffusion assays were conducted. Furthermore, ROS levels were measured with flow cytometry and with plate reader. We found that all azole drugs inhibit growth of H99 and identified the MIC of the drugs against H99. Isavuconazole, voriconazole, and propiconazole showed the largest zone of inhibitions and the smallest MIC among all the drugs tested. However, growth of H99 in presence of these drugs (voriconazole, propiconazole, and isavuconazole) and vitamin C was reversed. Other drugs that showed decreased inhibition of H99 in the presence of vitamin C were ketoconazole and cyproconazole. Inhibitory effects of miconazole, tebuconazole, and myclobutanil were not

affected by vitamin C. We found that all the drugs whose inhibitory functions were lowered by vitamin C also induced higher Reactive Oxygen Species (ROS) production in H99.

conclusions. We concluded that while all the eight azole drugs tested inhibited H99 growth, only five drugs (voriconazole, propiconazole, isavuconazole, ketoconazole, and cyproconazole) induced production of ROS in C. neoformans.

citation/acknowledgements. Support was provided by an SC INBRE Developmental Research Project Program grant awarded tp Dr. Srikripa Chandrasekaran.

CB-06

Effects of Antioxidants on Fluconazole Resistance in Cryptococcus neoformans

Neha Bhatnagar and Srikripa Chandrasekaran Biology Department, Furman University, Greenville, SC

introduction/background. Cryptococcus is an encapsulated yeast, which can infect immunocompromised individuals. The most commonly prescribed anti-fungal drug to combat Cryptococcus infection is fluconazole (FLC). FLC, a fungistatic drug, inhibits cell division and growth of Cryptococcus without killing fungal cells. FLC is implicated in the generation of ROS, which contributes to oxidative stress.

hypothesis. We hypothesized that if the ROS generated by FLC contributed to the inhibition of growth of Cryptococcus, treatment of Cryptococcus with a combination of FLC and antioxidants would reverse the inhibition of the anti-fungal drugs. Since FLC generates oxidative stress in C. neoformans, then the antioxidants rutin (125 µg/ml), resazurin (60 µM), coenzyme Q-10 (5µMl), thioctic acid (1 mM), trolox (1 mM), capsaicin (50 µg/ml), and lutein (0.04 µg/ml) should reverse the inhibition caused by FLC (32 µg/ml) and rescue Cryptococcus growth. We used previously tested antioxidants (vitamin A, vitamin C, and Pyrrolidine Dithiocarbamate) as controls.

methods and results. Our studies have shown that Cryptococcus is rescued from FLC-mediated inhibition in the presence of the natural antioxidant thioctic acid. We used spot assays to determine the extent of C. neoformans growth in rich YPD (Yeast extract, Peptone, Dextrose) media supplemented with FLC alone or FLC and an antioxidant.

conclusions. We can conclude that certain antioxidants, including thioctic acid, can recover C. neoformans from FLCmediated inhibition. The differences in the various antioxidants to facilitate rescue is likely due to the different modes of action of each of the antioxidants in their ability to reverse types of oxidative stressors.

citation/acknowledgements. Support was provided by an SC INBRE Developmental Research Project Program award presented to Dr. Srikripa Chandrasekaran at Furman University.

Interaction Between the Viral RNA Leader Sequence and nsp1 in SARS Coronavirus

Kaitlin Marie Caughman, Jonathan Luke Cromer, Anita Nag Department of Natural Sciences, University of South Carolina Upstate, Spartanburg, SC

introduction/background. Nonstructural protein 1 (nsp1) of severe acute respiratory syndrome coronavirus (SARS-CoV), inhibits host translation by blocking the translation initiation site on the 40S ribosome and cleaving host mRNA. Stem-Loop 1 (SL1) of the viral RNA leader sequence has been identified to bind to nsp1, allowing viral RNA to escape translation repression. However, the specific residues on nsp1 and the specific sequences on SL-1 important to binding have not been experimentally verified.

hypothesis/goal of study. To investigate this binding, we used gel-shift assay and RNA pull-down to verify the binding between nsp1 and SL1. By mutating SL-1 and nsp1, we seek to map the sequence responsible for this interaction. Interestingly, nsp1 is a small protein with intrinsically unstructured regions at both C- and N- terminal ends of the protein. Based on recent literature we hypothesize that the C-terminal domain binds to the stem structure of SL1 and disrupting the stem region of SL1 will decrease binding between nsp1 and SL-1.

methods and results. To investigate the binding of nsp1 to SL1, we used nsp1 purified from bacterial overexpression using glutathione beads followed by precision protease cleavage of GST-nsp1, and biotinylated RNA. We used chemiluminescence to detect the RNA in a complex with nsp1 using a gel shift assay. Contrary to our hypothesis, we found an increase in nsp1 binding to the RNA carrying stem mutation and a decrease in nsp1 binding to the RNA with the loop mutation. Currently, we are further investigating several mutations in SL1 to identify the actual binding site.

conclusions. We confirmed the complex formation by nsp1 directly on SL1 using a reconstituted system. Since disrupting the stem-loop region of the RNA decreased binding to nsp1, this region may be critical in allowing viral RNA to escape translation repression.

citation/acknowledgements. Dr. Nag is supported by the SC INBRE DRP award (2020-23). The nsp1 plasmid was a kind gift from Dr. Makino at UTMB.

Pd Catalyzed Cross-Coupling of Alkylbisboronic Esters

Ellie Kraichely and Timothy J. Barker Department of Chemistry and Biochemistry, College of Charleston, SC

introduction/background.

Suzuki-Miyauru cross-coupling reactions are important in organic synthesis because of their ability to synthesize various biaryl compounds relatively easily.

hypothesis/goal of study.

The purpose of this research has been to develop a method for selective monocoupling of an alkylbisboronic acid pinacol ester with identical C-B bonds with an aryl halide.

methods and results. The alkylbisboronic acid pinacol ester used was 1,2-bis(4,4,5,5-tetramethyl-1,3,2dioxaborolan-2-yl)ethane which was synthesized via hydroboration reaction conditions using vinyl boronic acid pinacol ester. The alkylbisboronic acid reagent was used in reactions with varying aryl bromides along with a palladium catalyst and ligand. Results found that reactions with 4-bromotoluene had the most significant yields at a 0.2 mmol scale. Optimization efforts using various ligands and palladium sources were performed to broaden the scope of aryl halides.

conclusions. Monocoupling one of two identical C-B bonds was achieved without the need of protecting groups or the use of steric differences between C-B bonds. Additional optimization efforts are needed to increase the scale of the reaction.

citation/ acknowledgements. This project was supported by SC INBRE institutional funds to the College of Charleston’s Chemistry and Biochemistry department.

Identification of Small Regulatory RNA UspS Associated with the Universal Stress Protein in Lactobacillus Species

Zarah M. Fowler1,2, Sasha S. Bronovitskiy2, Finn K. Rose2,3, Gabriela C. Pérez Alvarado2, and Brian M. Lee2

1Department

of Biology, Coastal Carolina University, Conway, SC, 2Department of Chemistry, Coastal Carolina University, Conway, SC, 3Scholars Academy, Coastal Carolina University, Conway, SC

introduction/background. The gut microbiome is a complex habitat for a variety of bacterial species. These bacteria play vital roles in regulating several physiological processes in the body such as metabolism, digestion, and immune response. With the rise in use of probiotics to combat human disease, it is important to understand the mechanisms by which probiotic bacteria regulate host interactions. Common bacteria in probiotics include lactic acid bacteria: Lactobacillus acidophilus and Lactobacillus bulgaricus. Further exploration of physiological functions of probiotic bacteria will elucidate the role of small regulatory RNAs or non-coding RNAs in regulating gene expression within the human body.

hypothesis/goal of study. The goal of this project was to identify and explore the conservation of function and structure of the sRNA UspS in two probiotic bacteria and to further analyze the role of UspS in host interactions.

methods and results. A candidate sRNA in Lactobacillus species was chosen based on its association with a downstream universal stress protein. A sequence alignment between Lactobacillus species was performed. The sequence was characterized using computational methods to predict secondary structure, tertiary structure, and mRNA interactions of UspS. Genes for UspS were isolated from two Lactobacillus species, and target sRNAs were isolated using a T7 RNA transcription. A thermal melt assay was performed to confirm the presence of secondary structure.

conclusions. Sequence alignment of UspS shows conservation between Lactobacillus species. Secondary and tertiary structure predictions show a conserved pseudoknot region of the P4 region of UspS. The conserved P4 region of UspS may correspond to 6S RNA found in E. coli. The presence of a universal stress protein in both bacterial species suggests UspS may regulate the expression of usp. UspS was successfully synthesized by in vitro transcription, and a thermal melt assay of UspS suggests the presence of secondary structure.

citation/acknowledgements. This project funded by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health (NIH), Grant P20GM103499 (SC INBRE).

Meet Zarah Fowler

Coastal Carolina University, Biology

My name is Zarah Fowler. I am the Women's Club Volleyball President at Coastal Carolina University. It is honestly great to be in the classroom, and involved in athletics at our school! In the position of club president, I typically have many leadership roles. I collaborate with other schools to create organized tournaments, serve as a liaison between club sports and campus recreation, arrange meetings and fundraisers, and plan practices for our week! We are actually currently in the process of creating a fundraiser with an app called Scoreboard to raise money to participate in the collegiate club national tournament next year. As a club we do a lot of travel during the semester to other schools. This semester we will be playing in tournaments at USC, UNCW, and ECU. It is a really good time because we stay in hotels together, have team dinners, and participate in highly competitive volleyball. My team is my second family here at Coastal, and my research professors are my third. I literally go from labs that end at 5:30 pm to running around the gym playing volleyball by 6 pm. It is an amazing thing, and I am glad to hear of other students participating in research and athletics at their schools!

As far as my research goes, I do research under the supervision of Dr. Brian Lee at Coastal Carolina University in the chemistry department. My research focuses on sRNAs in probiotic bacteria that regulate host interactions within the gut microbiome.

Studying the Transposition Mechanism of the mJing Miniature Inverted Repeat Transposable Element in Yeast

Megan Collins and C. Nathan Hancock Department of Biology and Geology, University of South Carolina Aiken, Aiken, SC

introduction/background.

Transposable elements are DNA segments that can jump in and out of the genome, causing mutations. The mJing miniature inverted repeat transposable element found in rice is mobilized by proteins encoded by the transposable element Jing. Studies with a related element, mPing, showed a cut and paste mechanism of mobilization catalyzed by the ORF1 and Transposase proteins.

hypothesis/goal of study. We hypothesize that mJing transposition requires two proteins encoded by Jing, however, the exact sequences are unknown. We set out to develop a yeast transposition assay to further define the Jing coding sequences and investigate the mJing transposition mechanism.

methods and results. We designed and cloned Jing ORF1 and Transposase expression constructs based on predicted splicing from the consensus across three Jing elements. We also developed a reporter plasmid, containing mJing inserted into the ADE2 gene. These plasmids were transformed into yeast, and a transposition assay was performed. by plating liquid cultures onto galactose plates lacking adenine. Transposition of mJing should have resulted in a functional ADE2 gene, allowing colonies to grow. Unfortunately, no colonies were observed from the yeast transposition assay.

conclusions. We were successful in creating the expression clones of the predicted Jing ORF1 and TPase proteins, as well as the plasmid containing the mJing reporter construct. The fact that we didn’t observe any transposition events suggests that we incorrectly predicted the splice sites. To identify the splice sites, we will express a genomic version of Jing in Arabidopsis to obtain Jing cDNA sequences. This will allow us to identify the coding sequences needed for transposition of the mJing element.

citation/acknowledgements. This work funded by SC INBRE, USCA Biology and Geology.

