The 11th Annual DSAC Student Research Symposium Friday ...biomed.emory.edu/news-events/documents/2014-DSAC... · Session II: Immunity to Infection 10:30 am 10:30 Maria Georgieva (MMG)

THE ELEVENTH ANNUAL DSAC STUDENT RESEARCH SYMPOSIUM [1]

The 11th Annual DSAC Student Research Symposium Friday, January 10

Woodruff Health Sciences Center Administration Building (WHSCAB) Auditorium

8:30 – 9:00 Breakfast – WSHCAB Lobby

Session I: Microbial Systems 9:00 am

9:00 Phillip Zakas (MSP) Reduced Antigenicity of Ovine Factor VIII in Human Hemophilia A Inhibitor Plasmas

9:15 Jennifer Frediani (NHS) Plasma High-Resolution Metabolomic Profiling Reveals Upregulation of Specific Resolvins in Patients with Pulmonary Tuberculosis

9:30 Mary Bushman (PBEE) Within-Host Competition of Malaria Parasites in Humans and the Fitness Cost of Drug Resistance

9:45 Brigid O’Flaherty (MMG) Viral Rta and Cellular IRF4 Synergistically Regulate MHV68 M1 Gene Expression in Plasma Cells

10:00 Emily Kuiper (BCDB) Interrogating RNA Structural Dynamics Upon Substrate Recognition by the Thiostrepton-Resistance Methyltransferase

10:15 – 10:30 Break

Session II: Immunity to Infection 10:30 am

10:30 Maria Georgieva (MMG) Mycobacterium tuberculosis Hip1 is a Serine Protease that Cleaves GroEL2 and Modulates Macrophage Responses

10:45 David Pinelli (IMP) Rapamycin Augments the CD8 T Cell Response to a Latent Viral Infection in the Presence or Absence of CTLA4-Ig

11:00 Victor Band (IMP) Recognition of B. anthracis Cell Wall Polysaccharide by the Host Innate Immune System

11:15 Justin Kandler (MMG) Neisseria gonorrhoeae Infection of Macrophages Impacts the Host Iron-Limiting Innate Immune Defense and Upregulates Gonococcal Iron-Acquisition Genes


Session III: Translational Models 11:30 am

11:30 James Cordova (MSP) Quantitative, Unbiased Tumor Segmentation for Evaluation of Extent of Brain Tumor Resection to Facilitate Multisite Clinical Trials

11:45 James Burkett (NS) Neural Mechanisms of Empathy-based Behavior in Monogamous Prairie Voles

12:00 – 1:30 Poster Sessions & Lunch– WHSCAB Lobby

Poster Session 12:00 – 12:45pm Odd Numbered Poster Presentations 12:45 – 1:30pm Even Numbered Poster Presentations

1:30 – 1:45 Break

Session IV: Life of a Cell 1:45 pm

1:45 Chantel Cadwell (BCDB) Cadherin Endocytosis, Adhesion, and Cytoskeletal Linkage Cooperatively Regulate Cell Migration

2:00 Jason Conage-Pough (CB) Phosphorylation Influences the Preferential Binding of Bim to Anti-Apoptotic Proteins in Multiple Myeloma Session V: Receptors and Signaling 2:15 pm

2:15 Gina Alesi (CB) RSK2-Mediated Phosphorylation of Stathmin Promotes Microtubule Polymerization and Provides a Pro-invasive Advantage to Metastatic Cancer Cells

2:30 Jessica Konen (CB) LKB1 Regulates a Molecular Switch that Drives #-D Lung Cancer Invasion

2:45 Ariana Mullin (NS) NSF Regulates Synaptic Homeostasis at the Drosophila Neuromuscular Junction

3:00 George Kannarkat (IMP) The rs3129882 Single Nucleotide Polymorphism Affects MHC-ii Expression and May Increase Risk for Parkinson’s Disease by Heightening Immune Responses

3:15 – 3:30 Break

Session VI : DNA and Gene Expression 3:30 pm

3:30 Will Hudson (MSP) The Structural Basis of Direct Glucocorticoid-Mediated Transrepression

3:45 Paul Musille (MSP) Divergent Sequence Tunes Ligand Sensitivity in Phospholipid-Regulated Hormone Receptors


4:00 Kevin Van Bortle (BCDB) A Subclass of Insulator Binding Sites Establish Dense Clusters to Delineate Topologically Associating Domains

4:15 Mariana Mandler (BCDB) A Cytoplasmic Quaking Isoform Regulates an hnrnpf/h-dependent Alternative Splicing Pathway in Myelinating Glia

4:30-Leila Myrick (NS) Pathogenesis of Novel FMR1 Mutations in Fragile X Syndrome

4:45-Jordan Morreall (GMB) TNF-α Inhibits DNA Repair Activity of BER Variant OGG1 Through Oxidative Stress

5:00-5:15pm Break

Keynote Address

5:15-5:45 pm

Adam M. Katz Adam M. Katz is the Policy and Advocacy Specialist at Research!America, where he leads a variety of advocacy initiatives to make science and medical research a higher national priority. Prior to joining Research!America, Adam held positions at the University of Rochester, the University of California at Irvine and the National Institute of Mental Health. He holds a masters degree in biomedical science policy and advocacy from Georgetown University and undergraduate degrees in brain and cognitive sciences as well as clinical and social psychology from the University of Rochester.

5:45pm Reception and Awards Presentation – WHSCAB Lobby

Images

Front Cover: Image Contest Winners 1st Place: Todd Deveau (NS) Page 5 2nd Place: Ana Montiero (BCDB) Page 24 3rd Place: Amanda York (BCDB) Page 58

All images were judged by the Integrated Cellular Imaging center




Session I

Microbial Systems

9:00 am


Phillip Zakas, MSP 9:00 am

REDUCED ANTIGENICITY OF OVINE FACTOR VIII IN HUMAN HEMOPHILIA A INHIBITOR PLASMAS

Philip M. Zakas1, Kristine Vanijcharoenkarn2, Rebecca Markovitz3, Shannon Meeks, MD4 and Christopher B. Doering, PhD41Graduate Program in Molecular and Systems Pharmacology, Emory University, Atlanta, GA, 2Emory University School of Medicine, MD candidate, 3Emory University School of Medicine, MD PhD candidate, 4Aflac Cancer and Blood Disorders Center, Emory University/Children’s Healthcare of Atlanta, Atlanta, GA Neutralizing antibodies (inhibitors) develop in approximately 30% of severe hemophilia A patients and present the greatest obstacle to the treatment of patients with access to care. Currently, a recombinant porcine factor VIII (pFVIII) product (OBI-1™, Baxter International) is undergoing phase III clinical trials due to reduced cross-reactivity of anti-human FVIII (hFVIII) inhibitors to pFVIII. Herein, we investigate the antigenicity of orthologous ovine FVIII (oFVIII), and its potential utility for inhibitor patients. Analysis of 38 hemophilia A inhibitor patient plasmas revealed significantly reduced inhibition of oFVIII coagulant function compared to hFVIII, with some patients showing lower titers against oFVIII than pFVIII. Inhibitors primarily target the functionally essential A2 and C2 domains of FVIII. Characterization of A2 and C2 inhibitors within patient plasmas by competition ELISA with anti-hFVIII murine monoclonals revealed a high prevalence of potent inhibitors overlapping with known epitopes. Using an indirect ELISA, we demonstrate that these inhibitors do not cross-react with oFVIII with the exception of specific C2 domain targeting inhibitors. Therefore, we hypothesize that non-conserved residues within oFVIII alter the clinically important surface epitopes and thereby diminish cross-reactivity. These data demonstrate that oFVIII could have potential as a third line treatment in the FVIII inhibitor patient setting.

Jennifer Frediani, NHS 9:15 am

PLASMA HIGH-RESOLUTION METABOLOMIC PROFILING REVEALS UPREGULATION OF SPECIFIC RESOLVINS IN PATIENTS WITH PULMONARY TUBERCULOSIS Jennifer K. Frediani1, Dean P. Jones1, Nestan Tukvadze2, Eka Sanikidze2, Maia Kipiani2, Karan Uppal1, Shaheen S. Kurani1,

Gautam Hebbar1, Romain A. Colas3, Jesmond Dalli3, Charles N. Serhan3, Vin Tangpricha1, Henry M. Blumberg1, Thomas R. Ziegler1 1Emory University, Atlanta, GA; 2National Center for Tuberculosis and Lung Disease, Tbilisi, Georgia; 3Harvard University, Boston, MA Background: Characterization of metabolites altered by tuberculosis (TB) disease may identify pathophysiologic pathways. Resolvins are lipid mediators derived from endogenous EPA and DHA critical for resolution of inflammation. Methods: Plasma samples from 17 patients (35±12 yrs old) with new pulmonary TB and 17 matched household contacts without TB (controls) were analyzed using high-resolution LC-MS. False discovery rate (FDR; q=0.05) and hierarchal cluster analysis was used to distinguish accurate mass ions (metabolites) significantly different between groups. The METLIN Metabolite Database was used to match ions to known metabolites. Identification of specific D-series resolvins (RvD1 and RvD2) was confirmed by LC-UV-MS/MS. Results: Over 23,000 metabolites were detected in untargeted metabolomic analysis, of which 61 were significantly different between the two groups by FDR. Eight metabolite clusters were evident containing several metabolites quantitatively upregulated in patients with TB (e.g. glutamate, choline derivatives, anti-TB drug metabolites and several lipids, including likely Mtb cell wall lipids and resolvins RvD1 and RvD2. Conclusions: This pilot study revealed metabolites that are upregulated during newly diagnosed TB potentially involved in pathogenesis. Upregulation of specific resolvins may mediate recovery from inflammation in TB disease.


Mary Bushman, PBEE 9:30 am

WITHIN-HOST COMPETITION OF MALARIA PARASITES IN HUMANS AND THE FITNESS COST OF DRUG RESISTANCE Mary Bushman1, Nancy O. Duah2, Neils Quashie2, Benjamin Abuaku2, Kwadwo A. Koram2, Jacobus de Roode1, Venkatachalam Udhayakumar3

1Emory University, Atlanta, GA, United States, 2Noguchi Memorial Institute for Medical Research, University of Ghana, Legon, Ghana, 3Centers for Disease Control and Prevention, Atlanta, GA, United States Most infections with the malaria parasite Plasmodium falciparum consist of multiple strains, which presumably occupy the same ecological niche; therefore, we expect within-host competition to occur in mixed-strain infections. Competition may suppress transmission of drug-resistant strains, which could significantly slow evolution and/or accelerate decline of resistance following retirement of a failing drug. To test the hypothesis that within-host competition occurs in mixed infections, we used quantitative real-time PCR to determine parasite density of two strains of P. falciparum (one chloroquine-sensitive, one chloroquine-resistant) in 1,423 blood samples from three African countries. We found that total parasite density did not differ between single-strain and mixed-strain infections, while each strain exhibited lower densities in mixed infections that those achieved in single-strain infections. These findings were replicated in all three countries, and suggest that competition does indeed occur in mixed-strain infections. Further investigation also revealed a significant fitness cost of resistance to chloroquine. First, chloroquine-resistant parasites attained lower densities than sensitive parasites in both single and mixed infections. Second, in one country, the frequency of resistance dropped from 78% to 43% over six years following the retirement of chloroquine. These findings constitute a significant advance in our understanding of the evolution of drug-resistant malaria.

Brigid O’Flaherty, MMG 9:45 am

VIRAL RTA AND CELLULAR IRF4 SYNERGISTICALLY REGULATE MHV68 M1 GENE EXPRESSION IN PLASMA CELLS

Brigid M. O’Flaherty1,2, Tanushree Soni1,2, Samuel H. Speck1,2 1Department of Microbiology and Immunology, 2Emory Vaccine Center, Emory University School of Medicine Atlanta, Georgia 30322 MHV68 is a murid gammaherpesvirus that infects laboratory mice and provides a small animal model for gammaherpesvirus biology and pathogenesis. Co-evolution with their hosts has resulted in acquisition of virus specific unique genes important in modulating infection. One such gene, M1, unique to MHV68, encodes a secreted protein with no known homolog. Previously M1 was identified as the stimulatory ligand responsible for expansion of cytolytic IFN/TNFα secreting Vβ4+ CD8+ T cells following MHV68 infection. In the absence of M1, Vβ4+ T cell expansion fails to occur and viral reactivation is poorly controlled. However, the mechanism of action and regulation of M1 are poorly understood. Utilizing a recombinant virus expressing M1 promoter driven YFP, we identified of splenic plasma cells as the predominant site of M1 expression. We extended these findings to characterize the transcriptional regulators of the M1 promoter, identifying cellular interferon regulatory factor 4 (IRF) and viral replication and transcription activator (Rta) as synergistic partners which regulate M1 expression through protein-protein interaction. These findings shed light on the function and mechanism of M1 mediated Vβ4+ CD8+ T cell expansion, and suggests a conserved means of transcriptional regulation for MHV68 genes linked to viral reactivation and plasma cell differentiation.


Emily Kuiper, BCDB 10:00 am

INTERROGATING RNA STRUCTURAL DYNAMICS UPON SUBSTRATE RECOGNITION BY THE THIOSTREPTON-RESISTANCE METHYLTRANSFERASE Emily G. Kuiper1,2, Jennifer Hernandez3,4, Graeme L. Conn1. 1Department of Biochemistry, 2Biochemistry, Cell and Developmental Biology (BCDB) Program, 3SURE Program, Emory University School of Medicine, Atlanta, GA, 4Georgia State University, Atlanta, GA. As bacteria become increasingly resistant to antibiotics in the clinic, it is critical to understand the molecular mechanisms of resistance in order to slow its spread or develop new antimicrobial therapies. Antibiotic-producing bacteria are a reservoir for new resistance determinants that could appear in the clinic, as they must contain mechanisms to protect themselves from their toxic products. In the thiostrepton producer Streptomyces azureus, the thiostrepton-resistance methyltransferase (Tsr) methylates the drug binding site on the 23S ribosomal RNA thereby inhibiting its binding and action against the bacterial ribosome. Resistance by this mechanism is common in drug producers and is now becoming increasingly prevalent in the clinic, e.g. providing resistance to macrolide and aminoglycoside antibiotics. To understand how antibiotic-resistance conferring enzymes recognize their RNA substrates we have biochemically dissected Tsr’s substrate recognition mechanism. Using RNA structure probing, binding and enzymatic experiments we show that the carboxyl-terminal domain (CTD) of Tsr docks on the RNA, then engages its amino-terminal domain (NTD) to bind and unfold the RNA tertiary structure in a critical step necessary prior to catalysis. These findings provide new insights into the intricacies of substrate recognition for antibiotic resistance conferring and other RNA modifying enzymes that recognize the bacterial ribosome.


Session II

Immunity to Infection

10:30 am


Maria Georgieva, MMG 10:30 am

MYCOBACTERIUM TUBERCULOSIS HIP1 IS A SERINE PROTEASE THAT CLEAVES GROEL2 AND MODULATES MACROPHAGE RESPONSES Maria Georgieva1,*, Jacqueline L. Naffin-Olivos2,*, Nathan Goldfarb3, Ranjna Madan-Lala1, Dagmar Ringe2, Ben M. Dunn3, Gregory A. Petsko2 and Jyothi Rengarajan1, 4 1Emory Vaccine Center, Emory University, Atlanta, GA, USA, 2Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA, USA, 3Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, FL, USA, 4Division of Infectious Diseases, Department of Medicine, Emory University, Atlanta, GA, USA *These authors contributed equally Mycobacterium tuberculosis (Mtb) employs multiple strategies to evade host immune responses. We have previously shown that the cell envelope-associated Mtb serine hydrolase, Hip1, dampens macrophage responses. We now provide key mechanistic insights into the molecular and biochemical basis of Hip1 function. We establish that Hip1 is a serine protease and show that the Mtb GroEL2 protein is a substrate of Hip1 protease activity. Cleavage of GroEL2 occurs between Arg12 and Gly13 and is optimal at intraphagosomal pH conditions. Interestingly, we discovered that Hip1-mediated cleavage of GroEL2 converts the protein from a multimeric to a monomeric form. Moreover, ectopic expression of cleaved GroEL2 monomers into the hip1 mutant restored wild type levels of cytokine responses in infected macrophages. Thus Hip1-dependent proteolysis is a novel regulatory mechanism that helps Mtb respond rapidly to immune environments during infection. These findings position Hip1 as a target for developing immunomodulatory therapeutics against Mtb. David Pinelli, IMP 10:45 am

RAPAMYCIN AUGMENTS THE CD8 T CELL RESPONSE TO A LATENT VIRAL INFECTION IN THE PRESENCE OR ABSENCE OF CTLA4-IG David F Pinelli1, Maylene E Wagener1, Mandy L Ford1

1Department of Surgery, Emory University School of Medicine

Latent viral infections are a major concern among immunosuppressed transplant patients. Treatment with belatacept, a CTLA4-Ig fusion protein, is contraindicated in EBV- patients, due to an increased risk of PTLD associated with primary EBV infection. Recent reports have shown that treatment with rapamycin (rapa) paradoxically enhances CD8 T cell responses to certain pathogen infections, raising the possibility that rapa could be used to enhance responses to EBV in belatacept-treated patients. We infected mice with gammaherpesvirus 68 (gHV), the murine homolog of EBV, and treated with rapa +/- CTLA-4 Ig. Rapa treatment alone increased the number of CD8 T cells response to gHV at Day 20 compared to mice that received no treatment, and also led to an increase in IFN-γ production. While CTLA4-Ig treatment alone dampened the T cell response to gHV, the addition of rapamycin was able to restore the response to baseline levels by day 20 post-infection. In addition, all rapamycin treated animals showed increased levels of CD127 on virus-specific cells, indicating that rapa may generate higher quality memory T cells regardless of treatment with CTLA4-Ig. These results suggest that rapamycin may be able to be used in combination with CTLA4-Ig in order to augment protective immune responses.


Victor Band, IMP 11:00 am

RECOGNITION OF B.ANTHRACIS CELL WALL POLYSACCHARIDE BY THE HOST INNATE IMMUNE SYSTEM Victor Band1,4, Nazia Kamal2, Elke Saile2, Scott Forsberg3, Conrad Quinn2, Geert-Jans Boons3, David Weiss4 1Emory University Immunology and Molecular Pathogenesis, 2Centers for Disease Control and Prevention, 3University of Georgia Carbohydrate Research Center, 4Emory Vaccine Center B. anthracis is a Gram positive bacterium that is the causative agent of the acute disease anthrax. It contains a protein capsule that protects it from antimicrobials and is linked to a unique cell wall polysaccharide. We show that this cell wall polysaccharide is a novel activator of the immune system, acting as a pathogen associated molecular pattern (PAMP). Using macrophages from knockout mice lacking various innate immune receptors, we determined that this polysaccharide is sensed by Toll like receptor 4 (TLR4), which also detects Gram negative lipopolysaccharides. After characterizing the structures of the cell wall polysaccharides of various B. anthracis strains, we determined that this polysaccharide is modified by galactose additions in virulent strains, which we show reduces TLR4 activation. Further, we discovered that an avirulent strain whose basis for attenuation is not known, does not have these galactose additions and is much more inflammatory in vitro. These data suggest that modification of this cell wall polysaccharide may be critical to the ability of B. anthracis to avoid immune detection and to cause disease, with important implications for the treatment of anthrax and our understanding of host-pathogen interactions.

Justin Kandler, MMG 11:15 am

NEISSERIA GONORRHOEAE INFECTION OF MACROPHAGES IMPACTS THE HOST IRON-LIMITING INNATE IMMUNE DEFENSE AND UPREGULATES GONOCOCCAL IRON-ACQUISITION GENES Justin L. Kandler1, Susu M. Zughaier1, and William M. Shafer1,2 1Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA 30322, 2Laboratories of Microbial Pathogenesis, VA Medical Center, Decatur, GA 30033 Neisseria gonorrhoeae (the gonococcus) is the causative agent of the disease termed gonorrhea and a strict human pathogen. Though many mechanisms of gonococcal iron-piracy are well-established, little is known about how intracellular gonococci acquire sufficient iron within the iron-limited host to sustain viability during infection. Due to the crucial role of macrophages in host iron homeostasis, we investigated the impact of gonococcal infection of macrophages on host and gonococcal iron-related genes. We found that phagocytosis of wild-type gonococci by mouse and human macrophages increased expression of host genes encoding hepcidin, NRAMP1, and NGAL, but decreased expression of host genes encoding ferroportin and BDH2. These changes were associated with an increased level of labile (free) iron within host cells. Analysis of gonococcal transcripts revealed that expression of iron-responsive genes (including iron-acquisition genes) was increased during intracellular residence within macrophages. Furthermore, approximately 15% of the live, internalized gonococci remaining after 1 hour survived the next 4 hours of the phagocytic killing process. Taken together, these data suggest that gonococci modulate the host iron-limiting innate immune defense and upregulate their own iron-acquisition genes to survive intracellularly. We conclude that this mechanism contributes to gonococcal survival in the infected host.


Session III

Translational Models

11:30 am


James Cordova, MSP 11:30 am

QUANTITATIVE, UNBIASED TUMOR SEGMENTATION FOR EVALUATION OF EXTENT OF BRAIN TUMOR RESECTION TO FACILITATE MULTISITE CLINICAL TRIALS James S. Cordova 1,2, Eduard Schreibmann3, Constantinos G. Hadjipanayis4, Ying Guo,5 Tim Fox3 , Hui-Kuo Shu3,6, Hyunsuk Shim1,6,*, and Chad A. Holder1,* 1Dept. of Radiology, 2Medical Scientist Training Program, 3Dept. of Radiation Oncology, 4Dept. of Neurosurgery, 5Dept. of Biostatistics, 6Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA 30322 RATIONALE: Generating tumor volume measurements is a difficult component of neurosurgical extent-of-resection (EOR) assessment. We utilized data from an ongoing trial to develop a semi-automated method for the generation of T1-weighted (T1W) contrast-enhancing tumor volumes pre- and postoperatively. EXPERIMENTAL DESIGN: T1W MR images from glioblastoma cases and controls were segmented using Otsu multi-level thresholding and Fuzzy C-means clustering for comparison to a specialist’s manual contours. Agreement and inter-rater variability between segmentations was assessed using the concordance correlation coefficient (CCC), and spatial accuracy was determined using the Dice similarity index (Dice) and Hausdorff distance (HD). RESULTS: Fuzzy C-means clustering with three classes (Fuzzy3) was the best method, generating volumes with high agreement to manual contours pre- and postoperatively (CCCs = 0.996 and 0.986, respectively) and between readers pre- and postoperatively (CCCs = 0.990 and 0.983, respectively). Spatial agreement with manual contours was high preoperatively for Fuzzy3 with mean Dice and HDs of 0.916 and 0.142 mm, respectively; and postoperatively, Fuzzy3 exhibited the highest spatial agreement of all algorithms. CONCLUSION: The proposed segmentation method allows tumor volume measurements of magnetic resonance images in the unbiased, reproducible fashion necessary for quantifying EOR in multicenter clinical trials. James Burkett, NS 11:45 am

NEURAL MECHANISMS OF EMPATHY-BASED BEHAVIOR IN MONOGAMOUS PRAIRIE VOLES James Burkett1,2, Daniel Curry1,2, Melissa Reyes1,2, Larry Young1,2

1Emory University, Atlanta, GA, 2Center for Translational Social Neuroscience.

Consolation is a common human expression of sympathy for another’s pain. Consoling behavior has also been naturally observed in several animal species, including some great apes, canids, corvids, and elephants. However, no laboratory model has been established to study this behavior. Here I present the first experimental data showing that male prairie voles (Microtus ochrogaster) express consoling behavior under laboratory conditions. Males significantly increase their partner-directed allogrooming toward stressed female partners, as compared to baseline and to unstressed controls. Promiscuous meadow voles do not show this stress-evoked increase. Prairie males who observe their stressed partners show an increase in self-grooming and an elevation of plasma corticosterone that is highly correlated to the corticosterone of their female partner, suggesting that consolation is based on an empathy mechanism. Finally, injection of an oxytocin receptor antagonist (OTA) into the cerebral ventricle prevents the stress-evoked increase in allogrooming. We are also exploring the role of OTA in specific brain regions in modulating consolation. These experiments are the beginning of a foundation for an animal model that will inform us about the neurobiology of consolation and empathy.