Isolation and Analysis of Extracellular Vesicles from Lactic Acid Bacteria

Isabel Myers, Brooke Busby, Klea Hoxha, Gabriela C. Pérez Alvarado, and Brian M. Lee Department of Chemistry, Coastal Carolina University, Conway, SC

introduction. Lactic acid bacteria have probiotic properties and are found within the microbiome of the human gut having a significant influence on human health. The extracellular vesicles are produced by the probiotic bacteria of the gut and are thought to be involved in intercellular communication with other bacteria and with the host. Extracellular vesicles (EVs) are produced by prokaryotic bacteria and derived from the outer membrane in Gram-negative bacteria. The thick layer of peptidoglycan present in Gram-positive bacteria was thought to prohibit release of EVs. Recent studies have suggested that autolysin may allow EVs to be released from Gram-positive bacteria.

hypothesis/goal of study. The goal of the study was to develop an isolation procedure and observe the contents of the isolated EVs from lactic acid bacteria. It was hypothesized that the protein and RNA content of the EVs could provide greater insight into the mechanisms of intercellular communication.

methods and results. Streptococcus thermophilus was grown in culture. The growth cultures were centrifuged to pellet the cells. The supernatant was filtered to remove the remaining bacterial cells. The filtrate was concentrated by ultrafiltration. The retentate was presumed to contain EVs, and gel electrophoresis was run to determine protein content. Scanning electron microscopy was used to observe the membrane surface of cells isolated from growth cultures and to look for budding EVs.

conclusions. The protein content observed during the process of isolation is indicative of the presence of EVs. Future studies include identification of the protein and RNA content of the EVs as well as further development of methods to improve isolation techniques.

citation/acknowledgements. Funded by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health (NIH), Grant P20GM103499 (SC INBRE)

Peanut allergen reduction using phytic acid functionalized magnetic chitosan nanoparticles

Johanna Bravo, Qurian Davis, Shemyia Smith, and Anselm Omoike Department of Chemistry, University of South Carolina Upstate, Spartanburg, SC

Peanuts (Arachis hypogea) are a nutritious plant protein that strengthens carbohydrates in foods widely consumed. Peanuts are an affordable and reliable source of proteins, vitamins, minerals, and other nutritional benefits. Although versatile, peanuts contain various proteins that are responsible for allergic reactions to predisposed individuals. In this study, phytic acid (PA) was used to functionalize magnetic nanoparticles (MNP) that were coated with low molecular weight chitosan (CH). The synthesized MNP were exposed to extracts from three varieties of peanuts: Runner, Spanish, and Virginia. Allergen extraction was monitored by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE)

and bicinchoninic acid (BCA) analysis. The SDSPAGE and BCA displayed the removal of Ara h 1 and Ara h 2 allergens. BCA results shows over 69%, 44%, and 59% removed from Runner, Spanish, and Virginia, respectively. MNP-CHPA may serve as an inexpensive method for removing protein allergens in peanuts.

citation/acknowledgements. This

Detecting Sequences Pseudo-nitzchia Species Using HRM Primers

Zachary Padgett1, Aramis Lawson1, Elaina Renshaw2 and Megan Cevasco1

1Department of Chemistry and Biochemistry, Coastal Carolina University, Conway, SC, 2Department of Biological Sciences, University of South Carolina, Columbia, SC

introduction/background. Diatoms are photosynthetic single-celled microalgae which inhabit both marine and freshwater systems. The marine diatom genus Psuedo-nitzchia is found in public shellfish harvesting grounds along South Carolina coast. Pseudo-nitzchia is important to study because it has the potential to produce the neurotoxin domoic acid that causes amnesic shellfish poisoning in consuming taxa.

objective/hypothesis. The objective of this study is to test the environmental DNA (eDNA) of the water samples in the public shellfish harvesting ground at Huntington Beach State Park with specific primers to see if the genus Pseudo-nitzchia is present. The amplicons recovered will be analyzed via nanopore sequencing to determine if the Pseudonitzchia taxa found are domoic acid producers.

methods and results. The eDNA was collected, and then extracted and quantified PCR and QPCR methods were employed with the specific primers (HRM F/R). After the PCR was complete, gel electrophoresis was used to determine if there was Pseudo-nitzcha present in the sample. Upon confirmation, a nanopore sequencing method was used to determine which species within the genus Pseudo-nitzchia were present. These data were assembled in a phylogenetic tree to observe taxonomic diversity patterns. The sequences indicate that some of the Psuedo-nitzchia present at HBSP have the potential to produce domoic acid.

conclusion. The genus Pseudo-nitzchia was found in the waters of Huntington Beach State Park. The sequencing of the water samples shows that some of the Pseudo-nitzchia present are potential domoic acid producers.

citation/acknowledgements.

This presentation was supported in part by grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health.

Identification and Analysis of the Regulatory RNA TRMS in the Probiotic Bacteria Streptococcus Thermophilus

Finn K. Rose1,2, Zarah M. Fowler2,3, Sasha S. Bronovitskiy2, Gabriela C. Pérez Alvarado2, and Brian M. Lee2 1Scholars Academy, 2Department of Chemistry, 3Department of Biology, Coastal Carolina University, Conway, SC

background. The human microbiome contains billions of bacteria, primarily found in the gut. Many of these bacteria are non-pathogenic and could have a beneficial relationship with our cells. Some non-pathogenic species of interest include Lactobacillus bulgaricus, Lactobacillus acidophilus, and Streptococcus thermophilus for their frequent use in the dairy industry and therefore their likelihood of being found in the gut. A way of further understanding these bacteria is to study how they regulate essential functions related to survival, reproduction, and potential host interaction. Genes can be regulated at the transcriptional level by regulatory RNA which can fold into hairpin structures that can interact with other RNA, proteins, or both.

goal of study. The goal of this study is to identify and characterize regulatory RNA in S. thermophilus along with their associated proteins to characterize potential communication between probiotic bacteria and host cells.

methods and results. Candidate RNA TrmS was identified based on a potential cis-regulatory element that is downstream of a tRNALeu methyltransferase in S. thermophilus. Secondary and tertiary structures were predicted. TrmS DNA was isolated and amplified through PCR, and RNA was synthesized through T7 transcription reactions, which was confirmed using gel electrophoresis. Thermal melt assays were performed to confirm the presence of secondary structures.

conclusions. TrmS was successfully isolated from S. thermophilus. Predicted secondary structures were confirmed through thermal melt assays, allowing further prediction of the mechanism of interaction between TrmS and the neighboring gene that encodes for a predicted tRNALeu methyltransferase protein. TrmS sequence alignments identified conserved loops regions in TrmS across pathogenic and non-pathogenic bacterial species, suggesting an evolutionary purpose for TrmS. Protein alignment of the neighboring gene with proteins of known structure and function in E. coli suggests a similar function in methylating the anticodon loop of tRNALeu in the protein found in S. thermophilus.

citation/acknowledgements. Funded by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health (NIH), Grant P20GM103499 (SC INBRE).

Synthesis of 1-phenylpent-4-en-2-ol

Alex Martin and Timothy Barker Department of Chemistry and Biochemistry, College of Charleston, Charleston, SC

introduction/background.

Reactions involving pinacol boronic esters are not well understood or researched. Therefore, there is a piqued interest in these types of reactions. From previous experiments, the pinacol boronic ester used in this reaction has been synthesized.

hypothesis/goal of study.

The goal of this synthesis is to prepare 1-phenylpent-4-en-2-ol in good yield.

methods and results. The reaction starts by stirring styrene oxide, silver tetra borohydride, and allyl boronic acid pinacol in THF. During this study the solvent, limiting reagent, catalyst, and temperature have varied to optimize the reaction. To confirm the product was successfully produced proton NMR is used to characterize.

conclusions. The synthesis of 1-phenylpent-4-en-2-ol has been successfully prepared. The reaction optimization is still in process to improve the yield of this reaction by increasing mole percent of the catalyst as well as mole ratios of reagents.

citation/acknowledgements. This work funded by SC INBRE.

Nucleic Acid Aptamer Au/Ag Nanoparticle

Conjugates as TrojanHorse drug Delivery Vehicles in the Fight Against Bacterial Infections

Kierra McCall1, Morgan Hunter1, Joshua Quarles1, Ashley Wood1, Allen Livingston1, Victoria Frost2 and Timea Gerczei Fernandez1

1Department of Chemistry, Physics, Geology and the Environment and 2Department of Biology, Winthrop University, Rock Hill SC

Illnesses caused by bacteria are a major public health concern since microorganisms became increasingly resistant to available antibiotics. At the same time big pharma gradually shifted its focus from developing drugs that cure diseases to those that treat chronic conditions. Thus, rediscovering old drugs and using them for new purpose became more important. The goal of this project is to use nucleic acid aptamernanoparticle conjugates as vehicles to deliver the antibiotics to cells that are resistant to them.

Currently, we are investigating the therapeutic potency of nucleic acid-silver/gold nanoparticle conjugates as carriers of tetracycline, ampicillin, and kanamycin as treatment against E Coli infection. We hypothesize that by attaching nucleic acids that bind to these antibiotics to silver or gold nanoparticles the resulting conjugates will work as a “Trojan-horse” drug-delivery vehicle that smuggles the antibiotic into the cell without being detected by cellular defense systems. Moreover, we reason that silver or gold ions released by the nanoparticles add to the antimicrobial effects of the antibiotics.

To test the viability of this idea we used a tetracycline and an ampicillin binding DNA aptamer that were developed for detection of these antibiotics. We optimized conditions to attach these DNA aptamers to silver and gold nanoparticles. We are currently testing antimicrobial effect of these aptamer nanoparticle conjugates using E. Coli 29522.

citation/acknowledgements. This research was supported by SC INBRE P20GM103499.

dynamics changes in epigenetic modifications

Charlotte McGuinness1, Megan A. Wilson1, Maria Ouzounova2, Hasan Korkaya3, Austin Y. Shull1 1Department of Biology, Presbyterian College, Clinton, SC, 2Cancer Research Center, University of Lyon, Lyon, France, 3Georgia Cancer Center, Medical College of Georgia at Augusta University, Augusta, GA

introduction/background. Metastatic potential of basal-like breast cancers typically is initiated by genetic alterations that lead to a process known as epithelia-mesenchymal transitioning (EMT).

hypothesis/goal of study. With this premise in mind, it is important to better understand the intracellular consequences that ultimately lead to the EMT phenotype. Previous work from our lab has demonstrated the role epigenetic modifications through DNA methylation changes play in promoting the invasive characteristics of metastatic breast cancers.

methods and results. Based on the previous notion connecting epigenetic changes to breast cancer metastasis, we performed DIA-based mass spectrometry of isolated histones from an isogenic panel of MCF10A breast cell lines where the tumor suppressor genes TP53 and PTEN were silenced and induced EMT. With this approach, we determined which histone modifications were differentially enriched in the non-EMT and EMTinduced MCF10A cell lines. From approximately 72 histone modifications identified and annotated from our mass spectrometry, we were able to identify 5 histone events that were differentially enriched in our MCF10A cell line panel. Two events of note were histone H3 lysine-14 acetylation (H3K14ac) significantly increasing and histone H4 arginine 55 dimethylation (H4R55me2) significantly decreasing in our EMT-transformed MCF10A p53-/PTEN- cell lines when compared to the parental, non-tumorigenic MCF10A cell line.

conclusion. Collectively, our results demonstrates two histone modification events (H3K14ac &amp; H4R55me2) differentially affected during the EMT process in breast cancer cells. These

Histone proteomic profiling of EMTtransformed MCF10A breast cells reveal

two modifications highlight the potential for two targetable epigenetic drivers in breast cancer metastasis.

citation/acknowledgements. Research reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number P20GM103499. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Determining if the Bases Adjacent to the mPing Element Impact Transposition

M. Elizabeth Colón-LaBorde and C. Nathan Hancock Department of Biology and Geology, University of South Carolina Aiken, Aiken, SC

introduction/background. A transposable element is a segment of DNA that can move from one part of the genome to another. mPing is a small, transposable element that actively transposes in rice. Our laboratory studies the transposition mechanism of mPing because it is currently altering the rice genome and is being developed into a gene tagging tool for plant gene discovery. The established method for measuring the transposition of mPing in Arabidopsis uses a GFP reporter which only allows expression of the fluorescent biomarker when mPing mobilizes.

hypothesis/goal of study. We are attempting to determine if the bases adjacent to the mPing element affect its transposition. The consensus sequence that normally flanks mPing is TTA:TAA. Experiments in yeast show that the element excises more when flanked by TTA:TAA and less when flanked by GGT:ACC. Our goal was to make constructs that would allow us to test the effect of these flanking sequences in Arabidopsis.

methods and results. Using PCR and NEB Builder cloning, we created four different plasmids containing mPing or mmPing20 (a hyperactive version of mPing) with adjacent TTA:TAA or GGT:ACC sequences. Preliminary results showed evidence of transposition for all four constructs. We will transform these constructs into an Arabidopsis line that contains the ORF1 and Transposase genes needed for transposition. This will allow us to use PCR and fluorescence microscopy to determine the relative frequency of transposition.

conclusion. We predict that elements with adjacent TTA:TAA will have higher transposition rates, indicating that these bases are contributing to the transposition mechanism. This is important as it will help us to understand how transposable elements function and interact in all genomes.

citation/acknowledgements. This research was supported by SC INBRE P20GM103499.

The RhoG guanine nucleotide exchange factor SGEF controls junctional localization of desmosomal proteins

Hannah C. Lee1, Agustin Rabino2, Rafael Garcia-Mata2, and Adi D. Dubash1 1Biology Department, Furman University, Greenville SC, 2Department of Biological Sciences, University of Toledo, Toledo OH

introduction/background.

The desmosome is a cell-cell adhesion complex which facilitates the mechanical stability of tissues. Disruption of the desmosome is observed in a range of different cutaneous syndromes, several types of cardiomyopathy, and in cancer, where loss of desmosomal proteins accompany epithelial to mesenchymal transition (EMT). Determining the signaling mechanisms which control desmosome structure is therefore relevant for a wide range of different pathologies.

hypothesis/goal of study.

Recent work has uncovered an important role for the protein Src Homology 3 (SH3)containing GEF (SGEF) in maintaining the structure of both adherens junctions and tight junctions in epithelial cells. SGEF is a guanine nucleotide exchange factor specific for the RhoG GTPase, as it enhances the activity of the GTPase via promotion of GDP-GTP exchange. In our study, we analyzed a potential role for SGEF in regulation of the desmosome.

methods and results. Control and SGEF KD MDCK cells (obtained from Dr. Rafael Garcia-Mata, University of Toledo) were cultured on coverslips, fixed at steady state and processed for immunofluorescence for a range of different desmosomal proteins.

Images were acquired using a Leica SP8 Confocal Microscope. Compared to control MDCKs, SGEF knockdown cells demonstrate a significant reduction in the junctional localization of several

different desmosomal proteins, such as Desmoplakin, Desmoglein-2 and Plakoglobin. These changes are not due to alteration of gene expression, as total protein levels (assessed by western blot) are not changed upon SGEF knockdown. Interestingly, we observe a similar reduction in localization (but not expression) of desmosomal proteins upon knockdown of RhoG.

conclusions. These studies have uncovered an important role for SGEF and RhoG in maintaining the localization of desmosomal proteins. Future work will focus on whether regulation of desmosomal structure by SGEF requires its guanine nucleotide exchange function and whether SGEF-mediated changes in desmosome protein localization affect junctional strength and integrity.

citation/acknowledgements. This project was funded by an NIH South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE) grant #5P20GM103499. These funds provided a Faculty Fellows award to AD Dubash. All experiments described here were conducted at Furman University by H.C. Lee (supported by INBRE Faculty Fellows Award) and AD Dubash.

Josh M. Powers, Maya M. Mrus, Beatriz T. Mateo, and Adi D. Dubash Biology Department, Furman University, Greenville SC

introduction/background.

Desmosomes are protein complexes crucial for maintaining cell-cell adhesion and integrity of tissues. These complexes are made up of proteins from three families: transmembrane cadherins (Desmoglein and Desmocollin) link adjacent cells in the extracellular space, armadillo proteins (Plakophilin and Plakoglobin) stabilize the intracellular plaque, and the cytolinker Desmoplakin (DP) connects the plaque to the intermediate filament network.

hypothesis/goal of study. Desmosomal proteins have also been shown to coordinate gene expression pathways required for processes such as proliferation, differentiation and cell migration. In particular, several lines of evidence have linked desmosomal proteins to gene expression of extracellular matrix (ECM) proteins. In our study, we sought to investigate the role of desmosomal cadherins in ECM gene expression.

methods and results. Compared to control A431 cells, Dsg2 knockout cells demonstrated an increase in expression of Fibronectin (FN1) mRNA, and minor changes in expression of Collagen 1 (COL1A1) and Collagen 2 (COL2A1) mRNA. An increase in expression of Fibronectin protein levels in Dsg2-deficient cells was also observed via western blot. Increased expression of ECM genes was also observed upon siRNA-mediated knockdown of Dsg2 in both A431 cells and HaCaT keratinocytes. An extensive analysis of signaling pathways known to be involved in regulation of ECM gene expression uncovered an important signaling role for Src kinase and the NF-kB transcription factor, as abrogation of either protein (via knockdown or pharmacological inhibition) resulted in a rescue of Dg2KOmediated increases in FN1 gene expression. Moreover, we have documented a significant increase in localization of NF-kB to the nucleus of Dsg2-deficient cells, indicating an increase in NF-kB transcriptional activity.

conclusions. Taken together, our study highlights a novel role for Dsg2 in mediating ECM gene expression via Src/NF-kB signaling, adding significant insight into the mechanisms by which desmosomal cadherins control the adhesive behavior of cancer cells.

citation/acknowledgements. This project was funded by an NIH South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE) grant #5P20GM103499. These funds provided a Faculty Fellows award to AD Dubash.

MCB-05

The desmosomal cadherin Desmoglein-2 controls extracellular matrix gene expression via Src and NF-kB signaling

Profiling the response of individual gut microbes to common nutritional supplements used in the neonatal intensive care unit (NICU)

Megan E. Waller1, Caroline J. Eichhorn1, Taylor D. Ticer2, Janiece S. Glover1, John E. Baatz3,4, Carol L. Wagner3,4, Katherine E. Chetta3,4, Melinda A. Engevik1,2

1Department of Regenerative Medicine & Cell Biology, Medical University of South Carolina, 2Department of Microbiology & Immunology, Medical University of South Carolina, 3Department of Pediatrics, C.P. Darby Children’s Research Institute, Medical University of South Carolina, 4Department of Pediatrics, Division of Neonatal-Perinatal Medicine, Medical University of South Carolina, Shawn Jenkins Children’s Hospital, Charleston, SC

introduction. For pre-term infants in the neonatal intensive care unit (NICU), adequate nutrition requires various enteral products, including human milk and sometimes formula. Human milk is typically fortified to meet increased protein and calorie goals and infants commonly receive other supplements.

hypothesis. Because diet is a significant driver of the gut microbiota, we hypothesized that various enteral products would impact the growth of common microbial strains found in the infant gut.

methods and results. To address how individual gut microbes respond to additives commonly used in the NICU, we cultured 8 distinct Bifidobacterium and 8 Lactobacillus strains as well as 10 gut pathobionts from the genera Streptococcus, Staphylococcus, Enterococcus, Acinetobacter, Proteus, Escherichia, Enterobacter, and Klebsiella all found commonly in the premature infant gut. All bacteria were grown in chemically defined bacteria medium and monitored for growth after 20 hrs of incubation. Increased bacterial growth was seen in almost all organisms after supplementation with term HMOcontaining formula. We identified significantly higher growth between unfortified and fortified freshly-pasteurized milk and donated, indicating that both groups could use the nutrients in multicomponent fortifiers to grow. In general, supplementation with thickeners or NaCl did not stimulate the growth of the tested microbes. Generally, vitamin mix promoted the growth of commensal microbes, while iron promoted pathobionts over commensal microbes.

conclusions. Pathobionts in the preterm gut have significant growth with preterm formula, fortified donor milk, and supplemented iron. These microbes do not have the same growth advantage with fresh milk, but the addition of multicomponent fortifiers to fresh milk promoted growth universally. We also found that commensals are adaptable and grow well with formula, donor and fresh human breast milk, with and without fortification, and supplemented vitamins. These data indicate that the choice of milk and supplements may collectively impact the infant gut microbiota.

citation/acknowledgements. This project was funded by SC INBRE grant #5P20GM103499 and the Medical University of South Carolina.

Meet DRP Recipient Mindy Engevick

(FY2022-2023)

Institution/Department: Medical University of South Carolina, Regenerative Medicine & Cell Biology

Project Title: Microbial suppression of intestinal inflammation and oxidative stress

How SC INBRE funding will help accomplish project goals: The SC INBRE grant will allow me to explore the novel hypothesis that the diet influences microbial metabolites that can suppress intestinal inflammation and provide preliminary data for an R01 application. This award will also provide my research with increased visibility and be a valuable steppingstone for my career.

Outside the Lab/Classroom: I love to travel!

Learn more about Dr. Engevik at https://bit.ly/scinbre-spotlights

signature in breast cancer stem cells

1Department of Biology, Presbyterian College, Clinton, SC, 2Cancer Research Center, University of Lyon, Lyon, France, 3Georgia Cancer Center, Medical College of Georgia at Augusta University, Augusta, GA

introduction/background. Metastatic potential of basal-like breast cancers typically correspond with enrichment of EpCAM-/CD49f- cancer stem cells (CSC). With this premise in mind, it is important to better understand the mechanistic drivers that allow these aggressive CSC populations to expand. Previous work from our lab found the IL32 promoter to be differentially hypomethylated in CSC-enriched cell lines, leading to its overexpression and potential downstream role in CSC expansion.

hypothesis/goal of study. Based on this previous work, we further sought to characterize IL32’s role in breast cancer aggressiveness through differential gene expression analysis via RNAseq and a multi-pathway phosphorylation protein array that evaluated the MAPK, AKT, JAK/STAT, TGFB, and NFKB pathways.

methods and results. We first identified several overarching mechanisms altered in siIL32 treated CSCenriched cells by RNAseq (FDR p-value <0.01). Most notable was the enrichment of upregulated pathways involved in extracellular matrix (ECM) organization as well as enrichment of downregulated pathways involved in cellular and replicative stress responses. Furthermore, IL32 suppression decreased cell invasion in both an ECM-matrix cell invasion assay and a chick CAM xenograft/angiogenesis model. Furthermore, our RNAseq results corresponded with our protein phosphorylation array where we observed a decrease in phospho-JNK and phospho-NFKB in siL32-treated cells, both of which are well-established events that can coordinate both cell stress responses and cellular invasion.

conclusions. Collectively, our results reflect the notion that differential IL32 expression by promoter hypomethylation in breast CSCs plays a role in mitigating intracellular stress and subsequently allowing for breast cancer cell invasion.

Cytotoxicity in Human Breast Cancer MCF-7 Cells following Exposure to Polycyclic Aromatic Hydrocarbons, Estrogen, and Tamoxifen

Steve Samson1, Selby Artis1, and Samir Raychoudhury2 1Ridge View High School, Columbia, SC, 2Department of Biology, Chemistry and Environmental Health Science, Benedict College, Columbia, SC

introduction/background.