Session IV

Life of a Cell

1:45 pm


Chantel Cadwell, BCDB 1:45 pm

CADHERIN ENDOCYTOSIS, ADHESION, AND CYTOSKELETAL LINKAGE COOPERATIVELY REGULATE CELL MIGRATION Chantel M. Cadwell1, Benjamin A. Nanes1, and Andrew P. Kowalczyk1,2,3

Departments of Cell Biology1 and Dermatology2, and the 3Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA

Endothelial cell adhesion is dynamically regulated during angiogenesis, a process important for development. Angiogenesis occurs by migration and proliferation of endothelial cells, activities that require the modulation of adhesion. The adherens junction protein VE-cadherin mediates adhesion through homophilic interactions via a conserved tryptophan (Trp2) in its extracellular domain. VE-cadherin is linked to the cytoskeleton through cytoplasmic interactions with catenins. These interactions stabilize the cadherin at the cell surface and increase adhesive strength. Previously, we determined that mutation of specific amino acids in the VE-cadherin cytoplasmic tail prevents endocytosis of the cadherin and inhibits cell migration. Interestingly, we now report that the inhibition of migration by this mutant is dependent upon both adhesion and cytoskeletal linkage. Mutation of the critical Trp2 residue disrupted adhesion and increased VE-cadherin endocytosis relative to wild type cadherin. Additionally, we observed that forced lateral dimerization of VE-cadherin prevented endocytosis even in the absence of homophilic adhesion. Importantly, dimerized VE-cadherin also inhibited migration in an adhesion dependent manner. Lastly, inhibition of VE-cadherin endocytosis disrupts Golgi reorientation, a process that is important for directed cell migration. These findings suggest that endocytosis, adhesion, and cytoskeletal linkages of VE-cadherin cooperate to control endothelial cell migration by regulating endothelial cell polarity.

Jason Conage-Pough, CB 2:00 pm

PHOSPHORYLATION INFLUENCES THE PREFERENTIAL BINDING OF BIM TO ANTI-APOPTOTIC PROTEINS IN MULTIPLE MYELOMA Jason E Conage-Pough1,2,3, Vikas A Gupta, MD PhD1,3, Shannon M Matulis, PhD1,3 and Lawrence H Boise, PhD1,2,3,4 1Winship Cancer Institute, 2Graduate Program in Cancer Biology, 3Department of Hematology and Medical Oncology, 4Department of Cell Biology, Emory University School of Medicine, Atlanta GA

Myeloma cells are strongly dependent on the anti-apoptotic protein Mcl-1 for survival. However, studies have shown that some myeloma cells are sensitive to treatment with drugs that target Bcl-2 and Bcl-xL, but not Mcl-1. These studies suggest that sensitive cells have more of the pro-apoptotic protein Bim bound to Bcl-2/Bcl-xL than to Mcl-1. We have previously shown that the expression levels of pro- and anti-apoptotic proteins have little influence on which anti-apoptotic protein binds Bim, leading us to further explore what factors influence the preferential binding of Bim.

We determined that Bim is constitutively phosphorylated in unstimulated myeloma cells at multiple unknown sites. We also found that the phosphorylation status of Bim affects its binding to anti-apoptotic proteins, with phosphorylated species preferentially binding to Bcl-2/Bcl-xL, and not to Mcl-1. We observed that the phosphorylated Bim species bound to Bcl-2 and Bcl-xL were present at a significantly lower level in myeloma cells that were more dependent on Mcl-1, than in cells with greater Bcl-2/Bcl-xL dependency. Lastly, we showed that inhibitors targeting the MAP kinase family affect the levels of constitutive Bim phosphorylation, providing a potential avenue for influencing Bim distribution through targeting of kinases and phosphatases, and improving therapeutic efficacy.


Session V

Receptors and Signaling

2:15 pm


Gina Alesi, CB 2:15pm

RSK2-MEDIATED PHOSPHORYLATION OF STATHMIN PROMOTES MICROTUBULE POLYMERIZATION AND PROVIDES A PRO-INVASIVE ADVANTAGE TO METASTATIC CANCER CELLS Gina Alesi,1 Dan Li,1 Lingtao Jin,1 Georgia Z. Chen,1 Dong M. Shin,1 Fadlo Khuri,1 and Sumin Kang1 1Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA 30322

Protein kinases have been implicated in mediating pro-metastatic signaling in human cancers. We previously demonstrated that a serine/threonine kinase, p90 ribosomal S6 kinase 2 (RSK2) promotes cancer cell invasion and migration, anoikis resistance, and tumor metastasis in human cancers. We investigated the role of RSK2 in the regulation of microtubule dynamics and its potential implication in cancer cell invasion and tumor metastasis. SiRNA-mediated knockdown of RSK2 disrupted microtubule formation in diverse metastatic human cancer cell lines. Stathmin (STMN) is a microtubule destabilizing protein that regulates microtubule dynamics. We found that stable knockdown of RSK2 decreased phosphorylation of STMN at Serine 16. Moreover, RSK2 directly phosphorylated STMN at Ser16 in an in vitro RSK2 kinase assay. We observed that phosphorylation of STMN by RSK2 reduced STMN-mediated microtubule depolymerization in vitro, while stable expression of phospho-deficient STMN S16A but not STMN wild type led to decreased invasive potential of metastatic cancer cells. These data suggest STMN as a potential novel RSK2 substrate/effector, which may contribute to RSK2-mediated pro-metastatic signaling. Therefore, the RSK2-STMN pathway represents a promising therapeutic target in the clinical prognosis and treatment of metastatic human cancers.

Jessica Konen, CB 2:30 pm

LKB1 REGULATES A MOLECULAR SWITCH THAT DRIVES 3-D LUNG CANCER INVASION Jessica Konen1, Adam I. Marcus2 1Graduate Program in Cancer Biology, Emory University, Atlanta, GA, 2Hematology and Medical Oncology, Emory University, Atlanta, GA LKB1 is a tumor suppressor and serine/threonine kinase that is the third-most commonly mutated gene in NSCLC. Additionally, a KrasG12DLkb1fl/fl knockout mouse had increased tumor burden and metastasis when compared to other clinically-relevant mutant mice. Together, these data solidify LKB1 as an important regulator of lung cancer metastasis; however, the mechanism by which LKB1 mutation results in increased metastasis is still unknown. To dissect how LKB1 loss increases the metastatic potential of cancer cells, we utilized a lung cancer cell spheroid model to analyze invasion into a 3-D microenvironment in real-time. We found shLKB1 cells switch their invasive phenotype, performing a mesenchymal to amoeboid transition (MAT). Quantification of live-cell imaging shows shLKB1 amoeboid cells have longer tracks and significantly greater velocity than LKB1 wildtype amoeboid cells. Because RhoA, and its effector kinase ROCK, have been implicated in regulating amoeboid motility, we examined the effects of inhibiting myosin II (downstream of RhoA/ROCK signaling) on 3-D motility in control and shLKB1 spheroids. We found inhibition of myosin II inhibits amoeboid morphology and results in a significant decrease in shLKB1 amoeboid cell velocity. This work shows LKB1 loss results in MAT during invasion, potentially providing LKB1-depleted cells a migratory advantage through the microenvironment.


Ariana Mullin, NS 2:45 pm

NSF REGULATES SYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NEUROMUSCULAR JUNCTION Ariana P. Mullin1,2 Avanti Gokhale1, Subrabhata Sanyal3, Victor Faundez1

1Department of Cell Biology, Emory University School of Medicine, Atlanta, GA, USA, 2Graduate Program in Neuroscience, Emory University, Atlanta, GA, USA 3Biogen-Idec, 14 Cambridge Center, Cambridge, MA, USA Synaptic homeostasis describes the ability of the presynaptic cell to adapt to changes in the postsynaptic response. During this phenomenon, the presynaptic cell integrates feedback from the postsynaptic cell to regulate neurotransmitter release properties, thus maintaining appropriate postsynaptic excitation. This type of synaptic plasticity is crucial during neural development, as synaptic contacts are particularly dynamic. Bloc-1 (Biogenesis of lysosome-related organelles complex 1) is an octameric complex involved in synaptic membrane protein trafficking, but the role of Bloc-1 in neurotransmission is unclear. Recently, however, it was shown that loss of function dysbindin, one of eight proteins that comprise Bloc-1, results in deficits in synaptic homeostasis at the Drosophila neuromuscular junction (NMJ). Dysbindin has been identified by multiple independent GWAS analyses as a schizophrenia-susceptibility factor, and loss of function in dysbindin is consistently implicated as a risk factor across multiple neurodevelopmental disorders. Here, we identify a novel role for N-ethylmaleimide sensitive factor (NSF) in synaptic homeostasis at the NMJ. NSF biochemically and genetically interacts with the Bloc-1 complex to regulate synaptic homeostasis. Our results suggest that interactions between protein complexes within a biochemically defined network regulate complex neural phenotypes, such as synaptic homeostasis, necessary for proper neuronal development.

George Kannarkat, IMP 3:00pm

THE RS3129882 SINGLE NUCLEOTIDE POLYMORPHISM AFFECTS MHC-II EXPRESSION AND MAY INCREASE RISK FOR PARKINSON’S DISEASE BY HEIGHTENING IMMUNE RESPONSES George T. Kannarkat1, Darcie A. Cook1, Jianjun Chang1, Jaegwon Chung1, Jae-Kyung Lee1, Elaine M. Sperin2, Stewart Factor2, Jeremy M. Boss3, and Malu G. Tansey1. 1Department of Physiology, 2Emory Movement Disorders Clinic, and 3Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA The non-coding rs3129882 HLA-DRA gene single nucleotide polymorphism (SNP) was associated with altered risk for Parkinson’s disease (PD) in genome-wide association studies and is part of the major histocompatibility complex-II (MHC-II) locus that mediates antigen presentation. People homozygous for the G (high risk) allele at this locus have a 1.7 fold higher risk for Parkinson’s disease. This association suggests that alteration in antigen presentation is a potential mechanism to link environment and genetics in conferring disease susceptibility. We have demonstrated that this SNP is linked to specific expression patterns of MHC-II molecules in B cells and interferon-�-stimulated monocytes from healthy controls and PD patients. These findings suggest that the SNP is an immune biomarker that may predict susceptibility for PD based on antigen presentation responses and engagement of adaptive immunity. Furthermore, we observed a positive correlation between MHC-II expression on B cells and disease severity but not disease duration suggesting a link to pathogenesis. This novel finding is the first direct evidence in humans that antigen presentation via MHC molecules plays a role in sporadic PD risk.


Session VI

DNA and Gene Expression

3:30 pm


Will Hudson, MSP 3:30 pm

THE STRUCTURAL BASIS OF DIRECT GLUCOCORTICOID-MEDIATED TRANSREPRESSION William H Hudson1, Christine Youn1, Eric A Ortlund1 1Emory University School of Medicine, Department of Biochemistry A newly discovered negative glucocorticoid response element (nGRE) mediates DNA-dependent transrepression by the glucocorticoid receptor (GR) across the genome and has a major role in immunosuppressive therapy. The nGRE differs dramatically from activating response elements, and the mechanism driving GR binding and transrepression is unknown. To unravel the mechanism of nGRE-mediated transrepression by the GR, we characterized the interaction between GR and an nGRE in the thymic stromal lymphopoietin (TSLP) promoter. We show using structural and mechanistic approaches that nGRE binding is a new mode of sequence recognition by human GR and that nGREs prevent receptor dimerization through a unique GR-binding orientation and strong negative cooperativity, ensuring the presence of monomeric GR at repressive elements.

Paul Musille, MSP 3:45pm

DIVERGENT SEQUENCE TUNES LIGAND SENSITIVITY IN PHOSPHOLIPID-REGULATED HORMONE RECEPTORS Paul M. Musille1, Manish Pathak1, Janelle L. Lauer2, Patrick R. Griffin2, and Eric A. Ortlund1 1 Emory University School of Medicine, Atlanta, GA 30322, USA 2 The Scripps Research Institute, Jupiter, Florida, USA.

The NR5A subfamily family of nuclear receptors are important regulators of pluripotency, lipid and

glucose homeostasis, and steroidogenesis. Liver receptor homologue 1 (LRH-1) and steroidogenic factor 1 (SF-1), have therapeutic potential for the treatment of metabolic and neoplastic disease; however, a poor understanding of their ligand regulation has hampered the pursuit of these proteins as pharmaceutical targets. In this study, we dissect how sequence variation among LRH-1 orthologs affects phospholipid (PL) binding and regulation. Both human and mouse LRH-1 (mLRH-1) respond to newly discovered medium chain PL agonists to modulate lipid and glucose homeostasis. These PLs activate human LRH-1 by altering receptor dynamics in a newly identified alternate activation function region. Mouse and Drosophila orthologs contain divergent sequence in this region potentially altering PL-driven activation. Structural evidence suggests that these sequence differences in mLRH-1 and Drosophila FTZ-f1 confer at least partial ligand independence, making them poor models for human LRH-1 studies; however, the mechanisms of ligand independence remain untested. We show using structural and biochemical methods that the recent evolutionary divergence of the mLRH-1 stabilizes the active conformation in the absence of ligand, yet does not abrogate PL-dependent activation. We also show by mass spectrometry and biochemical assays that FTZ-f1 is indeed a ligand independent receptor. This work provides a structural mechanism for the differential tuning of PL-sensitivity in NR5A orthologs and supports the use of mice as viable therapeutic models for LRH-1-dependent diseases.