Polycyclic Aromatic Hydrocarbons (PAH) are ubiquitous and persistent environmental contaminants. Some of them are suspected carcinogens and may affect the reproductive systems as potential endocrine disruptors. The compounds in question for this experiment were estrogen (E2), tamoxifen (TAM), and two polycyclic aromatic hydrocarbons benzo(A)pyrene (BAP), and fluoranthene (FLA,). Estrogen activates a transduction pathway that leads to cell proliferation. Tamoxifen is a selective estrogen receptor modulator. It prevents estrogen from binding to the receptors on breast cells.

hypothesis/goal of study.

The purpose of this experiment was to examine the effect of different treatments on lactate dehydrogenase (LDH) activity. LDH is an enzyme involved in anaerobic respiration and its activity is a good indicator of cell damage. It was hypothesized that PAH would significantly increase LDH activity compared to the control.

methods and results. The

citation/acknowledgements.

Research

reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number P20GM103499. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

MCF-7 cells were cultured in 75cm2 flasks containing, 10ml of growth media. After confluence, the cells were trypsinized, counted, and distributed at 20,000 cells per well, in 96 well plates. We monitored the cells under

IL32 expression contributes to a cell stress response and ECM-remodeling expression

Megan A. Wilson1, Elayne M. Benson1, Emma V. Gray1, Paris L. Rizzo1, Paige Cairns1, Maria Ouzounova2, Hasan Korkaya3, Austin Y. Shull1

a microscope, and on day 2, we added exposure media, and media containing BAP, FLA, E2, and TAM (125ng/ ml or 500ng/ml). After 24 hours, the media was separated, and the conditioned media was saved in centrifuge tubes for the LDH assay using an ELISA plate reader to measure the absorbance of the samples at 490 nm. A one-factor ANOVA on the data showed that the results were statistically significant (p<0.05). We have performed multiple comparison tests using GraphPad Prism 9.5. The average absorbance values of all treatments were significantly greater than the absorbance of the media control (p<0.05). It is also important to note that for BAP, or FLA, LDH activity increased when the concentrations were increased from 125ng/ml to 500ng/ml. In contrast, LDH activity decreased when the exposure concentrations of E2 and TAM were increased from 125ng/ml to 500ng/ml. Additional experiments are needed to examine the mechanism of action of these varied toxic effects.

citation/acknowledgements. This project was funded by SC INBRE grant #5P20GM103499.

Testing HIV-dependency using firefly luciferase reporter plasmids

Kaitlynn Cook and William J. Jackson, Jr. Department of Biology and Geology, University of South Carolina Aiken, Aiken, SC

introduction. We are developing a cell culture model to test inducibility using a HIV-dependent plasmid to express firefly luciferase. This plasmid, p(LIR)LucF.pA, was constructed by cloning the HIV-1 promoter enhancer, the Rev Response Element, and the Gag Inhibitory Sequence into pcDNA3. Together, these elements should control reporter expression by inducing expression in the presence of HIV-1 regulatory proteins Tat and Rev. hypothesis. We hypothesize that p(LIR)LucF.pA will express firefly luciferase only in the presence of HIV-1 Tat and Rev and that expression will increase with the levels of Tat and Rev.

methods and results. Reporter gene expression studies were carried out using HEK293T cells transfected with either pCMV-LucF, a constitutive expression plasmid, or p(LIR)LucF.pA. Inducible expression was measured following co-transfection with pCMVR8.74, which expresses Tat and Rev. The response to Tat and Rev was measured using a range of pCMVR8.74 DNA (0-500 ng). Forty-eight hours post-transfection, cells were lysed, and luciferase activity was measured. Results showed low levels of luciferase activity in the absence of Tat and Rev. Luciferase activity increased in pCMVR8.74 co-transfected cells up to 250 ng pCMVR8.74; however, there were no significant changes in reporter expression between 250 ng and 500 ng pCMVR8.74.

conclusions. Cell culture results using p(LIR)LucF.pA showed very low levels of firefly luciferase expression in the absence of HIV-1 Tat and Rev. Importantly, reporter expression increased with the addition of Tat and Rev. Results also indicated HIV-dependent expression using p(LIR)LucF.pA is responsive to low levels (50 ng) of Tat and Rev, but expression levels flatten above 250 ng. This is likely due to the use of pCMVR8.74, which expresses high levels of Tat and Rev, and the comparatively limited availability of p(LIR)LucF.pA in co-transfected cells. It is expected that the addition of higher levels of p(LIR)LucF.pA would result in higher expression levels.

citation/acknowledgements. This work funded by SC INBRE grant #P20GM103499.

Investigating the Role of RNA Polymerase V in mPing Excision Site Repair

Kaili Renken and C. Nathan Hancock Department of Biology and Geology, University of South Carolina Aiken, Aiken, SC

introduction. Recent studies indicate that RNA plays a role in eukaryotic DNA repair. The plant-specific RNA polymerase V (Pol V) is mostly known for its role in gene silencing and can also be found aiding in the repair of double-stranded DNA breaks (DSB). The excision of DNA transposons creates DSB, but little is known about Pol V’s role in the repair of these sites. The transposable element mPing, is an active miniature inverted repeat transposable element that can be induced to transpose in Arabidopsis.

goal of study. Our goal is to test if Pol V protein is required in mPing excision site repair. Because Pol V has been shown to be required for efficient repair of other DSB, we predict that DNA repair of mPing excision sites will also be affected.

methods and results. We measured mPing excision site repair of a mPing:GFP reporter construct in multiple DNA methylation mutants including pol V and pol V 1-3 . We screened for GFP expression by fluorescence microscopy and analyzed the excision site by PCR and sequencing. We found that pol V and pol V 1-3 plants had significantly less GFP expression than the other lines tested, indicating that either mPing excision isn’t occurring or the DNA isn’t being repaired correctly. We detected mPing excision in all plants by PCR, but sequencing of the excision sites showed large deletions in plants lacking Pol V.

conclusions. These results suggest that RNAs produced by the Pol V protein play a role in the repair of mPing excision sites. Improper DNA repair in pol V mutants was the likely the cause of decreased GFP expression.

citation/acknowledgements.

Funding for this project was provided through SC INBRE and was supported by a grant from the National Institutes of Health National Institute of General Medical Sciences (P20 GM13499-20). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH.

Inducing

Cell Death

using a HIV-Dependent Expression System

Andrew P. Gregory and William H. Jackson Department of Biology and Geology, University of South Carolina Aiken, Aiken, SC

introduction/background. Elimination of infected cells in HIV-positive individuals is a challenge of current treatments. HIV-dependent expression of therapeutic genes might be used to specifically target/ eliminate HIV-infected cells. To investigate this, we created a HIV-inducible plasmid using the HIV-1 promoter/enhancer, the Rev Response Element, and gag inhibitory sequence. This plasmid was further modified by the addition of the Bcl-2-associated X protein (Bax), a pro-apoptotic gene that induces apoptosis when overexpressed. In this plasmid, p(LIR)Bax.pA, Bax expression is dependent on the presence of HIV-1 regulatory proteins Tat and Rev.

hypothesis/goal of study. Pro-apoptotic Bax expression from p(LIR)Bax.pA is expected to increase in the presence of HIV Tat and Rev, resulting in a parallel increase in Bax-induced cell death.

methods and results. Bax expression was measured in HEK293T cells transfected with either pCMVBax, which constitutively expresses high levels of Bax, or p(LIR)Bax.pA, which is inducible in the presence of Tat and Rev. To mimic HIV infection, p(LIR)Bax.pA transfections included pCMVR8.74, which expresses Tat and Rev. Forty-eight hours post transfection, cells were assayed for evidence of Bax expression. Cells transfected with pCMVBax showed decreased viability compared to negative controls transfected with pcDNA3. Cells co-transfected with p(LIR)Bax.pA and pCMVR8.74 showed decreased viability in a dose-dependent manner, while p(LIR)Bax.pA transfections in the absence of pCMVR8.74 were similar to negative controls. Similarly, caspase 3/7 activity increased in cells co-transfected with p(LIR)Bax.pA and pCMVR8.74, while transfections of p(LIR)Bax.pA alone showed caspase 3/7 activity equivalent to negative controls.

conclusion. In this study, HIV-dependent expression was observed in cells transfected with a plasmid constructed using three viral regulatory elements: the 5’ LTR, the RRE, and the gag inhibitory sequence. Together, these elements positively regulate expression of pro-apoptotic Bax in the presence of HIV Tat and Rev.

citation/acknowledgements. Funding for this project was provided through SC INBRE grant P20 GM13499-20.

The Effect of Cellular Differentiation on AdenoAssociated Viral Vector Genome Status

Farouk Chatila and Jennifer Lyles Departament of Francis Marion University, Florence, SC

introduction/background. Gene therapy is used to treat genetic disorders by introducing a functional copy of a mutated or absent gene. This requires a vector for gene delivery, and among the most successful gene therapy vectors is Adenoassociated virus (AAV). The genome of AAV vectors persists in the host cell nucleus in an episomal circular state independent of the host cell chromosomes. Previous research suggests that different molecular mechanisms are employed in undifferentiated (dividing) and differentiated (non-dividing) cells regarding AAV transduction and that this may have an impact on the final status, or confirmation, of the vector genome.

hypothesis/goal of study. The purpose of this study is to define the status of AAV vector genomes in undifferentiated and differentiated cells and to better understand the transition of vector genome conformation during the process of cellular differentiation.

methods and results. C2C12 cells (murine myoblasts) can be induced to terminally differentiate into myotubes via serum starvation. To establish a definitive timeline for analysis, cell morphology and myosin heavy chain (MHC) expression were analyzed daily for 14 days post-serum starvation using light microscopy and Western Blot analysis, respectively. Signs of cell elongation, alignment, and fusion, as well as MHC expression, were observed beginning 2 days postserum starvation.

conclusions. Elongation, alignment, and fusion of cells take place during C2C12 differentiation. Additionally, MHC is expressed only in myotubes and not in myoblasts. Therefore, these serve as markers for differentiation. Based on changes observed in cell morphology and MHC expression, it is likely that C2C12 differentiation begins approximately 2 days post-serum starvation. Saturation of these observations indicates differentiation may be complete by 7-9 days postserum starvation, but further analysis is required to confirm. Future work will focus on characterization of the molecular status of the AAV vector genome in these cell populations via qPCR and Southern Blot analysis.

citation/acknowledgements. This study was funded by SC-INBRE and the REAL Program at FMU.

Investigations of Nucleotide Modifications in Winthrop’s Bacteriophage Collection

Gabrielle Walker and Victoria Frost Department of Biology, Winthrop University, Rock Hill, SC

Bacteriophages (phages) are viruses that replicate in bacteria and have co-existed with their host in a complicated, evolutionary arms race for approximately three billion years. Both partners continuously evolve attack and defense mechanisms to ensure their own survival. Bacteria employ an array of defense mechanisms, including a Restriction-Modification System that utilizes restriction endonucleases (REs) to destroy infecting phage genomes. Methylation of the DNA bases at RE recognition sites is often used by bacteria to protect their own genomes. Phages can utilize methylation, or as appears to be more common, using noncanonical nucleobase substitutions. Purine and pyrimidine precursor molecules are modified by phage encoded genes. They are then incorporated into phage genomes during replication, and effectively “camouflage” their RE sites. The objective of this study was to reveal possible phage DNA modifications by comparing patterns of endonuclease cleavage of Winthrop’s collection of phage genomes, to virtual profiles of predicted endonuclease activity. Our protocols utilized methylation-insensitive enzymes, to reveal possible RE site protection in the phage genomes. In parallel, an in-silico tool, available at New England Biolabs (NEB), was used to generate virtual digestion patterns using the same suite of enzymes. Our results highlighted that several of our phages’ DNA was blocked from digestion, especially those grouped in the EA cluster. These EA phages show resistance to similar REs, suggesting that phages within the same cluster may display similar nucleobase substitutions. Further investigations are underway to identify modification similarities between phages within related or unrelated clusters. Biochemical analysis to reveal the presence and molecular structure of noncanonical nucleobase substitutions is a future goal of these investigations. Detailed knowledge of phage genome protective mechanisms is important, especially as it may contribute a competitive advantage to the phage when using in a therapeutic setting.

citation/acknowledgements.

Howard Hughes Medical Institute, Winthrop University Ronald E. McNair Post-Baccalaureate Program, SC INBRE grant 5P20GM103499 NIH/NIFMS/SC.

Generating a Gene Library of Bacteriophage Phayonce

Kyla Thomas, Matthew Defreitas, Jacqueline Gould, and Daniel C. Williams

Department of Biology, Coastal Carolina University, Conway, SC

background. Due to their toxicity for bacterial hosts, bacteriophages have an emerging importance in treating medically significant bacterial species. Generating a gene library of Phayonce, a bacteriophage that infects Mycobacterium smegmatis, allows for systematic analysis of individual gene function and possible cytotoxic effects on host cells.

goal of study. This work aims to generate a library that contains each of Phayonce’s 77 genes in an inducible expression vector. Individually cloned genes can then be introduced into M. smegmatis host cells for subsequent analysis of gene function.

methods and results. Using gene specific primers, each Phayonce gene was amplified by PCR. Amplified genes were then assembled into the pExTra plasmid, which contains a tetracycline inducible promoter to drive Phayonce gene expression. After assembly and transformation in to E. coli, colonies containing putative plasmids were analyzed by PCR to verify the insert. All 77 genes of Phayonce’s genome were successfully cloned into pExTra and subsequent sequencing established they are error-free.

conclusions. Generating a complete gene library of the bacteriophage Phayonce has allowed for all 77 expression vectors to be used in phenotypic analysis of toxicity of each Phayonce gene. Toxicity of Phayonce genes on host M. smegmatis may have medically significant implications on bacteriophage therapies for bacterial species that have shown to be resistant to treatment.

citation/acknowledgements.

This project was supported by grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health. Additional support was provided by the SEA-GENES project of the Howard Hughes Medical Institute.

The Effect of Fibronectin on Epithelial-Normal and Cancer-Cells using Electrical Impedance Sensing

Brooke Daniels, Jessica Holder, and Maddy Behravan Department of Biology, Chemistry, and Physics, Converse University, Spartanburg, SC

introduction. Investigation of the epithelial cell attachment and cells layer formation is a crucial step in understanding how they are affected by cancer cell invasions. This work reports the effects of fibronectin on cell membrane attachment and layer formation using an Electric Cell-Substrate Impedance Sensing (ECIS) system in real-time. The impedance data is correlated to the cell attachment, spreading, and layer formation.

goal of study. To examine the effects of fibronectin on normal epithelial cells, cancer cells, and cell layer formation using impedance measurement.

methods and results. Healthy human epithelial cells (HaCAT) and cancer cells (A431) were used for this investigation. Fibronectin was added at various dosages of 2.5, 5, and 15 ug/ml to the cells. An ECIS system (Applied BioPhysics) was used to measure the electrical impedance of the cells.

conclusions. For the normal cells (HaCAT) with additive fibronectin, the impedance was initially increased but that after hour 20 the levels returned to equilibrium. Overall, there was no significant effect observed in relation to the fibronectin addition to the normal cells (HaCAT) at the applied dosages. For cancer cells (A431), a concentration of 15 ug/mL of fibronectin, there was a greater resistance but a lower concentration had little to no effect. After a 40-hour time period, the resistance of cancer cells (A431) with the addition of 15 ug/mL fibronectin was decreased. These greater resistances and decreased resistances are attributed to an increased and decreased cell attachment.

citation/acknowledgements. We thank SC INBRE for supporting this research. This project is supported by a grant from the South Carolina INBRE funded by the National Institutes of Health National Institute of General Medical Sciences (P20GM103499).

Investigating Protein Interactions of Mycobacteriophage Cain Genes 2, 9, and 55

Dallas Nivens, Laela Walker and Victoria Frost

Department of Biology, Winthrop University, Rockhill, SC

introduction. Previous investigations by SEA-GENES students at Winthrop University revealed bacteriophage Cain genes 2, 9 and 55 inhibit host growth when overexpressed inside Cain’s host Mycobacterium smegmatis. This phenotypic result is due to a protein-protein interaction occurring between a singular phage gene product and a corresponding host protein(s).

hypothesis/goal of study.

Discovering these interactions is important to describe the phage gene functions of Cain 2 and 55, and confirm the gene function of Cain 9 which is predicted to be a MuF-like minor capsid protein.

methods and results. The bacterial 2-hybrid assay allows us to target and identify the host protein interactions with our phage gene product. Our most recent B2H assays identified a number of possible host protein interactions of Cain 2 and 9, which included several transcription regulators. Prior protein-protein interaction assays have confirmed Cain 55 strongly interacts with the NusA and GntR transcriptional regulators of M. smegmatis. Therefore, we have the goal of performing truncation assays with this gene to produce shorter protein variants in order to better understand how the protein product of Cain 55 interacts with these two transcriptional regulators.

conclusions. Cloning gene 55 in this way allows for individual testing of its terminal domains and comparison to homologous regions in other identified K cluster phage gene sequences. Learning more about how bacteriophage proteins interact with their host’s proteome can expose the possible functions of phage genes, such as Cain 2 and 9. Since 65% of phage genes have no predicted functions, this work is important for future use of phage and phage gene products in the medical, biotech and agricultural fields.

citation/acknowledgements.

This work funded by SC INBRE grant # 5P20GM103499.

Systematic Characterization of Mycobacteriophage Gene Function on Bacterial Cell Growth

Matthew Defreitas, Jackie Gould, Kyla Thomas, and Daniel C. Williams

Department of Biology, Coastal Carolina University, Conway, SC

introduction/background. Bacteriophages are ubiquitous viruses containing diverse genomes. Many phage genomes have been bioinformatically annotated, however many genes lack wet-bench functional characterization. Elucidating individual phage gene function on host growth and the phage-host protein interactions causing such phenotypes allows for the exploration of novel antibacterial therapies within the context of phage-host biology.

hypothesis/goal of study. Our goal is to systematically characterize the genome of the temperate mycobacterophage Phayonce through the investigation of individual gene expression on host cell growth and the phagehost protein interactions underscoring their cytotoxicity.

methods and results. A library of inducible expression vectors was generated that contains each of Phayonce’s 77 genes. Individual plasmids were transformed into the host cell Mycobacterium smegmatis. When plated and induced on selective media, the resulting host colony phenotype was observed for signs of cytotoxicity. A total of 29 genes were identified as cytotoxic. Of these genes, genes 41 and 64, which lack a characterized function, showed neartotal inhibition of colony formation. To identify host proteins that interact with these genes, we performed a bacterial twohybrid screen and isolated numerous host protein fragments of possible interaction partners.

conclusions. Phayonce’s genome contains multiple cytotoxic genes, many of which are poorly characterized. Given their effect on host colony formation, these cytotoxic genes serve as a reservoir of potential candidates to be exploited in antibacterial therapeutic development. Since novel genes 41 and 64 exhibit extreme cytotoxicity, they represent prime examples of genes that can be utilized in antibacterial therapy development. Isolated phage-host protein interaction candidates will be verified and subsequently sequenced to identify the host interaction partners that mediate cytotoxicity. Once identified, these interaction partners can also be exploited in antibacterial therapeutics.

citation/acknowledgements. This project was supported by grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health. Support has also been provided by the SEA-GENES project of the Howard Hughes Medical Institute.

Determining the Cytotoxic Effects of Phayonce Genes on Growth of Mycobacterium smegmatis

Jacqueline Gould, Matthew Defreitas, Kyla Thomas, and Daniel C. Williams Department of Biology, Coastal Carolina University, Conway, SC

introduction/background. Bacteriophages are viruses that infect bacteria. The result of bacteriophage infection is host cell and subsequent release of the phage copies. Because the population of bacteriophages is vast and diverse, they provide an untapped reservoir of genes that may have deleterious effects on bacterial cell function. Our work focuses on the genome of mycobacteriophage Phayonce, which contains 77 genes.

hypothesis/goal of study. This study investigates how individual Phayonce genes impacts growth of the host Mycobacterium smegmatis. Evaluating cytotoxicity of Phayonce genes will provide insight to phage biology and possibly identify phage candidates for antibacterial therapeutics.

methods and results. Using genes specific primers, each Phayonce gene was amplified by PCR and assembled into the inducible expression vector pExTra. Purified plasmids were transformed into M. smegematis using electroporation. Individual Phayonce genes were expressed by plating transformants on induction media and then analyzing them for growth defects. We categorized genes based on their impact on growth and found 38 that didn’t affect have an affect on cell growth, 11 genes that caused a slight reduction of cell growth, and 10 genes that severely inhibited colony formation and growth.

conclusion. This study found numerous Phayonce genes that reduced cell growth when expressed in M. smegmatis and many of these cytotoxic genes have not been characterized bioinformatically. Genes that inhibit bacterial cell colony formation are particularly interesting and may represent effective bactericidal agents. Future studies are aimed at characterizing the mechanism of cytotoxicity and identification of host proteins that interact with these genes.

citation/acknowledgements.

This presentation was supported in part by grant P20GM103499-20(SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health. Additional support provided by SEA-GENES and the Howard Hughes Medical Institute.

Characterization and Analysis of Mycobacteriophage Cain’s Genes

Laela Walker, Jessica Morgan and Victoria Frost Department of Biology, Winthrop University, Rockhill, SC

Mycobacteriophage Cain has been of interest to SEA students at Winthrop University for almost two years. The K6 sub-cluster phage contains 100 diverse genes, each of which have been individually cloned into the pExTra plasmid and amplified before transformation into its bacterial host Mycobacterium smegmatis. The cytotoxicity assay is then carried out to observe possible inhibition of host growth after inducing the expression of each phage gene. This phenotypic method reveals whether interactions are occurring between the phage gene product and its host’s proteome. As a result of our work, six additional genes have been demonstrated to exhibit these traits: 10, 19, 35, 43, 58, and 67 (a total of 17/73 tested to date), the majority of which have no predicted function. All pExTra plasmids were clone verified to check for the presence of the gene insert, then sequenced. The results were compared against each known Cain gene sequence using in silico NCBI sequence alignment tools. Following the analysis of these results, using SnapGene Viewer software, we can conclude that most of Cain’s genes match the anticipated gene sequence. This suggests that the phenotypic observations are a reflection of the effect of a specific phage gene’s expression. Our next steps, with these and Cain’s other cytotoxic genes, consist of performing bacterial-2hybrid assays to investigate the protein-protein interactions occurring between a phage and its host. Elucidating specific interactions at the phage-host interface may provide opportunities to exploit previously unknown proteins, or manipulate the outcome of the microbial coexistence.

Howard Hughes Medical Institute, Science Education Alliance Gene-function Exploration by a Network of Emerging Scientists, SC INBRE grant 5P20GM103499 NIH/NIGMS/SC.