Kevin Van Bortle, BCDB 4:00 pm

A SUBCLASS OF INSULATOR BINDING SITES ESTABLISH DENSE CLUSTERS TO DELINEATE TOPOLOGICALLY ASSOCIATING DOMAINS Kevin Van Bortle1, Mike Nichols, Li Li1,2, Chintong Ong1, Naomi Takenaka1, Zhaohui S. Qin2, Victor G. Corces1. 1Department of Biology, Emory University, Atlanta, GA 30322, USA, 2Department of Biostatistics and Bioinformatics, Emory University, Atlanta GA 30322, USA Chromatin insulators represent a unique class of regulatory elements proposed to play a role in chromosome domain organization. Recent advances in 3C – based assays have led to novel insight into the three dimensional arrangements of chromatin, and suggest that genomes are organized into tissue-invariant sub-megabase (Mb) scale structures call Topologically Associating Domains (TADs). Insulator proteins are enriched at the boundaries of TADs, yet a majority of insulator proteins localize within domains, suggesting boundary-associated insulator elements represent a unique subclass. By mapping the genome-wide binding profiles for several insulator factors in D. melanogaster, we demonstrate that insulator proteins establish dense clusters analogous but distinct from HOT regions, called Architectural Cluster Elements (ACEs). ACEs associate with the SMC-containing cohesin (as shown for HOT regions) and condensin complexes, suggesting these factors may be necessary for the co-occupancy of several proteins. ACEs correlate strongly with topological domain boundaries as well as the strength of domain interaction separation, and represent open chromatin regions that are stably accessible throughout development, consistent with the tissue invariant nature of TADs in humans. Finally, we uncover a similar relationship between insulator occupancy and TADs in humans, suggesting a conserved role for clustered architectural proteins in sub-Mb scale chromatin organization. Mariana Mandler, BCDB 4:15 pm

A CYTOPLASMIC QUAKING I ISOFORM REGULATES AN HNRNPF/H-DEPENDENT ALTERNATIVE SPLICING PATHWAY IN MYELINATING GLIA Mariana Mandler1, Li Ku1, Yue Feng1 1Department of Pharmacology, Emory University The selective RNA-binding protein Quaking I (QKI) plays important roles in controlling alternative splicing (AS). Three QKI isoforms are broadly expressed, which display distinct nuclear-cytoplasmic distribution. However, molecular mechanisms by which QKI isoforms control AS, especially in distinct cell types, still remain elusive. The quakingviable (qkv) mutant mice carry deficiency of all QKI isoforms in oligodendrocytes (OLs) and Schwann cells (SWCs), the myelinating glia of central and peripheral nervous system (CNS and PNS), respectively, resulting in severe dysregulation of AS. We found that the cytoplasmic isoform QKI-6 regulates AS of polyguanine (G-run)-containing transcripts in OLs and rescues aberrant AS in the qkv mutant by repressing expression of two canonical splicing factors, heterogeneous ribonuclearproteins (hnRNPs) F and H. Moreover, we identified a broad spectrum of in vivo functional hnRNPF/H targets in OLs that contain conserved exons flanked by G-runs; many of which are dysregulated in the qkv mutant. Surprisingly, AS targets of QKI-6 and hnRNPF/H in OLs are differentially affected in SWCs, suggesting that additional cell-type-specific factors modulate AS during CNS and PNS myelination. Together, our studies provide the first evidence that cytoplasmic QKI-6 acts upstream of hnRNPF/H, forming a novel pathway that controls AS in myelinating glia.


Leila Myrick, NS 4:30 pm

PATHOGENESIS OF NOVEL FMR1 MUTATIONS IN FRAGILE X SYNDROME Leila Myrick1, Mika Nakamoto-Kinoshita1, Xuekun Li1, Jeannie Visootsak1,2, Noralane Lindor4, Salman Kirmani5, Peng Jin1, Xiaodong Chen3, Stephen Warren1,2,3 1Department of Human Genetics, Emory University, Atlanta, GA, 2Department of Pediatrics, Emory University, Atlanta, GA, 3Department of Biochemistry, Emory University, Atlanta, GA, 4Department of Medical Genetics, Mayo Clinic, Rochester, MN, 5Department of Health Science Research, Mayo Clinic, Scottsdale, AZ Fragile X Syndrome (FXS) is the most common form of inherited intellectual disability (ID) and a leading known cause of autism spectrum disorder. FXS is most often caused by a trinucleotide repeat expansion within the FMR1 gene. In more than the two decades since FMR1 was discovered, only a single pathological missense mutation has been reported (I304N). Recently we identified 2 novel variants, G266E and R138Q, in males with ID who tested negative for repeat expansion. To determine if these variants are pathological, we used lentivirus to infect Fmr1 KO cells with either G266E-FMRP or R138Q-FMRP. We found that G266E behaves like a functional null and is unable to rescue AMPAR trafficking, associate with polyribosomes, or bind mRNA. Crystallographic modeling of G266E also indicates considerable steric structural disruption. Conversely, R138Q-FMRP was able to rescue all these phenotypes. However, when neuronally expressed in transgenic dfmr1-deficient Drosophila, R138Q failed to rescue synaptic overgrowth at the neuromuscular junction, suggesting this mutant impairs presynaptic function while retaining its postsynaptic function. In conclusion, G266E is a pathological null mutation, identified in a patient with classic FXS phenotype. R138Q retains postsynaptic FMRP function but may exhibit presynaptic specific defects. These data demonstrate both the investigational and clinical utility of screening for conventional FMR1 mutations in individuals with ID. Jordan Morreall 4:45 pm

TNF-α INHIBITS DNA REPAIR ACTIVITY OF BER VARIANT OGG1 THROUGH OXIDATIVE STRESS Jordan Morreall1,2, Clayton Sheppard4, Erica Werner1, Yoke Wah Kow4, Paul W. Doetsch1,3,4,5 1Department of Biochemistry, 2Graduate Program in Genetics and Molecular Biology, 3Emory Winship Cancer Institute, and 4Department of Radiation Oncology and 5Hematology and Medical Oncology, Emory University School of Medicine, Atlanta, GA

8-oxoguanine (8OG) is a common, mutagenic, oxidative DNA lesion. 8OG is repaired by the base excision repair (BER) pathway, initiated by 8-oxoguanine glycosylase 1 (Ogg1). A common Ogg1 allelic variant is S326C-Ogg1, associated with cancer, and inhibited by oxidative stress. Oxidative stress arises during inflammation, for which a key mediator is tumor necrosis factor alpha (TNF-α), a cytokine driving oxidative stress and contributing to all stages of cancer. However, inflammatory cytokines’ role in modulating BER glycosylase variant phenotypes is poorly understood. We hypothesized that TNF-α drives oxidative stress causing DNA damage and inhibits S326C-Ogg1 glycosylase activity through cysteine 326 oxidation. To measure Ogg1 activity, we incubated whole-cell lysates with 32P-labeled 8OG oligonucleotides, quantifying intact vs. cleaved oligonucleotide fragments. In vitro, S326C-Ogg1 activity is inhibited by oxidative stress caused by treatment with H2O2 or TNF-α, but not after treatment with the antioxidant N-acetylcysteine. Our in vivo model of S326C-Ogg1 activity uses the comet assay, measuring the migration of cellular DNA via electrophoresis after Ogg1 treatment. In vivo, TNF-α drives oxidative stress that impairs S326C-Ogg1 activity. Our experiments suggest that TNF-α may promote tumorigenesis in S326C-Ogg1 backgrounds by promoting oxidative stress that inhibits S326C-Ogg1 activity, while possibly driving mutagenic oxidative DNA damage.



Poster Presenters

Poster Presenter Program 1 Allen, Megan BCDB 2 Alter, Shawn MSP 3 Barnes, Dawn BCDB 4 Barnhart, Katelyn CB 5 Bauer, Nicholas BCDB 6 Bizzell, Erica MMG 7 Blanchard, Kristen GMB 8 Braykov, Nikolay PBEE 9 Brown, Harrison MSP 10 Brown, Nicole MSP 11 Byun, Phil GMB 12 Calderon, Brenda BCDB 13 Carter, Richard GMB 14 Christopher, Michael GMB 15 Cutler, Alicia BCDB 16 Deveau, Todd NS 17 Deymier, Martin MMG 18 Donlin-Asp, Paul BCDB 19 Dunn, Amy NS 20 Gaudette, Brian IMP 21 Gavile, Catherine IMP 22 Harrell, Constance NS 23 Huang, Brenda GMB 24 Hunter, Emily BCDB 25 Hwang, Juyeon BCDB 26 Infield, Daniel MSP 27 Kohn, Jordan NS 28 Limpose, Kristin CB 29 Lohr, Kelly NS 30 Meng, Jia MMG

Poster Presenter Program 31 Miller-Kleinhenz, Jasmine CB 32 Morris, Kevin BCDB 33 Noble, Donald NS 34 Peake, Bridgette MSP 35 Perez Diaz, Maylen NS 36 Perszyk, Riley MSP 37 Reding, Katherine NS 38 Rha, Jennifer BCDB 39 Rose, Samuel NS 40 Ruppersburg, Chelsey BCDB 41 Rye, Emily BCDB 42 Salgueiro, Alessandra CB

43 Sanabria-Figueroa, Eduardo MSP

44 Schmidt, Karl NS 45 Schureck, Marc BCDB 46 Seladi-Schulman, Jill MMG 47 Shah, Shardule IMP 48 Sia, Jonathan IMP 49 Stout, Kristen MSP 50 Torres, E. Shannon GMB 51 Umstead, MaKendra CB 52 Vachon, Virginia MMG 53 Vinal, Kellie MMG 54 Wasson, Jadiel BCDB 55 Weir, David CB 56 Wendt, Amanda NHS 57 Wilkinson, Scott CB 58 Williams, Kathryn BCDB 59 Woodworth-Hobbs, Myra NHS

POSTER SESSION: Odd Numbers, 12:00 – 12:45PM & Even Numbers, 12:45 – 1:30PM

GDBBS Programs

Biochemistry, Cell & Developmental Biology( BCDB) Cancer Biology (CB) Genetics & Molecular Biology (GMB) Immunology & Molecular Pathogenesis (IMP) Microbiology & Molecular Genetics (MMG)

Molecular & Systems Pharmacology (MSP) Neuroscience (NS) Nutrition & Health Sciences (NHS) Population Biology, Ecology & Evolution (PBEE)

The Eleventh Annual DSAC Student Research Symposium [26]

Megan Allen, BCDB Poster #1 POST-TRANSCRITIONAL REGULATION OF CDK5 ACTIVITY IN BRAIN NEURONS BY THE RNA BINDING PROTEIN HUD Megan Allen, Wenqi Li, Guanglu Liu, Andrew Bankston, Wei Feng, Yue Feng Department of Pharmacology, Emory University, Atlanta, GA 30322 Activity of Cyclin dependent kinase 5 (Cdk5) is essential for normal brain function and is dysregulated in neurological diseases. The abundance of p35 and p39, either of which can bind and activate Cdk5, controls Cdk5 activity. In neurons, the role of p35 is well-characterized, while p39 function remains unknown. We demonstrate that up-regulation of p39 expression is responsible for Cdk5 activity during neuronal development. Additionally, we show that the neuronal RNA-binding protein, HuD, binds p39 mRNA and regulates abundance of the p39 transcript through AU-Rich elements in the p39 3’UTR. In mice which harbor elevated hippocampal HuD expression (HuDtg+), p39 mRNA and protein are significantly up-regulated and we observe increased Cdk5 activity. Consistent with a role for Cdk5 in promoting axonal growth, mossy fibers of the hippocampus are over-projected in HuDtg+ mice, a phenomenon commonly observed in epilepsy models. We found that HuDtg+ mice respond more severely to kainic acid induced seizure than WT littermates. Importantly, elimination of p39 rescues mossy fiber and seizure phenotypes in HuDtg+ mice. These studies identify a functional role for p39 and a novel mechanism controlling Cdk5 activity in neurons, providing clues for future studies investigating Cdk5 in the normal brain and in neurological disease. Shawn Alter, MSP Poster #2 SEROTONERGIC DISRUPTIONS IN THE VMAT2-DEFICIENT MOUSE MODEL OF PARKINSON'S DISEASE Shawn Alter, Tonya N. Taylor, Minzheng Wang, and Gary Miller Department of Environmental Health, Emory University, Atlanta, GA Serotonergic denervation and loss of the raphe nuclei occur in the progression of Parkinson’s disease (PD). The selective vulnerability of monoaminergic neurons in PD is in part thought to be due to their susceptibility to oxidative stress and the instability of their neurotransmitters in the cytosol. Mice with very low expression (5%) of the vesicular monoamine transporter 2 (VMAT2) undergo progressive nigrostriatal degeneration, have decreased monoamines, increased monoamine turnover, and exhibit both motor and nonmotor symptoms of PD. Neurochemically, VMAT2- deficient mice have reductions in tissue serotonin in the striatum (85%), hippocampus (90%), and cortex (80%), and increased turnover to 5-hydroxyindoleacetic acid, as measured by HPLC. Here we report the effects of VMAT2 hypomorphy on the integrity of the serotonergic system. Immunohistochemical analyses of serotonergic neurons in the dorsal and median raphe nuclei indicate normal morphological development. We will present evaluation of serotonin transporter expression in efferent regions throughout the brain. Additionally, we will show that VMAT2-deficient mice have increased behavioral indices of depressive behavior in the forced swim and tail suspension tests that are responsive to the SSRI fluoxetine. This work demonstrates the critical role of vesicular sequestration to the viability of monoaminergic neurons.