Effects of Estrogen, Antiestrogen, and Polycyclic Aromatic Hydrocarbons (PAHs) on Human Breast Cancer Cells

Christian Ward and Samir Raychoudhury Department of Biology, Chemistry, and Environmental Health Science, Benedict College, Columbia, SC

introduction/background. Polycyclic aromatic hydrocarbons (PAHs) are known to be environmental pollutants, carcinogenic, mutagenic, and bioaccumulate in fatty tissue over time. The existence of PAHs in air pollution is an important public health issue. The purpose of the research project was to study the exposure induced cellular changes, and ZO-1 and laminin levels in human breast cancer MCF-7 cells following treatment with Benzo(a)Pyrene (BaP), Fluoranthene (FLA), 17β-estradiol (E2) and tamoxifen (Tx). ZO-1 is a tight junction (TJ) protein that is located on cytoplasmic membrane surface precisely at sites of cell-cell contact. Laminin is a basement membrane glycoprotein that plays a role in cell adhesion and differentiation.

hypothesis/goal of study. The research project aimed to study the correlation between ZO-1 and laminin following treatment with 17beta-estradiol, tamoxifen, and PAHs.

methods and results. We have planned to study cell membrane permeability by measuring cellular polarity in a two-chamber culture conditions. In general, one million MCF-7 cells were cultured in 35mm dishes, then exposed to medium containing 0.1% dimethyl sulfoxide (control solvent), 1 and 10µg/mL BaP, 1 and 10µg/mL FLA, 1 and 10µg/ mL E2, or 1 and 10µg/mL Tx. Following the 24hour exposure, the protein was extracted, and the enzyme-linked immunosorbent assay (ELISA) was performed. There were no relative changes in laminin or ZO-1 levels as determined by ELISA. However, Tx treatment reduced ZO-1 to undetectable levels. Further studies will require quantifying the amount of protein and validation of the absorbance data.

conclusions. The results suggest that the anti-estrogenic effect of tamoxifen may be mediated through reduced ZO-1 level.

citation/acknowledgements. This work was supported by SC INBRE.

The Analgesic Effects of Arnica montana Extracts on Post-Operative Pain

Reagan B. Turner1, Deltrice T. Holmes1, Theodore J. Price2, Wei Lei3 1Department of Biology, College of Art and Science, Presbyterian College, Clinton, SC, 2School of Behavioral and Brain Sciences, The University of Texas at Dallas, Richardson, TX, 3 College of Pharmacy, Natural and Health Sciences, Manchester University, Fort Wayne, IN

introduction/background. Arnica Montana (AM) is an herbaceous perennial plant that has been traditionally used in treating trauma, bruises, inflammation, or tissue injuries. However, the molecular mechanisms of Arnica’s medicinal properties are largely unknown.

hypothesis/goal of study. The objective of this study is to evaluate the effects of the Arnica extracts on post-operative pain in CD-1 mice.

methods and results. The extracts were made by mixing Arnica powder with one of the following solvents: 100% ethanol, ethanol: water (7:3, v/v), methanol, and acetone. The extracts were dried at room temperature and mixed in PCCA VersaBase gel at 1% by weight. The analgesic effect of Arnica extracts was tested in a post-operative pain model in CD-1 male and female mice. Mice were treated with gel alone or gel with 1% Arnica extract for one hour under anesthesia after the paw incision surgery. After recovered, the von Frey assay was performed at 3, 5, 24, and 48 hours after surgery. We found that all Arnica extracts exhibited analgesic effects to reduce the post-operative pain. The tissues around the incision were harvested for measuring the concentration of pro-inflammatory mediators, such as interleukin (IL)-1β, IL-6, and tumor necrosis factor alpha (TNF-α) using Western Blot and ELISA. We found that treatment with AM acetone extract reduced IL-1β, but not TNF-α concentration by ~50%.

conclusions. These findings further confirm the activity of Arnica on pain relief and provide a better understanding of the mechanism of the analgesic effect of Arnica.

citation/acknowledgements.

This work was funded by institutional funds from Presbyterian College and Manchester University, and by the South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE) Summer Research Intern Program.

The Impact of Taste Experience during Early Development on Later Taste and Neural Processing

Kaliana Reifsnyder and Veronica L. Flores Furman University, Greenville, SC

introduction. Experiences with taste can impact how future taste decisions are formed later in life. For example, in latent inhibition (LI), familiarity with a taste prevents the same taste from being associated with a negative consequence that would lead to an aversion. Here, we ask whether early life experiences can also cause LI-like effects later in life. Previous research has shown that taste and odor experiences during different stages of development can influence decisions made toward the same taste and odor later in life. The impact of early taste experience on later taste learning, independent from odor and fat to our knowledge, has not been examined.

methods. We address the question of how taste experiences, specifically sucrose, during gestation, lactation, or weaning impact later latent inhibition in aversion learning towards sucrose later in life in Long Evans Rats. We explore how early life sucrose experience targets taste preference and threshold formation by investigating how aversions toward sucrose generalize to different concentrations of sucrose, saccharin, and fructose. Data was collected using a gustameter to measure consumption and licking behavior.

hypothesis. Previous research leads us to the hypothesis that early life experience with sucrose will lead to latent inhibition of aversion learning later in life. This research is essential for a better understanding of how early life taste experience impacts taste decisions in adulthood which can provide insight into better treatments and preventative measures for consumption disorders that can result in obesity, depression, hypertension, and diabetes.

conclusions. Animals who had experience with sucrose during gestation and lactation are better able to learn about sucrose as adults. On the other hand, animals who had self-experience with sucrose as juveniles demonstrated mild latent inhibition to sucrose as adults. Furthermore, animals lacking taste experience generalized their aversions to other sweet tastes.

citation/acknowledgements. This project was supported by grant P20GM103499-20 from the South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE) from the National Institutes of Health National Institute of General Medical Sciences.

Meet DRP Recipient

Veronica Flores

(FY2022-2023)

Institution/Department: Furman University, Psychology

Project Title: The impact of innocuous taste experience on long-term taste-learning and memory persistence

Project Information: Humans and animals alike often learn to make food decisions by factoring in the consequences of their past eating experiences. However, my research has shown that even inconsequential taste experiences can impact future food decisions. In this project, we will use a rodent model of learning to shed light on how inconsequential taste experience impacts neural plasticity mechanisms that can ultimately lead to enhanced taste-related learning and memory.

How the SC INBRE grant funding will help accomplish your goal(s): SC INBRE funding allows my undergraduate students to earn hands-on research experience that will help prepare them for their future career goals. Furthermore, this award allows me to form collaborations with renowned scientists around the US and acquire the necessary equipment, supplies, and solutions to perform our behavioral experiments and analyze neural tissue.

Outside the Lab/Classroom: When not working, I like to hike with my husband and dog, travel, bake, play video games, and make espresso drinks.

Learn more about Dr. Flores at https://bit.ly/scinbre-spotlights

The Impact of Innocuous Taste Experience on LongTerm Taste Learning and Memory Persistence

Dallas Shuman, Annika Patterson, Parker Pack and Veronica Lee Flores Department of Psychology, Furman University, Greenville, SC

introduction/background. The evolutionary basis of learning and memory is influenced by an organism’s experience with sensory stimuli. For example, animals learn to avoid poisons after negative experience. However, even benign previous experiences can shape the way that new incidents with stimuli are perceived. The study of how experience influences future taste processing is important to consider, especially in laboratory taste research where naturalistic taste experiences are lacking, to best apply findings to the natural world. A previous study showed that rats that have had prior benign experience with tastes (TE-Taste Exposure) learned a stronger aversion to a novel taste (sucrose) 24 hours later compared to naive rats (Flores et. al, 2016) suggesting that taste experience enhances rats’ ability to learn about new tastes in short-term memory.

hypothesis/goal of study. The current study expands upon previous research and explores how prior experience with tastes affects taste learning in long-term memory through behavioral and immunohistochemistry analysis in Long Evans Rats. Because experience enhances learning, we expect TE rats to maintain a stronger aversion at long-term memory time points; 24 hours, 72 hours, 1 week, or 2 weeks postaversion. Since early immediate gene protein Npas4 is expressed when neuronal connections are strengthened, we expect higher levels of this protein in gustatory cortex in TE rats as a neurological indicator of stronger learning.

methods and results. Our results show that taste aversions weaken overtime, as expected. The experience-enhanced learning phenomenon seen before leads us to hypothesize that aversions post TE will be stronger and thus retained better over time.

conclusions. Humans have inconsequential taste experiences from the moment they are born, so examining the effects of these experiences on future taste learning will help form a better understanding of the neural mechanisms behind everyday consumption and appetite-related disorders.

citation/acknowledgements. This work was funded by the South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE) Program.

The Effects of Adenosine on Temporal Perception

Kelsy White1, Anna Joe Powell1,2, Nathaly Camargo1, Kelsie Glass1, Richard Keen2 , 1Neval Erturk

1Department of Biology, Chemistry, and Physics, Converse University, Spartanburg, SC, 2Department of Psychology, Converse University, Spartanburg, SC

introduction. Time perception plays a crucial role in many daily tasks. This cognitive process is used for walking, eating, and many forms of communication. Certain neuromodulators and neurotransmitters have been shown to alter this physiological process. Our lab previously tested and determined that caffeine speeds up temporal perception by acting as an adenosine antagonist. The reverse effect was the focus of our most recent experiment, which focused on adenosine and its potential to slow down time perception. Adenosine, an inhibitory neurotransmitter, plays an important role in depressing the nervous system and can increase the pressure for sleep. This study tested whether various doses of adenosine would alter time perception by slowing it down.

goal of study. To determine if adenosine slows down temporal perception, and if there is a dose-dependent relationship.

methods and results. Twenty four female Wistar rats were trained in an operant chamber using the Stubbs Procedure to discriminate between long (x > 4 seconds) and short intervals (x < 4 seconds). After this discrimination was learned, adenosine (2.5, 5, 7.5, or 10 mg/kg) or a sham control was injected intraperitoneally 15 minutes prior to beginning the timing procedure using a counterbalanced, within-subjects design. No statistically significant slowing down of temporal perception was observed although one condition did approach significance, t(23) = 1.323, p = 0.099. In addition, no effect was observed on overall accuracy after adenosine injection.

conclusions. No significant results were found with half of the planned data collected. We plan to finish the project this year to determine if adenosine slows down temporal perception.

citation/acknowledgements. This project is supported by a grant from the South Carolina INBRE funded by the National Institutes of Health National Institute of General Medical Sciences (P20GM103499).

Regulation of food intake: leptin and serotonin interactions

Daniel C. Giles, Nicholas D. Maxwell, DeWitt M. Coleman, Victoria M. Glass, James R. Fadel, Claudia A. Grillo Department of Pharmacology, Physiology and Neuroscience, University of South Carolina School of Medicine, Columbia, SC, WJB Dorn VA Medical Center, Columbia, SC

In 1998, the World Health Organization first deemed global obesity as an “epidemic”. As of 2019, the epidemic affected more than 2 billion people, roughly 30% of the world’s population. One contributing factor of obesity is overeating which has been found to positively associate with weight gain. Appetite is controlled by a myriad of neurochemical signals, with two key players being leptin and serotonin (5-HT). Leptin—an adipocyte-derived hormone— acts on hypothalamic neurons, but its impact has been shown to extend to other areas of the brain, such as the dorsal raphe nucleus (DRN), which is the primary site of 5-HT synthesis. In our laboratory, we have characterized the anatomical connection between leptin sensitive neurons in the DRN and the arcuate nucleus (ARC) of the hypothalamus. Further, we have shown that 5-HT affects leptin’s ability to regulate food intake and are continuing to explore the signaling mechanism.

The goal of our current study was to test the hypothesis that DRN neurons that respond to leptin can release 5-HT in the ARC which contributes to decreased food intake.

To accomplish this goal, we performed stereotaxic surgery and implanted two cannulas in the brain: one in the ARC and one in the DRN. In male Sprague-Dawley rats, we performed in-vivo micro dialysis and injected leptin or vehicle into the DRN and collected dialysate samples from the ARC. Once complete, we analyzed the dialysates via high performance liquid chromatography (HPLC) with electrochemical detection to measure 5-HT concentration. We found leptin injections in the DRN stimulates a triphasic release of 5-HT into the ARC.

This study demonstrated that activation of the DRN leptin receptors stimulates the 5-HT efflux in the ARC defining a novel circuit to inhibit food intake. Further, we suggest that this extra-hypothalamic pathway may be a future target for controlling appetite.

citation/acknowledgements. This work was supported by the University of South Carolina SPARC Award (NDM), grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health (NDM), and the National Science Foundation grant number IOS-1656626 (CAG).

Conformational Analysis of μ-Opioid Receptor Agonists

Leah Juechter, Brenna Outten, Lauren Jones, and George Shields Department of Chemistry, Furman University, Greenville, SC

The abuse of prescription and illicit opioids is a major health crisis as it is the leading cause of death by overdose in Europe and North America. When an opioid agonist binds to the μ-opioid receptor (MOR), it elicits both positive and adverse effects. In order to develop novel pain treatments, we need to understand how opioids bind to the MOR. We aim to use computational methods to gain insight into the unbound conformations of opioid ligands in solution. We considered the hypothesis that ligands have their lowest free energy conformations already arranged prior to being bound to the MOR such that they will bind tightly. A set of rough geometries were generated using a PM7 semi-empirical genetic algorithm (GA) that is implemented in CREST. The geometries were optimized at the ωB97XD density functional with the 6-31++G** basis set, and electronic energies of the final set of lowenergy conformations were calculated at the DLPNO-CCSD(T)-cc-pVnZ//wB97XD/6-31++G**/ SMD (N= D, T, Q) level of theory at 298 K and 310 K in order to assess the biological viability of these conformers. All calculations were computed using the SMD solvation node to produce structures that are optimized in aqueous solution. The ligands we analyzed in this study are BU72, morphine, and fentanyl. Results showed little conformational variation in structures within 1 kcal/mol of the global minima for BU72 and morphine. For fentanyl, the hydrophobic phenol rings tend to cluster together across all structures, but there was a greater amount of conformational diversity overall. These data show that the ligands are changing to adopt their lowest energy conformation in solution based on their intramolecular forces and steric interactions with water. Future research includes docking these ligands in the MOR using a program called Autodock to evaluate any conformational and energetic changes within the binding pocket.

citation/acknowledgements.

This presentation was supported in part by grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health and the Furman University Department of Chemistry.

Blocking the Expression of PTSD in a Female Rodent Model

Alex O’Neal, Kirsten Folger, Jason Hayden and Onarae Rice Department of Psychology, Furman University, Greenville, SC

introduction. Posttraumatic stress disorder (PTSD) is a psychological disorder that develops in response to experiencing one or more traumatic events. Symptoms of the condition include re-experiencing events, heightened startle response, negative emotional states, sleep disturbance, increased anxiety, and avoidance behaviors. Past studies have suggested that the dopamine D3 receptor plays a critical role when it comes to stress-induced behaviors. Additional studies have indicated that the D3 antagonist SB-277011A can be used to block the expression of PTSD-like symptoms in male rats. However, males and females experience the effects of PTSD differently.

hypothesis. We were interested in investigating the effects of a D3 antagonist in a female rodent model. We hypothesized that female mice exposed to the D3 antagonist SB-277011A would express lessened PTSD-like behaviors when compared to mice who did not receive the antagonist.

methods and results. Our experiment consisted of 50 female mice and utilized a modified single prolonged stress (mSPS) model of PTSD. Mice were subjected to baseline behavioral testing and were then put through the mSPS procedure, which consisted of a 20-minute forced swim, 2-hour physical restraint, and a 20-minute electric foot shock session. After nine days, mice were injected with SB-277011A and were then put through a second round of behavioral testing. Throughout behavioral testing and mSPS procedures, a tone and scent were used as trigger stimuli. Additionally, vaginal lavages were taken before behavior testing and mSPS to gather estrous cycle phases. Our data was inconclusive, but results from female mice subjected to SB-277011A was different when compared to male rats.

conclusion. PTSD is a complex mental disorder that affects males and females differently. D3 antagonists have been shown to affect PTSD-like behaviors in rodents. Continued work in researching the effects of D3 antagonists on both males and females is vital to understanding and even preventing PTSD.

citation/acknowledgements.

Research

reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number P20GM103499. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Over-expression of ABC Transporters by Reflux Medication Induces Drug Resistance and Promotes Cancer Growth in Esophageal Tumor Cells

Mayah Lee and Christopher Farrell School of Pharmacy, Presbyterian College, Clinton, SC

background. The high mortality rate of gastro-intestinal cancers, such as esophageal cancer, has been linked to patients' resistance to chemotherapy. Past literature suggests that medications that are proton-pump inhibitors (PPis) — used to alleviate symptoms of chemotherapy and gastrointestinal disorders — correlate to poorer prognoses in cancer. The administration of PPis can disrupt the absorption of chemotherapy in cancer cells and has been linked to causing multidrug resistance (MDR). ABC transporters are responsible for transporting drugs out of the cell; the change in drug response may be correlated to the administration of PPis, such as omeprazole, an ABC transporter substrate.

hypothesis. Our objective was to identify an association of chemo-naïve cells treated with PPis as having higher levels of expression for ABC transporters. Methods: The PPi studied, omeprazole, was selected due to its frequent prescription in gastrointestinal disorders, specifically Gastro-Esophageal Reflux Disease (GERD) and Barrett's Esophagus (BE). Multiple gene expression analyses were conducted in a colorectal cell line, Caco2, via PCR (Polymerase Chain Reaction) and cytotoxicity assays. The colorectal cell line was used as a baseline for the expression of ABC transporters.

results. Our results exhibited a correlation of higher expression levels of these transporters in cells that were treated with omeprazole. Changes in

cells expressing drug-resistance were verified in esophageal cell lines, OE19 and OE33. Caco2 cell lines from the experimental group showed higher levels of drug resistance as compared to the untreated group.

conclusions. Cell lines with induced expression of ABC transporters showed a lower drug response to chemotherapy, as compared to the control.

citation/acknowledgements. This work was funded by institutional funds to Presbyterian College from SC INBRE grant P20GM103499.

PH-02

Modeling COVID-19 in Spartanburg County, SC with a Susceptible-Vaccinated-Infected-RecoveredDeceased (SVIRD) Model

D. Chloe Griffin1 and Amanda Mangum2 1Appled Mathematics, Brown University, Providence, RI, 2Department of Mathematics and Computer Science, Converse University, Spartanburg, SC

introduction. Epidemic mechanistic models are governed by biological principles of disease spread. Standard Susceptible-InfectedRecovered (SIR) and Susceptible-Exposed-Infected-Recovered (SEIR) compartmental models are too simplistic to capture the behavior of the COVID-19 pandemic. Therefore, a model with time-dependent parameters is needed to model the behavior of this pandemic.

goal of study. In our work, we seek to capture the overall behavior and multiple peaks of the various COVID-19 waves that occurred in Spartanburg Country, SC, as seen in the data gathered from USA Facts and SC DHEC.

methods and results. After smoothing the data corresponding to COVID-19 positive-test counts, hospitalization, death, and vaccination counts by computing a moving average, we fit parameters to the experimental data using the MATLAB fmincon function. We solve the system of differential equations created by the Susceptible-Vaccinated-Infected-Recovered-Deceased (SVIRD) model using ODE45. We explore the results of the fit SVIRD model that also incorporates time-dependent parameters, a level of caution factor in response to increased infection in the population, and a sense of safety factor for increased vaccinations.

conclusions. The model provides a good fit for the data in Spartanburg County. The addition of the timedependent parameters in the model gives insight into the changing social response toward the COVID-19 pandemic within Spartanburg County. The model illuminates what COVID-19 levels may have been at the beginning of the pandemic before testing became widely available. The model and the fitted parameters may also help provide forecasting for the behavior of future waves now that testing rates have declined.

citation/acknowledgements. This project is supported by a grant from the South Carolina INBRE funded by the National Institutes of Health National Institute of General Medical Sciences (P20GM103499).

South Carolina Research Experiences for Teachers (RET) Program Summer 2023

The SC RET program, with funding provided by National Institutues of Health-funded South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE) and National Science Foundation-funded South Carolina Established Program to Stimulate Competitve Research (SC EPSCoR), enables teachers to engage in a six-week immensive research experiences at an institution in their local area. These experiences, the first for many of South Carolina’s teachers, expose teachers to modern research methods and allow them to link their research activities to classroom activities designed to increase student knowledge and awareness of science. Up to 14 teachers per year are directed in carrying out unique, individualized research projects resulting in scientific presentation at the SC INBRE Science Symposium and/or SC EPSCoR Annual Meeting.

This program represents a full time commitment (35 to 40 hours per week) and provides financial support ($3,000 each) to participants, as well as a supplies budget (up to $500) to translate summer experiences into deliverables during the participant’s academic year instruction.

It is the intention that the efforts of the RET program in training teachers will strengthen the pipeline of South Carolina K-12 students to become ready to engage in biomedical and advanced materials research.

ED PD 662: Research Experiences for STEM Teachers (optional): Teachers selected for the research experience may register for 4-6 credit hours of recertification/professional development credit. Course serves to connect summer research experience with tangible teaching artifacts to bring back into the classroom. Course credit is offered through the Furman University Office of Graduate Studies.

RET is managed through SC INBRE network institution, Furman University. To learn more about Summer 2023 opportunities, contact Dr. John Kaup, SC RET Coordinator (Furman University), (864) 294-3773, john.kaup@furman.edu or visit online at

https://bit.ly/SC-RET

GLV effects on herbivory by Tobacco Hornworms & Beet Armyworms on Tomato Plants

Lakshmi Sunitha Arava1, Harshita Negi2, Johannes Stratmann2

1Fairfield Central High School, Winnsboro, SC, 2Department of Biology, University of South Carolina, Columbia, SC

introduction/background. Plants are under constant stress as they are being attacked by microorganisms and insects. Plants communicate with their environment by chemical language. This means when the plants are wounded by insects (herbivores), they release volatile danger signals that neighboring plants can ‘smell.’ These Volatiles include green leaf volatiles (GLVs), such as the small alcohol hexenol. GLVs trigger “priming”-, a state of alertness that enables the receiver plant to forcefully respond to the attack of herbivores. (Ref: Int. J. Mol. Sci. 2013, 14, 17781-17811; doi:10.3390/ijms140917781)

hypothesis/goal of study. We hypothesize that tomato plants that were treated with a GLV are more resistant to subsequent attack by caterpillars as compared to plants that were not treated with the GLV.

methods and results. Beet Armyworm caterpillars-(Spodoptera exigua) were used as the herbivore, a small tomato cultivar (Micro-Tom) as the plant, and Z-3-hexenol as the GLV. Armyworm eggs were cultured on artificial food. Five to eight weeks-old tomato plants were placed in airtight containers. Four were treated (priming) with 74 microliters of pure Z-3-hexenol, and four others were left untreated. One day after the GLV treatment, five armyworms of equal weight were placed onto each tomato plant on the 10th day of hatching.