Dawn Barnes, BCDB Poster #3 COMPLEXITY OF TROPOMYOSIN ISOFORMS IN C. ELEGANS GENERATED BY ALTERNATIVE SPLICING OF THE LEV-11 GENE Dawn Barnes1,2, Shoichiro Ono1,2 1Pathology, Emory University; 2Cellular Biology, Emory University, Atlanta, GA, USA Tropomyosin is a coiled-coil dimer that lies along actin filaments in both muscle and non-muscle cells. Dimers can connect to form an elongated polymer. Tropomyosin is involved in cellular processes including cell motility, cell division, and muscle contraction. In mammals, more than 40 isoforms of tropomyosin are expressed from 4 genes through extensive alternative splicing. We utilized Caenorhabditis elegans to study functional diversity of tropomyosin isoforms. In C. elegans, previous studies reported 4 isoforms are expressed from a single gene, lev-11, by alternative promotors and splicing. Two high-molecular-weight (HMW) isoforms are expressed from the upstream promoter and encoded by 9 exons; whereas 2 low-molecular-weight (LMW) isoforms are expressed from the downstream promoter and encoded by 7 exons. Through analysis of Expressed Sequence Tag sequences and RT-PCR, we identified a novel exon (Exon 7a) that is alternative to Exon 7b and HMW transcript containing Exon 7a. Additional RT-PCR analysis suggests that Exon 7a is also utilized in a transcript driven by the downstream promoter to produce an LMW isoform. To characterize biochemical properties, we established a system for the expression and purification of recombinant tropomyosin isoforms. Two HMW isoforms containing Exon 7b bound actin filaments with similar affinity. Katelyn Barnhart, CB Poster #4 DUAL INHIBITION OF MCL-1 BY THE COMBINATION OF CARFILZOMIB AND TG02 IN MULTIPLE MYELOMA Katelyn G. Barnhart1*, Shannon M. Matulis, Ph.D.2*, Sadae Hitosugi, MS3*, Cathy Sharp, R.N.4*, Francis Burrows, PhD5*, Ajay K. Nooka, MD, MPH6, Jonathan L. Kaufman, MD6, Sagar Lonial, M.D.6 and Lawrence H. Boise, PhD7 1Cancer Biology Graduate Program, Emory University, Atlanta, GA; 2Hematology and Medical Oncology and Winship Cancer Institute, Emory University, Atlanta, GA; 3Hematology and Medical Oncology, Emory University, Atlanta, GA;4Winship Cancer Institute of Emory University, Atlanta, GA; 5Tragara Pharmaceuticals, San Diego, CA; 6Winship Cancer Institute, Emory University, Atlanta, GA; 7Hematology and Medical Oncology, Winship Cancer Institute of Emory University, Atlanta, GA Carfilzomib (Kyprolis®) is an FDA-approved proteasome inhibitor for single-agent use in relapsed/refractory multiple myeloma (MM). We combined carfilzomib with TG02, a multi-kinase inhibitor. Both independently target Mcl-1 and MM cells are dependent on it for survival. Four different human myeloma cell lines were treated for 24 hours with carfilzomib +/- TG02. Co-treatment resulted in additive apoptosis in all lines. Carfilzomib pulse dosing was performed to mimic pharmacokinetics. Similar activity was observed with continuous and pulse dosing. Activity of the combination was also observed in freshly isolated samples from relapsed/refractory patients. Co-culture with Hs-5 cells protected against combination treatment in three lines, while addition of Hs-5 conditioned medium was protective in RPMI-8226. We then determined the molecular basis for increased apoptosis. Treated cells were isolated for western blot and RT-qPCR analysis to determine Bcl-2 family protein levels. Carfilzomib treatment caused an increase in NOXA mRNA. TG02 treatment resulted in decreased Mcl-1 protein but not mRNA expression. Mcl-1 loss at the protein level occurs in the presence of carfilzomib, therefore, the effect of TG02 is unlikely due to increased degradation. Mcl-1 translational regulation is a likely mechanism. These data suggest Mcl-1 dual inhibition is active in myeloma and warrants further preclinical testing.


Nicholas Bauer, BCDB Poster #5 AUTOMATED QUANTIFICATION OF THE SUBCELLULAR LOCALIZATION OF MULTI-COMPARTMENT BASE EXCISION REPAIR PROTEINS VIA Q-SCAN Nicholas C. Bauer1,2, Anita H. Corbett1,3, Paul W. Doetsch1,3,4,5 1Department of Biochemistry, 2Graduate Program in Biochemistry, Cell and Developmental Biology, 3Winship Cancer Institute, 4Department of Radiation Oncology, 5Department of Hematology and Medical Oncology, Emory University School of Medicine, Atlanta, GA Defects in various components of base excision repair (BER) lead to increased mutation frequencies and genetic/genomic instability. Although the biochemical mechanism of BER is well-characterized, few studies have addressed how this critical pathway is regulated. Many BER proteins localize to both the nucleus and mitochondria, and changes in localization have been observed in Saccharomyces cerevisiae Ntg1 in response to oxidative stress. We hypothesize that dynamic localization is a general mode of regulation for BER. However, quantifying the extent of DNA repair protein localization is a major barrier to elucidating this potential regulatory mechanism. To address this issue, we developed a novel technique termed Quantitative Subcellular Compartmentalization Analysis (Q-SCAn). This analysis technique uses markers to define subcellular compartments and then quantifies the relative amount of target protein in each compartment of each cell. To validate Q-SCAn, we analyzed relocalization of the transcription factor Yap1 following oxidative stress and then extended the approach to multi-compartment localization by examining two DNA repair proteins critical for the base excision repair pathway, Ntg1 and Ung1. Our findings demonstrate the utility of Q-SCAn for quantitative analysis of the subcellular distribution of multi-compartment proteins, and has revealed new information about the regulation of Yap1, Ntg1 and Ung1. Erica Bizzell, MMG Poster #6 CHARACTERIZATION OF THE SECRETED SERINE PROTEASE, RV2223C OF MYCOBACTERIUM TUBERCULOSIS. Erica Bizzell,1,2, Maria Georgieva1,2 and Jyothi Rengarajan1,3

1Emory Vaccine Center. 2Microbiology and Molecular Genetics Program, Graduate Division of Biological and Biomedical Sciences, Emory University. 3Division of Infectious Diseases, Emory University. Mycobacterium tuberculosis (Mtb) has evolved multiple strategies to evade host immune defenses. Several pathogenic bacteria use extracellularly secreted proteases to regulate processes ranging from repression of cytokine production to degradation of surface-associated host proteins. Mtb encodes several putative secreted proteases whose functions are poorly understood. Our lab is characterizing the predicted Mtb protease, Rv2223c. Rv2223c is located in a predicted operon with cell wall-associated serine protease, Hip1 (Rv2224c), with which it shares 52% amino acid identity. Hip1 is involved in modification of the Mtb cell wall during infection, and functions by dampening normal immune responses to Mtb infection. Using the model system Mycobacterium smegmatis expressing Mtb Rv2223c, we found that Rv2223c is secreted from mycobacterial cells in a signal-sequence dependent manner and undergoes autoproteolytic cleavage upon secretion. Additionally, we have observed that Rv2223c interacts with the Hip1 physiological substrate, GroEL2, suggesting potentially overlapping or cooperative functions of these proteases. We hypothesize that Rv2223c is a secreted protease that functions by cleaving Mtb substrates and/or host cell substrates to modify host responses during Mtb infection. Further work is currently being conducted to determine the enzymatic activity of Rv2223c and to identify novel bacterial and host interactors and/or substrates.


Kristen Blanchard, GMB Poster #7 ENZYME EVOLUTION: EXAPTATION IN THE E. COLI GENOME Kristen Blanchard, Ichiro Matsumura Department of Biochemistry, Emory University How do new catalytic functions evolve? I hypothesize that enzymes often evolve through exaptation - proteins that have evolved towards one catalytic goal can be co-opted for a secondary purpose. Our lab has previously demonstrated that many proteins in E. coli have weak secondary activity that can be revealed through expression at high levels on multicopy plasmids. When these promiscuous enzymes are expressed at very high levels, their secondary activity is sufficient to rescue a lethal deficiency. I hypothesize that these promiscuous proteins can also be exploited in their native state on the E. coli chromosome. To this end, I have mutagenized E. coli conditional auxotrophs and selected for rescues of the lethal phenotype. By discovering which mutations rescue the auxotrophs, it should be possible to discover new examples of proteins with secondary activity. This work should demonstrate that the E. coli proteome contains ample candidates for exaptation; many proteins exist which have evolved towards one primary function, but under selective conditions may be evolved to perform a second function. Such a finding would aid in the understanding of how new catalytic functions emerge, and would also help change the way proteins are evolved in the lab. Nikolay Braykov, PBEE Poster #8 IMPACT OF ANTIBIOTIC EXPOSURE AND OTHER MANAGEMENT PRACTICES ON THE PHENOTYPIC AND GENOTYPIC COMPOSITION OF DRUG-RESISTANT E. COLI IN BROILER CHICKENS Nikolay Braykov1, Lixin Zhang2, William Cevallos2, Natalia Burbano, Nadia López, Carl Marrs2, Betsy Foxman2, Gabriel Trueba, Joseph Eisenberg2, Karen Levy4 1 Emory University Dept. of Biology, Atlanta, GA, United States, 2 University of Michigan Dept. of Epidemiology, Ann Arbor, MI, United States, 3 Universidad San Francisco de Quito, Quito, Ecuador, 4 Emory University Dept. of Environmental Health, Atlanta, GA, United States Non-therapeutic use of antibiotics in animal husbandry promotes the development of antibiotic resistance (AR), contributing to an environmental reservoir of resistance genes and potentially threatening human health. A factorial experiment was conducted in a chicken farm in Northern Ecuador to describe the emergence, phenotypic and genotypic diversity of AR-Escherichia coli in broilers raised under different conditions. Seventy-two newly hatched chickens raised in single, 6-chicken and 34-chicken cages were administered tetracycline via water on days 9 and 24, and sampled on days 1, 19-21 and 38. Susceptibility to twelve antibiotics was categorized using disk diffusion. Isolates were fully genotyped on a microarray platform. In preliminary data from 559 isolates, we found that >80% of samples were resistant to at least one antibiotic on day 1, suggesting chickens arrived at the facility pre-colonized. Prevalence of AR decreased over time without statistically significant difference between treatment and controls. There was a drastic increase in the number of unique genotypes after day 1. Time was more important than selection pressure or number of chickens/cage in explaining the phenotypic and genotypic community composition of samples, suggesting that physiological factors or external environmental colonization play a bigger role in shaping the ecology of AR E. coli.


Harrison Brown, MSP Poster #9 BIOENGINEERED COAGULATION FACTOR VIII ENABLES LONG-TERM CORRECTION OF MURINE HEMOPHILIA A FOLLOWING LIVER-DIRECTED ADENO-ASSOCIATED VIRAL VECTOR DELIVERY Harrison C. Brown, J. Fraser Wright, Shangzhen Zhou, Jordan E. Shields H. Trent Spencer, Christopher B. Doering Clinical data support the feasibility and safety of adeno-associated viral (AAV) vectors in gene therapy applications. Despite several clinical trials of AAV-based gene transfer for hemophilia B, a unique set of obstacles impede the development of a similar approach for hemophilia A. These include 1) the large transgene size of factor VIII (fVIII), 2) humoral immune responses to the transgene product, 3) inefficient biosynthesis of human coagulation factor VIII, and 4) dose limitations imposed by capsid mediated cytotoxic immunity. To address the latter two of these concerns, our laboratory has developed a bioengineered fVIII molecule, designated ET3, which demonstrates 10-100 fold improvement in fVIII biosynthesis in recombinant expression systems. When constructed within an oversized AAV vector expression cassette, the ET3-AAV genomes packaged into viral particles primarily as partial genome fragments. Despite the potential reduction in potency caused by genome fragmentation, a single peripheral vein injection the oversized ET3-AAV vector into immune-competent hemophilia A mice resulted in correction of the fVIII deficiency at vector doses lower than those of smaller, previously reported fVIII constructs, thus demonstrating that ET3 improves vector potency thereby mitigating two of the critical barriers remaining to AAV gene therapy for hemophilia A. Nicole Brown, MSP Poster #10 INTEGRATION OF H-RAS AND Gαi SIGNALING PATHWAYS BY REGULATOR OF G PROTEIN SIGNALING 14 (RGS14) Nicole E. Brown1, Christopher P. Vellano1, Devrishi Goswami2, Patrick R. Griffin2, and John R. Hepler1 1Department of Pharmacology, Emory University 2Department of Molecular Therapeutics, the Scripps Research Institute – Scripps Florida Regulator of G protein signaling 14 (RGS14) is a scaffolding protein that integrates G protein and H-Ras signaling pathways. RGS14 possesses an RGS domain that binds active Gαi/o-GTP subunits to promote GTP hydrolysis and a G protein regulatory (GPR) motif that binds inactive Gαi1/3-GDP subunits to form a stable complex at cellular membranes. RGS14 also contains two Ras-binding domains (RBDs), the first of which (RBD1) binds activated H-Ras. Here we show RGS14 interactions with H-Ras-GTP are regulated by inactive Gαi1-GDP. Using bioluminescence resonance energy transfer (BRET), we show RGS14-Luc and active H-Ras(G/V)-Venus exhibit a robust BRET signal that is enhanced in the presence of inactive Gαi1-GDP, but not active Gαi1-GTP. Moreover, experiments using hydrogen deuterium exchange (HDX) indicate binding of inactive Gαi1-GDP through the GPR motif stabilizes RBD1, potentially regulating interactions with activated H-Ras-GTP. HDX also indicated changes in the RGS domain upon binding Gαi1-GDP suggesting RGS activity may be regulated by interactions with Gαi. Further BRET studies examining the RGS domain using Luc-RGS14 and aluminum fluoride (AlF4

-) activated Gαi1-YFP demonstrate a robust BRET signal that is decreased in the presence of activated H-Ras(G/V). Together, these studies highlight the interdomain regulation of RGS14 by its binding partners, Gα and H-Ras.


Phil Byun, GMB Poster #11 GENETIC ANALYSIS OF TISSUE INVASION DRIVEN BY THE DROSOPHILA NCOA-1,2,3 HOMOLOG TAIMAN Phil K. Byun1, Kenneth H. Moberg1 1Emory University Cancer cells spread from the primary tumor and seed new tumors in other organs and tissues through a process called metastasis. While the gross morphologic changes associated with metastasis are well defined, specific pathways that trigger these ‘motogenic’ phenotypes in vivo are not. Model systems, such as Drosophila melanogaster, have been critical in providing powerful genetics tools to define such pathways. We have discovered that ectopic expression of the Drosophila steroid-receptor transcriptional coactivator protein Taiman (Tai) is sufficient to drive non-invasive pupal wing disc cells to invade into adjacent thoracic tissue. Tai normally supports invasive and motile behavior of ovarian border cells during the oocyte maturation but little is known about the transcriptional targets or interacting cofactors involved in this process. We find that Tai binds directly to the Hippo pathway target and transcriptional co-activator Yorkie, whose vertebrate homolog Yap1 promotes metastasis in mouse models of breast cancer. In light of these data, we hypothesize that Tai functions with specific factors such as Yorkie to promote the invasion of wing cells, and that a genetic screen for modifiers of this phenotype will define pathway(s) through which Tai controls cell motility. Brenda Calderon, BCDB Poster #12 STRUCTURAL ANALYSIS OF NC886, A CELLULAR NON-CODING RNA REGULATOR OF THE ANTIVIRAL KINASE PKR Brenda M. Calderon1,2, and Graeme L. Conn2

1Biochemistry, Cell and Developmental Biology Program, Emory University, Atlanta, GA 2Department of Biochemistry, Emory University, Atlanta, GA The double-stranded (ds)RNA-activated protein kinase (PKR) senses dsRNA produced during viral replication and acts as the first line of defense in the innate immune response to viral infection. Activated PKR phosphorylates the eukaryotic initiation factor 2 (eIF2), halting protein synthesis and viral replication. How PKR basal activity is regulated in the absence of viral infection is unknown. Recently, the cellular non-coding RNA 886 (nc886) was discovered and proposed to act as a novel, endogenous regulator of PKR. We hypothesize that nc886 regulates PKR by binding to the monomeric form and preventing the dimerization step of activation. This maintains PKR in an inactive state that is poised for a rapid response to viral infection. Our goal is to define the molecular mechanism of nc886-mediated PKR inhibition using RNA structure probing and mutational analyses combined with binding and activity assays. We show that nc886 RNA exists as two stable, structurally and functionally distinct conformations. One conformation binds and inhibits PKR autophosphorylation while the other lacks both activities. By delineating the structural basis for PKR interaction with the active nc886 conformer, we will uncover the structural motifs that control PKR inhibition and determine how PKR is poised to act in the antiviral response.


Richard Carter, GMB Poster #13 NEURAL PROGENITOR CELLS DERIVED FROM TRANSGENIC HUNTINGTON’S DISEASE MONKEY INDUCED PLURIPOTENT STEM CELLS DEVELOP DISEASE CELLULAR PHENOTYPES Richard L. Carter1,2,3 , Chen YJ2 , Kunkanjanawan T3,4, Chan AWS1,2,3* 1 Genetics and Molecular Biology Program, Graduate Division of Biological and Biomedical Sciences, Emory University 2 Department of Human Genetics, Emory University School of Medicine 3 Division of Neuropharmacology and Neurologic Diseases, Yerkes National Primate Research Center Emory University, Atlanta, GA 30329, USA. 4 Embryo Technology and Stem Cell Research Center, School of Biotechnology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand. Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder caused by an expansion of CAG repeats in the coding region of the Huntingtin (HTT) gene. A major focus of HD research is on understanding and modeling mechanisms contributing to the degeneration of neuronal populations. Pluripotent cellular models are emerging as a powerful tool to understand disease related changes as neurons develop, therefore we describe the use of neural progenitor cells (NPCs) derived from transgenic HD monkeys as a cellular platform for pathological and therapeutic study in HD. We have derived NPCs from iPSCs of an HD monkey (rHD-NPCs). rHD-NPCs were capable to differentiate to GABA positive neurons in vitro and in vivo. rHD-NPCs showed disease associated susceptibility to cell stress demonstrated by increased apoptosis. Upon differentiation to neurons, HD neural cells developed cellular features of HD including the formation of nuclear inclusions and accumulation of oligomeric mutant HTT aggregates. The cell death phenotype and mutant protein aggregation was rescued upon siRNA mediated reduction of HTT transcript, and after treatment with memantine in culture. These findings highlight the utility of iPSCs derived NPCs from transgenic HD monkeys in providing a unique non-human primate platform for studying HD pathogenesis and therapeutic strategies. Michael Christopher, GMB Poster #14 THE HISTONE DEMETHYLASE LSD1/KDM1A IS A MASTER REGULATOR OF THE STEM CELL FATE Michael A. Christopher1,3, Dexter A. Myrick2,3, and David J. Katz3 1Genetics and Molecular Biology, 2PREP, 3Department of Cell Biology During transcription, stem cell genes acquire the histone modification dimethylation of lysine 4 on histone H3 (H3K4me2). It has previously been shown that LSD1 is necessary to repress stem during stem cell differentiation by removing H3K4me2. Based on this, we investigated the role of LSD1 during neural stem cell (NSC) differentiation in the mouse. Deletion of Lsd1 in NSCs results in perinatal lethality within 24 hours of birth. We find that cultured motor neurons from these mutants continue to express the stem cell genes Nestin and Sox2. These findings suggest that LSD1 is generally required to repress the stem cell network during neural stem cell differentiation. To determine if LSD1 continues to represses the stem cell network in differentiated cells, we inducibly deleted LSD1 throughout the adult mouse. Surprisingly, loss of LSD1 results in paralysis and death in as little as 4 weeks with massive neuronal cell death in the hippocampus and surrounding cortex. Additionally, immunohistochemistry demonstrated the reactivation of the master pluripotency gene Oct4 exclusively in these dying neurons. These data suggest that LSD1 is continually required to repress the stem cell network in differentiated neurons. As a result, we conclude that LSD1 is a master regulator of the stem cell fate.


Alicia Cutler, BCDB Poster #15 NUCLEAR IMPORT AND PERMEABILITY IN MULTINUCLEATED SKELETAL MUSCLE CELLS Alicia Cutler1, Jennifer Jackson1, Justin Ho1, and Grace Pavlath1

1Department of Pharmacology, Emory University

Skeletal muscle is critical for survival and quality of life. Proper gene expression is essential for myogenesis and maintenance of skeletal muscle. Proteins that regulate gene expression are synthesized in the cytoplasm and cross the nuclear pore to access their targets in the nucleus. The intact nuclear pore allows free diffusion of small molecules between the nucleus and cytoplasm but excludes molecules larger >40kDa. To cross the nuclear pore, larger proteins must be bound to a nuclear transport receptor, which recognizes a nuclear localization signal (NLS) within the cargo protein. During myogenesis, myoblasts differentiate into myocytes, which fuse to form mature myotubes containing thousands of nuclei. Our results demonstrate that nuclear properties vary during myogenesis and aging. During myogenesis the ability of nuclei to import a protein containing the classical NLS (cNLS), changes with differentiation. Almost all myoblast nuclei are cNLS import-competent; however, only 38% of myocyte nuclei are cNLS import-competent. As myocytes develop into mature myotubes, cNLS import-competence is restored. Additionally, as skeletal muscle ages, the percentage of myonuclei permeable to >40kDa molecules not bound to nuclear transport receptors increases. These results suggest nuclear characteristics are connected to stages of muscle development and aging potentially affecting gene expression. Todd Deveau, NS Poster #16

INTRANASAL ADMINISTRATION OF INDUCED-PLURIPOTENT STEM CELL DERIVED NEURAL PROGENITORS IMPROVES FUNCTIONAL RECOVERY FOLLOWING ISCHEMIC STROKE IN MICE

Todd C. Deveau1, Katharine Yuengling1, Xiaohuan Gu1, Shan Ping Yu1, and Ling Wei1 1Department of Anesthesiology, Emory University School of Medicine, Atlanta, GA 30322 Induced Pluripotent Stem (iPS) cells are currently under extensive examination in cellular biology and disease treatment investigations. iPS cells can differentiate into neurons and non-neuronal cells in vitro and after transplantation into the ischemic brain of experimental stroke animal models. The present study investigates intranasal administration (iNA) of induced pluripotent stem cell-derived neural progenitors (iNA-iPS-NPCs) to improve functional recovery following ischemic stroke. Adult male mice were subjected to a focal ischemic insult primarily in the barrel cortex. Vehicle or iPS-NPCs were administered into the nasal passages of mice 1-3 hours after stroke. At day 3 and 7 after transplant, there was a trend in local cerebral blood flow (LCBF) increase in the ischemic region of animals that received iNA- iPS-NPCs compared to vehicle. Animals who received iNA- iPS-NPC administration showed significant improvements in functional recovery compared to vehicle at day 7. Current experiments are examining functional recovery at even delayed time points after stroke. Further analysis will examine gene regulation and signaling pathways using WB, RT-PCR, and histological techniques to gain insight into the mechanisms behind iNA-iPS-NPC treatment. Our available results suggest that intranasal administration of iPS-derived neural progenitors is a viable treatment approach for stroke and other CNS disorders.