Subsequently, caterpillar weight was measured every day until pupation. The weight of the caterpillars was measured from day 9 to 12. Activity is repeated with Tobacco hornworms (Manduca sexta). The only difference compared to Activity 1 was that only one caterpillar was dropped on each tomato plant. The caterpillars were weighed from day 10 to day 17.

conclusions. We found that the weight of both caterpillar species grown on GLV-treated plants is slightly higher than for caterpillars grown on untreated plants. This indicates that our hypothesis was rejected by the experiments. In the future, we can vary the concentration of the GLV or test another GLV for priming activity.

citation/acknowledgements. INBRE: This project was supported by grant P20GM103499-20 (SC INBRE) from the National Institute of General Medical Sciences, National Institutes of Health. EPSCoR: National Science Foundation (NSF) EPSCoR Program under NSF Award # OIA-1655740.

I thank Dr. Johannes Stratmann, and Harshita Negi for helping me explore “GLV effects on herbivory.” I thank Dr. John Kaup, Director of Science Education (Furman University), to present this research opportunity via the RET program.

References:

1. Int. J. Mol. Sci. 2013, 14, 17781-17811; doi:10.3390/ijms140917781

2. South-Carolina-college-and-career-ready-science-standards-2021

Evaluation of the Aegilops tauschii chromosome introgression lines in common wheat for reducedgluten content

Habibunnisa Begum1, Zachary T. Jones2 and Sachin Rustgi2

1Marion High School, Marion, SC, 2Department of Plant and Environmental Sciences, Clemson University, Clemson, SC

Wheat is an essential nutritional source worldwide, serving as a staple food for 35% of the global population. While wheat is a crucial protein and carbohydrate source for most, 1.4% of the world’s population suffers from a wheatassociated auto-immune disorder known as celiac disease, for which gluten consumption causes damage to the villi of the small intestine resulting in poor nutrient uptake. At this time, the only effective treatment for this condition is total avoidance, which can prove difficult due to unintended contamination of daily-use products. The D-genome of wheat has been found to contain the highest number of epitopes responsible for celiac disease. To identify reduced gluten wheat, a D-nested association mapping (DNAM) population was screened for its gluten content and composition. The DNAM population is a unique resource where the D-subgenome of the common wheat was replaced with eight Aegilops tauschii donors. The DNAM population is currently being analyzed with an expectation to identify lines completely lacking or accumulating the reduced quantity of immunogenic gluten proteins. Additionally, we intend to decipher the genetic regulation of immunogenic gluten protein accumulation in wheat grains via protein quantity loci mapping using this population. Information from these experiments will be used to further breeding programs to develop reduced immunogenic wheat varieties for celiac patients. The techniques learned during this research experience will be utilized to enhance students’ curriculum and experiential learning.

citation/acknowledgements. We acknowledge the financial support from the NIH INBRE RET Program and USDA National Institute of Food and Agriculture (GRANT13715753).

Exploring the LightInduced Transformation of Photoswitchable Compound in a High School Chemistry Classroom

Asia Shaik1, Gina Wilson2 and Natalia Shustova2 1Laurens District 55 High School, Laurens, SC, 2Department of Chemistry and Biochemistry, University of South Carolina, Columbia, SC

introduction/background. Photochromism is a unique property exhibited by molecules to alternate between two discrete states in response to light stimulation. Spiropyran is a type of photochromic material that undergoes a structural change that results in a shift in its photo-absorption and is being developed for various applications such as optical switches, bioadhesive mediators, and functional links. This technology is commonly used in ski-googles, motorcycle shields, and helmet wearing people.

hypothesis/goal of study. The goal of the research is to provide high school chemistry students with a practical understanding of photochromism and its applications. By conducting this simple experiment students can observe the transformation of a colorless solution to a different color upon exposure to light. This hands-on experience helps students to understand versality of these compounds for sun protection and other applications to make the study of chemistry more meaningful and relevant.

methods and results. Students can perform a simple experiment to observe the photochromic properties of Spiropyran by dissolving it in dimethylformamide and applying the solution to a plain white material such a paper, cotton, or synthetic material. The material will appear purple when exposed to UV light or sunlight. This experiment provides hands-on experience in observing the reversible change in the optical properties of Spiropyran in response to light exposure and can help students understand the concepts of photochromism.

conclusions. Overall,

the study of photochromism offers a valuable opportunity for students to explore the real world applications of chemistry and see the impact that scientific research can have on society. Through hands-on experimentation with Spiropyran, students can gain a deeper appreciation of this field and understand the relevance and significance of their studies.

citation/acknowledgements. National Science Foundation South Carolina EPSCoR Program under NSF Award #OIA-1655740.

Bioinformatics Pilot Project Program (BIPP)

Up to 5 awards of $10,000 each

BIOMEDICAL RESEARCH FUNDING OPPORTUNITIES 2022

Developmental Research Project Program (DRP)

Up to 6 awards for PUI faculty; Up to 1 for Clemson, 1 for UofSC faculty; $50,000 each

Student-Initiated Research Projects Program (SIRP)

Up to 5 awards of $3,000 each

SC INBRE announces $695,000 in funding awarded to FY2022 Project Program Grant Recipients

CALL FOR APPLICATIONS

The South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE), a statewide network of institutions of higher education funded by the NIH National Institute of General Medical Sciences, has an open Call for Applications for three grants programs for undergraduate students through faculty. The BIPP Program is designed to stimulate the application of genomics and bioinformatics methods by supporting research and student training through the SC INBRE and SCTR networks, as well as at Primarily Undergraduate Institutions across South Carolina. The DRP Program is designed to foster faculty research programs at SC INBRE network institutions. SIRP Program recipients apply bioinformatics tools to biomedical research questions. See our website for eligibility and more details.

South Carolina IDeA Networks of Biomedical Research Excellence (SC INBRE), a statewide $18.9 million, five-year renewable grant program funded by The National Institutes of Health (NIH) National Institute of General Medical Sciences (NIGMS), is pleased to announce our FY2022-2023 Project Program grants recipients.

FOA DATES

NEW THIS YEAR – LETTER OF INTENT required. Due 5 pm, March 4, 2022

Deadline for Submission: 5 pm ET, April 4, 2022

Period of Performance: September 1, 2022 through August 31, 2023

The 2022 BIPP Program recipients are:

• Linnea Freeman, Furman University, Biology, Fecal microbiome evaluation in a mouse model of autism

• Chakia McClendon, Columbia College, Biology, Kinome Expression Profiling of Airway Cells Exposed to Electronic and Conventional Cigarettes

SCINBRE

The Bioinformatics Pilot Project Program (BIPP) is designed to stimulate the application of genomics and bioinformatics methods by supporting research and student training through the SC INBRE and South Carolina Clinical & Translational Research (SCTR) networks. BIPP proposals focus on Biomedical Science and fit within the broad scientific focus areas of SC INBRE: Biochemistry/Cell and Molecular Biology, Bioinformatics, Bio/Biomedical Engineering and Regenerative Medicine, and Neuroscience. The program supports independent, faculty-driven research and provides research training to students and/or postdoctoral fellows in Bioinformatics. Each project awards up to $10,000 for one year.

scinbre.org/funding

• Kandy Velazquez, University of South Carolina School of Medicine Columbia, Pathology, Microbiology & Immunology, Development of pain biomarkers to assess analgesia and predict pain in pre-clinical models of cancer pain

The Developmental Resesarch Project Program (DRP) is designed to foster faculty research programs at SC INBRE network institutions. DRP projects must focus on Biomedical Science and fit within the broad scientific focus areas of SC INBRE: Biochemistry/Cell and Molecular Biology, Bioinformatics, Bio/ Biomedical Engineering and Regenerative Medicine, and Neuroscience. The program awards support independent

research programs and, in the process, provide research training to students and/or postdoctoral fellows in the Biomedical Sciences. Each project awards up to $50,000 for one year, renewable for a second and possible third year upon favorable review of the progress report by SC INBRE’ s External Advisory Committee.

The 2022 DRP Program recipients are:

• Timothy Barker (NEW), College of Charleston, Chemistry/Biochemistry, Synthesis and Biological Evaluation of 5-HT antagonists

• Meghan Breen (2nd YEAR), Furman University, Chemistry, Effects of Pdr1 phosphosites on azole resistance in Candida glabrata

• Heather Dunn (NEW), Clemson University, Bioengineering, Novel Investigations of Breast Cancer Racial Disparities based on Deep Learning and Bioinformatics Analysis

• Mindy Engevik (NEW), Medical University of South Carolina, Regenerative Medicine & Cell Biology, Microbial suppression of intestinal inflammation and oxidative stress

• Timea Fernandez (NEW), Winthrop University, Chemistry, Using Antibiotic-Binding Nucleic Acid Aptamers as Trojan-Horse Delivery Vehicles in the Fight Against Drug-Resistant Bacteria

• Veronica Flores (NEW), Furman University, Psychology, The impact of innocuous taste experience on long-term taste-learning and memory persistence

• Meredith Frazier (NEW), College of Charleston, Chemistry/Biochemistry, Structural and Functional Characterization of Viral Ribonucleases

• Chiara Gamberi (NEW), Coastal Carolina University Biology, Modeling renal cyst mechanisms in the fruit fly Drosophila melanogaster

• Jordon Gilmore (2nd YEAR), Clemson University, Bioengineering, Point-of-Care Colorimetric Biosensor for Detection of Pseudomonas aeruginosa Signaling Molecule 3OC12HSL in Wound Exudate from Negative Pressure Wound Therapy

• Anita Nag (3rd YEAR), USC Upstate, Chemistry, Connecting host shutoff by SARS coronavirus nsp1 to pre-mRNA processing and mRNA trafficking

• Subramanya Pandruvada (2nd YEAR), Medical University of South Carolina, Oral Health Sciences, Modeling macrophage response in periodontal infections

• Daniel Stovall (2nd YEAR), Winthrop University, Biology, Role and Regulation of Ring1 and YY1 binding protein in Glioblastoma Multiforme

• Kristy Welshhans (NEW), University of South Carolina, Biomedical Sciences, Molecular and cellular basis of altered neural development in Down syndrome

In addition to the two grants listed above which are focused on supporting faculty research, SC INBRE also offers a student-focused opportunity entitled Student-Initiated Resesarch Project Program (SIRP). The SIRP program is designed to enrich the academic experience of students and to better prepare the future generation of researchers, investigators, and entrepreneurs throughout the SC INBRE network. SIRP recipients apply bioinformatics tools to biomedical research questions that fit within the broad scientific focus areas of SC INBRE: Biochemistry/Cell and Molecular Biology, Bioinformatics, Bio/Biomedical Engineering and Regenerative Medicine, and Neuroscience. SIRP awards support bioinformatics training for undergraduate or graduate students. Each project awards up to $3,000 for one year.

The 2022 SIRP Program recipients are:

• Patrice Cunningham, graduate student (Dr. Kandy Velazquez, mentor), USC School of Medicine

Columbia, Pathology, Microbiology & Immunology, Identification of quercetin targets in skeletal muscle as a novel treatment of cancer cachexia

• Ethan Older, graduate student (Dr. Jie Li, mentor), University of South Carolina, Chemistry and Biochemistry, Discovery of selective inhibitors of proinflammatory sulfonolipids produced by human gut microbiota

• Mark Pitman, graduate student (Dr. Jessica Larsen, mentor), Clemson University, Chemical and Biomolecular Engineering, Applying a Citrate Biomaterial to Reduce Ferroptosis in Spinal Cord Injury

• Alia Sadek, medical student (Dr. Jennifer Grier, mentor), USC School of Medicine Greenville, Biomedical Sciences, Identifying Mechanisms of Resistance in Novel Acinetobacter baumannii Necrotizing Fasciitis Isolates

• Mihyun Waugh, graduate student (Dr. Melissa Moss, mentor), University of South Carolina, Biomedical Engineering, Blood Cytokine Levels and Healthcare Data as Predictors of Mesh Exposure After Pelvic Organ Prolapse Surgery