Martin Deymier, MMG Poster #17

GENETIC & PHENOTYPIC CHARACTERISTICS OF SUBTYPE C FULL LENGTH GENOME HIV-1 FROM LINKED HETEROSEXUAL TRANSMISSION PAIRS

Martin J. Deymier1, Zachary S. Ende1, Daniel T. Claiborne1, William Kilembe2, Susan Allen1,2, and Eric Hunter1,2. 1Emory University, Atlanta, GA, USA, 2Zambia Emory HIV Research Project, Lusaka, Zambia2

In 80-90% of heterosexual transmissions of HIV, a genetic bottleneck occurs, in which an individual, with a diverse viral quasispecies, transmits a single viral variant, the Transmitted/Founder (TF), to a naïve host. Previous genetic studies of the HIV envelope glycoprotein have suggested a non-stochastic process for transmission and evidence for selection of a specific TF variant. Although some genetic signatures have been found, no biological correlate for TF variants have thus far been elucidated. We have performed HIV near full-length single genome amplification followed by direct sequencing, from plasma of six subtype C acutely infected individuals and each of their chronically infected linked partners from the Zambia-Emory HIV Research Project. We provide genetic evidence for the selection of TF variants that more closely resembles the Most Recent Common Ancestor from the donor quasispecies than the median of the donor variants (p=0.0313). The generation of full-length infectious molecular clones from these viruses has provided us with the unique observation, for one transmission pair, that the TF virus is not necessarily the highest replicating virus from the donor quasispecies. The genetic bottleneck thus provides an opportunity for understanding the unique properties of TF viruses and may provide potential targets for intervention. Paul Donlin-Asp, BCDB Poster #18

THE SURVIVAL OF MOTOR NEURON PROTEIN IN mRNP COMPLEX ASSEMBLY Paul G. Donlin-Asp1,2, Claudia Fallini2,4, Gary J. Bassell2,3, Wilfried Rossoll2. 1Biochemistry, Cell and Developmental Biology Graduate Program, Emory University Laney Graduate School, 2Dep. of Cell Biology and Center for Neurodegenerative Disease, 3Dep. of Neurology, Emory University School of Medicine, Atlanta, GA, 4Dep. of Neurology, University of Massachusetts Medical School, Worcester, MA Spinal muscular atrophy (SMA) is a neuromuscular disease characterized by a specific degeneration of motor neurons. SMA results from a reduction in the survival of motor neuron (SMN) protein, which is ubiquitously expressed with a well characterized role in promoting the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs). While underlying defects in splicing have been observed in SMA models, these defects are not unique to motor neurons, leaving their role in motor neuron degeneration unclear. We and others have demonstrated that SMN colocalizes with various mRNA binding proteins known to play vital roles in mRNA transport. Additionally, a defect in localization of mRNA and mRNA-binding proteins in the axon terminus in motor neurons from the SMA mouse model has been observed. These and other data suggest that SMN plays a non-canonical role in mediating the assembly and delivery of mRNA and protein associated complexes (mRNPs). We have optimized trimolecular fluorescence complementation (TriFC) as a novel approach to visualize the assembly of mRNPs in cultured motor neurons derived from control and SMA embryos. Our results show a marked reduction in TriFC fluorescent signal in SMA motor neurons, suggestive that SMA motor neurons are deficient in mRNP complex assembly.


Amy Dunn, NS Poster #19

GENETIC MANIPULATION OF VESICULAR FUNCTION AS A POTENTIAL MEDIATOR OF DOPAMINERGIC NEURO-TOXICANT VULNERABILITY

Amy R. Dunn1,3, Kelly M. Lohr1,3, Kristen A. Stout2,3, Alison I. Bernstein3, Thomas S. Guillot3, Minzheng Wang3, Yingjie Li3, Ali Salahpour4, Gary W. Miller3,4.1Neuroscience Graduate Program, Emory University, 2Molecular and Systems Pharmacology Graduate Program, Emory University, 3Department of Environmental Health, Emory University, 4Center for Neurodegenerative Disease, Emory University, 5Department of Pharmacology and Toxicology, University of Toronto.

Proper vesicle function is critical for dopamine neuron health and contributes to neuron integrity after toxicant exposure. Dopaminergic vesicles have two important roles: packaging transmitter for synaptic transmission, and sequestering toxicants to prevent cell damage. Various proteins maintain vesicular function, and the current study focuses on two (vesicular monoamine transporter 2 (VMAT2) and the synaptic vesicle glycoprotein 2C (SV2C)). Our lab has previously shown that impaired vesicular function causes dopamine mishandling and cell death, but the converse—enhanced vesicular function—had been previously unexplored. The current investigation explores the consequences of overexpressing VMAT2 in a transgenic mouse model (VMAT2-HIs). Analysis of vesicular morphology by electron microscopy indicates that VMAT2 overexpression increases vesicular volume by a third, which likely underlies neurochemical changes and resistance to toxicants. A novel target, SV2C, is a pharmacologically-targetable, undercharacterized protein thought to be involved in vesicular storage and release. It is restricted to dopaminergic brain regions, and SV2C genotype mediates the protective effect of nicotine against Parkinson’s disease. Here, we describe and characterize our SV2C-KO mouse. VMAT2 overexpression and SV2C-KO are excellent tools with which we will investigate the effect of altered vesicular function on the vulnerability of dopaminergic degeneration after toxicant exposure.

Brian Gaudette, IMP Poster #20

HETEROGENEOUS BCL-2 FAMILY EXPRESSION IN WALDENSTRÖM MACROGLOBULINEMIA DETERMINES RESPONSE TO AGENTS OF INTRINSIC APOPTOSIS. Brian T. Gaudette1,3,4, Kasyapa S. Chitta, Ph.D.5, Stéphanie Poulain6, Bhakti Dwivedi, Ph.D.3, Doris Powell2,3, Kelvin P. Lee, M.D.7, Asher A. Chanan-Khan, M.D.8, Stephen M. Ansell, M.D., Ph.D.9, Xavier Leleu, M.D.10, Ph.D., Jeanne Kowalski, Ph.D.3, Paula Vertino, Ph.D.2,3, Sagar Lonial, M.D.1,3, Lawrence H. Boise, Ph.D.1,3,4 1Departments of Hematology and Medical Oncology and Cell Biology, 2Department of Radiation Oncology, 3Winship Cancer Institute, 4Graduate Program in Immunology and Molecular Pathogenesis - Emory University, Atlanta, GA 30322; 5Cancer Biology, Mayo Clinic, Jacksonville, FL; 6Service d'Hématologie Immunologie Cytogénétique, Hopital de Valenciennes, Valenciennes, France; 7Immunology, Roswell Park Cancer Institute, Buffalo, NY; 8Hematology and Oncology, Mayo Clinic, Jacksonville, FL; 9Hematology and Oncology, Mayo Clinic, Rochester, MN; 10Service des Maladies du Sang, Hopital Claude Huriez, CHRU Lille, Lille, France Waldenström Macroglobulinemia (WM) is a disorder of lymphoplasmacytoid cells that inhabit lymph nodes and the bone marrow. 91% of WM cases carry an activating mutation of MyD88 (L265P). Having observed previously that plasmablasts driven to differentiate using the TLR4 ligand LPS enter a Bcl-xL-dependent state, we sought to determine if WM cells were Bcl-xL-dependent. We treated three WM cell lines all harboring the MyD88 (L265P) mutation with ABT-737, bortezomib and arsenic trioxide (ATO) all agents that induce apoptosis through the intrinsic pathway. While all three lines were insensitive to ABT-737, one line was insensitive to all three agents. Further examination of the Bcl-2 family proteins showed dysregultion of the same key step in the intrinsic apoptosis pathway with downregulation of Bim in BCWM.1 and MWCL-1 cells and loss of Bax and Bak in RPCI-WM1 cells. Furthermore, we examined expression data from WM patients and determined that WM cells expressed lower levels of pro-apoptotic BH3 proteins than MM cells. These WM cell lines demonstrate that sensitivity to agents that kill through the intrinsic apoptotic pathway may vary within a disease that is characterized by a single activating mutation and suggests that additional heterogeneous events regulate the expression of Bcl-2 family proteins.


Catherine Gavile, IMP Poster #21

CD28 AND CD86 REGULATE INTEGRIN SURFACE EXPRESSION IN MYELOMA CELL LINES Catherine M. Gavile1, David Egas1, Gregory Doho1, Sagar Lonial1, Kelvin P. Lee2, Lawrence H. Boise1 1Departments of Hematology and Medical Oncology and Winship Cancer Institute, Emory University, Atlanta GA 30322 2Departments of Immunology and Medicine, Roswell Park Cancer Institute, Buffalo, NY 14263 CD28 and CD86 are better known for their roles in T-cell costimulation. Recent reports have shown that this signaling module also plays an important role in the maintenance of normal long-lived plasma cells (LLPCs). As myeloma cells retain most of the physiological characteristics of LLPCs, our lab is currently investigating the role of CD28 and CD86 in myeloma. Previously, we have shown that CD28 and CD86 are necessary for myeloma cell survival, as knockdown of expression of either CD28 or CD86 leads to cell death in 3 myeloma cell lines. We have also shown that blocking the ability of these molecules to interact using a chimeric soluble receptor CTLA4-Ig (Abatacept) also leads to cell death. We are currently investigating what downstream signals are mediated upon CD28-CD86 signaling. Based on results obtained via RNA-Seq, our data indicate that CD28-CD86 signaling plays a role in regulation of integrin expression in myeloma cell lines.

Constance Harrell, NS Poster #22

CONSUMPTION OF A HIGH-FRUCTOSE DIET DURING ADOLESCENCE INCREASES DEPRESSIVE-LIKE BEHAVIORS, ALTERS HPA AXIS FUNCTION, AND REMODELS HYPOTHALAMIC GENE EXPRESSION

Constance S. Harrell1,2, Zachary P. Johnson3,4, Jillybeth Burgado5, Gretchen N. Neigh1,6

1Department of Physiology; 2Molecules to Mankind Doctoral Pathway; 3Nonhuman Primate Genomics Lab, 4Yerkes National Primate Center, 5NET/Work Program, Center for Behavioral Neuroscience; 6Department of Psychiatry and Behavioral Sciences

The 25% increase in fructose consumption over the past thirty years has been linked to obesity. Fructose is implicated in dyslipidemia and insulin resistance but little is known about its impact on behavior and the brain. Given that adolescents are the highest consumers of fructose and in a “critical period” of development, we examined the effects of an adolescent high-fructose diet on behavior, the HPA axis, and whole-transcriptome hypothalamic expression. Male rats were assigned to either standard chow or a 55% fructose diet from weaning to adulthood. Weight, blood glucose and consumption were measured throughout while behavior and serum corticosterone were assessed in adulthood. Hypothalamic RNA was extracted for RNA-Sequencing. Fructose consumption resulted in elevated baseline corticosterone yet a blunted response to acute stress. Fructose also reduced struggle and increased immobility in the forced swim test. Moreover, fructose impacted hypothalamic gene expression, altering 966 genes, or 5.6% of those tested. Principal Components Analysis demonstrated clustering by diet type. Both the dopamine transporter and tyrosine hydroxylase were upregulated by fructose and highly ranked in the first component. Several pathways involved in dopamine signaling were significantly enriched by differentially expressed genes, suggesting a potential link among hypothalamic dopaminergic expression, consumption, and behavior.


Brenda Huang, GMB Poster #23

OLIGODENDROCYTE DYSFUNCTION IN HUNTINGTON’S DISEASE MICE

Brenda Huang1,2, Cindy Wei1, Marta Gaertig1, Xiao-Jiang Li1, Shihua Li1

1Department of Human Genetics, Emory University, Atlanta, GA 303222 Graduate Program in Genetics and Molecular Biology

Huntington’s disease is a neurodegenerative disorder that is caused by an expanded polyglutamine tract in the N-terminal region of huntingtin. It is characterized by cognitive and behavioral deficits, as well as movement disorders. While the effect of mutant huntingtin on neuronal function has been extensively studied, its effect on the function of glial cells remains to be fully investigated. There is evidence of white matter abnormalities in both HD patients and HD mouse models, suggesting that mutant huntingtin might affect oligodendrocyte function. We hypothesize that mutant huntingtin causes oligodendrocyte dysfunction, which contributes to the pathogenesis of the disease. To this end, we have generated transgenic mice that express an N-terminal fragment of mutant huntingtin exclusively in oligodendrocytes. These mice develop a tremor by 2 months of age and have locomotor and behavioral deficits, as well as body weight loss and reduced lifespan compared to control mice. Reduced myelination is present in the striatum of mutant mice, and preliminary data suggest that there is a dysregulation of myelin gene expression. These results suggest that the expression of mutant huntingtin in oligodendrocytes plays an important role in HD pathogenesis. Emily Hunter, BCDB Poster #24

MECHANISMS OF CILIARY ASSEMBLY

Emily L. Hunter1, Lea M. Alford1, Winfield S. Sale1* 1Department of Cell Biology, Emory University School of Medicine, Atlanta, GA Cilia, found on most human cells, play an essential role in development, cell signaling, and organ function. Defective ciliary assembly results in diseases collectively known as ciliopathies. The goal of our research is to determine mechanisms of ciliary assembly, focusing on the assembly of the axonemal radial spoke structure that is essential for regulating ciliary motility. The radial spoke assembles into a large precursor complex before entry into the cilium and subsequent transport to the distal tip for final assembly. To further assess radial spoke transport, we analyzed the Chlamydomonas mutant pf27 that is deficient in radial spokes. We determined the pf27 radial spokes that dock to the microtubule core are fully assembled and localize to the proximal end of the cilium. We postulate the radial spoke deficiency is caused by a defect in Intraflagellar Transport (IFT) of the radial spoke precursor. Consistent with this hypothesis, assembly of radial spokes occurs asynchronously from other axonemal components in pf27 and radial spoke proteins are greatly reduced in the pf27 membrane-matrix fraction of the cilium where IFT occurs. We are now testing the hypothesis that PF27p is required for normal radial spoke transport.


Juyeon Hwang, BCDB Poster #25

CONTROL OF PP2A FAMILY PHOSPHATASE METHYLATION BY LCMT-1 AND PME-1

Juyeon Hwang1,2, Jocelyn A. Lee1,2, and David C. Pallas1,2 1Department of Biochemistry and Winship Cancer Institute, Emory University School of Medicine, Atlanta, Georgia 30322, USA, 2Biochemistry, Cell, Developmental Biology Graduate Program, Emory University School of Medicine, Atlanta, Georgia 30322, USA. Protein phosphatase 2A (PP2A) and the PP2A-related protein phosphatases, PP4 and PP6, are multifunctional serine/threonine protein phosphatases involved in various cellular processes including cell cycle control, apoptosis, and development. Methylation of the PP2A catalytic (C) subunit is a reversible process tightly regulated by leucine carboxyl methyltransferase 1 (LCMT-1) and protein phosphatase methylesterase 1 (PME-1). The site of methylation on PP2A is conserved in PP4 and PP6, but the identities of the PP4 and PP6 methyltransferase and methylesterase enzymes are not known. We now show that LCMT-1 is the major methyltransferase for both PP4 and PP6 and that PME-1 is responsible for PP4 and PP6 demethylation. Analysis of PP2A-related protein phosphatase complex formation by blue native gel electrophoresis suggests that loss of LCMT-1 affects complex formation. Consistent with these results, loss of LCMT-1 leads to a reduction in the amount of PP2A B regulatory subunit and a decrease in association of PP2A C subunit with the remaining B subunit. Lastly, loss of LCMT-1 increased protein stability of PP4 C subunit without affecting translation initiation. Collectively, our data suggest that LCMT-1 and PME-1 regulate the methylation of PP2A-related phosphatases, and consequently, protein phosphatase complex formation and stability of individual subunits.

Daniel Infield, MSP Poster #26

USE OF FUNCTIONAL CROSSLINKERS TO STUDY CONFORMATIONAL CHANGES ASSOCIATED WITH CFTR GATING

Daniel T. Infield1,2,, Guiying Cui, MD Ph.D.2, Christopher Kuang2, and Nael A McCarty, Ph.D.1,2,3

1 Emory University Molecular and Systems Pharmacology Graduate Program, 2 Departmentof Pediatrics, Emory University School of Medicine, 3Childrens Healthcare of Atlanta The chloride ion channel CFTR is a member of the ABC transporter family. However, it is unknown whether, like transporters, the transmembrane domains of CFTR undergo an inward-to-outward facing transition upon NBD dimerization. To test this, we made cysteine substitutions in CFTR in extracellular loops (ECL) 1 and 4, which appose each other in inward-facing but not outward-facing structures of ABC transporters. We found the macroscopic current of D110C/K892C-CFTR channels (but not corresponding single mutants) was increased in the presence of DTT. Multichannel patch recordings revealed that DTT increased number of open double cysteine mutant channels, indicating that they were locked into closed state by a spontaneous disulfide bond prior to DTT. To further understand the kinetics of the interaction, we applied Cd2+ and found that D110C is transiently bound by Cd2+, causing a rapidly reversible inhibition of current. Under identical cumulative exposure, Cd2+ inhibited D110C/K892C-CFTR irreversibly. K892C and E115C-CFTR were only slightly functionally modified (<2%) by Cd2+, though E115C-CFTR was labeled by MTS-TAMRA, indicating accessibility. Neither D112C/K892C nor E115C/K892C CFTR were affected by DTT, and D112C/K892C was not affected by Cd2+. The data suggest ECL1 and ECL4 come into close proximity in a closed state of CFTR.


Jordan Kohn, NS Poster #27

THE IMPACT OF ACUTE PSYCHOSOCIAL STRESS ON GENOME-WIDE INFLAMMATORY SIGNALING IN THE FACE OF CHRONIC SOCIAL ADVERSITY. Jordan N. Kohn1,2, Steve W. Cole3,4, Mark E. Wilson1, and Zach P. Johnson1 1Division of Developmental and Cognitive Neuroscience, Yerkes National Primate Research Center, Atlanta, GA, 2Graduate Program in Neuroscience, Emory University, Atlanta, GA, 3University of California Los Angeles Molecular Biology Institute, 4The Norman Cousins Center, University of California, Los Angeles, CA. Chronic social stress leads to chronic inflammation, a well-established risk factor for disease and all-cause mortality. Understanding the relationship between stress and inflammation is therefore an important priority for improving human health. Studies suggest that chronic stress disrupts glucocorticoid (GC) signaling at multiple levels of the limbic-hypothalamic-pituitary-adrenal (LHPA) axis, leading to amplified inflammatory gene expression by immune cells. Chronic stress results from repeated exposure to acute stressors, so one may gain insight into the mechanisms of stress-induced inflammation by studying the immune response. In this study, we used a well-validated translational model of social subordination stress in female rhesus macaques to examine social status-dependent differences in the genome-wide inflammatory response to psychosocial stress challenge. Total mRNAs were sequenced from mononuclear cells, and promoter-based bioinformatics analysis of differentially expressed genes revealed core promoter enrichment for proinflammatory transcription factors, nuclear factor kappa B (NF-κB) and AP1, at all timepoints in social subordinates. Interestingly, there was a trend toward promoter enrichment of glucocorticoid receptor (GR)-binding motifs in up-regulated genes, in the absence of plasma cortisol or interleukin-6 differences. This finding suggests a unique mechanism by which GR-mediated transcriptional responses during acute stress are enhanced in chronic social adversity to dampen concurrent hyperinflammatory signaling. Kristin Limpose, CB Poster #28 REGULATION OF BASE EXCISION REPAIR IN GENOME MAINTENANCE: IMPLICATIONS IN CANCER Kristin Limpose1, Erica Werner2, Anita H. Corbett2, Paul W. Doetsch1,2,3,4

1 Department of Cancer Biology, Emory University School of Medicine, Atlanta, GA, USA; 2 Department of Biochemistry, Emory University School of Medicine, Atlanta, GA, USA; 3 Department of Hematology and Medical Oncology, Emory University, Atlanta, GA, USA; 4 Department of Radiation Oncology, Emory University, Atlanta, GA, USA Dysregulation of DNA repair and maintenance pathways contribute to cancer by increasing levels of DNA damage. Ultimately, unrepaired DNA damage results in the accrual of mutations and genomic instability. A common cause of DNA damage is reactive oxygen species (ROS) that are generated from normal cellular metabolism, inflammation, or through exogenous sources such as radiation. Importantly, tumor cells experience an increased ROS burden from these sources, potentially facilitating more mutations. Base excision repair (BER) is the main pathway for repairing ROS-induced DNA damage, and is initiated by the N- glycosylase proteins. How BER dysregulation contributes to cancer development is largely unknown. To investigate this question, the NTHL1 glycosylase will be used as a model to address the mechanisms of BER regulation and the role of BER in preventing cancer. Possible modes for NTHL1 regulation are compartment-specific localization and/or posttranslational modification(s) in response to ROS-induced DNA damage. The comet assay, immunoprecipitation, mass spectrometry, and cleavage assays are being employed to interrogate NTHL1 regulation in a panel of colon cancer cell lines. BER dysregulation is being tested through biochemical and cellular assays. This project will provide insight into BER mechanisms that contribute to genome integrity, and how BER dysregulation contributes to tumorigenesis.


Kelly Lohr, NS Poster #29 ENHANCED IN VIVO NEUROTRANSMISSION VIA INCREASED VESICULAR TRANSPORT: VMAT2 OVEREXPRESSION IN A MOUSE Kelly M. Lohr1, Alison I. Bernstein1, Kristen A. Stout1, Amy R. Dunn1, Xueliang Fan3, Ellen J. Hess3,4, Hong Yi5, David S. Goldstein6, Thomas S. Guillot1, Ali Salahpour7, Gary W. Miller1,2,3,4

1Dept, of Environmental Health, Rollins School of Public Health, 2Center for Neurodegenerative Diseases, 3Dept. of Pharmacology, 4Dept. of Neurology, 5Robert P. Apkarian Integrated Electron Microscopy Core, Emory University,6 National Institute of Neurological Disorders and Stroke, 7Dept. of Pharmacology and Toxicology, University of Toronto Several therapeutic strategies have been employed to enhance signaling of the monoamine neurotransmitters (dopamine, serotonin, and norepinephrine): administration of precursors to increase synthesis, inhibition of metabolic enzymes to prevent break down, inhibition of plasma membrane transporters to increase the synaptic lifespan, and administration of receptor agonists to directly activate postsynaptic targets. However, many of these interventions have deleterious side effects or lose effectiveness due to desensitization or altered regulation of receptors over time. These undesirable consequences likely occur because these strategies disrupt the temporal nature of regulated neurotransmitter release and uptake. We demonstrate that monoamine neurotransmission can be amplified by increasing vesicular packaging of neurotransmitter via overexpression of the vesicular monoamine transporter 2 (VMAT2). Mice with elevated VMAT2 display increased vesicular filling and size, higher stimulated release and extracellular levels of dopamine, and enhanced spontaneous and amphetamine-induced locomotion. This unexpected plasticity of the vesicle reveals a new therapeutic approach for treating neurological and neuropsychiatric disorders that involve impaired monoaminergic signaling. In support of this, VMAT2-HI mice also show both improved outcomes on measures of anxiety and depressive-like behavior and protection from nigral cell loss by the dopaminergic toxicant MPTP. Jia Meng, MMG Poster #30 FUNCTIONAL ANALYSIS OF THE RSV STRAIN A2001/2-20 ATTACHMENT GLYCOPROTEIN Jia Meng1,2, Michael, G Currier1, 2, Kate L Stokes1, 2, 3, Anne L Hotard1,2, and Martin L Moore1,2 1 Department of Pediatrics, Emory University, 2 Children’s Healthcare of Atlanta, Atlanta, GA, 3 Currently St. Jude Children’s Research Hospital RSV strain A2001/2-20 (2-20) induced more airway epithelial damage and pulmonary mucus expression in BALB/c mice than the laboratory A2 strain [Stokes et al. J. Virol. 2011; 85: 5782.]. We hypothesize that increased fusion caused by the fusion (F) and attachment (G) glycoproteins in infected cells is a determining factor for virulence. We transfected cells with RSV glycoprotein expression plasmids and used a dual-split protein fusion assay to quantify cell-cell fusion. There was no difference in fusion activity between A2 F and 2-20 F. Co-expression of 2-20 F and 2-20 G resulted in higher fusion activity than co-expression of A2 F and A2 G. We generated viruses that either express G and F or F only: A2-A2GA2F, A2-GstopA2F, A2-2-20G2-20F, and A2-Gstop2-20F. We determined infectivity and entry kinetics in BEAS-2B, CHO-K1, and pgsD-677 cell lines. In all these cell lines, A2-A2GA2F had higher infectivity and entry kinetics than A2-2-20G2-20F. Expression of G increased infectivity and entry more for 2-20F than for A2F-containing virus. Furthermore, viruses with both G and F produced larger plaques than viruses with F only. Taken together, the attachment glycoprotein G of RSV increased infectivity, entry kinetics and plaque size, depending on the strain its F protein is from.


Jasmine Miller-Kleinhenz, CB Poster #31 DECREASED EXPRESSION OF IGF-1R AND KI67 USING UPAR-TARGETED THERANOSTIC NANOPARTICLES IN DRUG-RESISTANT TRIPLE-NEGATIVE BREAST CANCER PATIENT-DERIVED XENOGRAFT Jasmine Miller-Kleinhenz1, Hongyu Zhou1, Weiping Qian1, Ruth O’Regan2, Amelia Zelnak2, Toncred Styblo1, Lily Yang1 1Winship Cancer Institute, Department of Surgery, Emory University School of Medicine, Atlanta, GA, 2Winship Cancer Institute, Department of Hematology and Medical Oncology, Emory University School of Medicine, Atlanta, GA Triple-negative breast cancer (TNBC) is a highly heterogeneous and aggressive disease with a poor clinical prognosis. While the primary therapeutic option for TNBC patients is pre-operative neo-adjuvant chemotherapy followed by surgical resection, about 60% of patients develop drug resistance and have a very high incidence of tumor recurrence and poorer prognosis. Given its poor survival and lack of available targeted therapeutics, the development of novel treatment options for chemo-resistant TNBC is critical. Our lab has developed urokinase plasminogen activator receptor (uPAR) targeted theranostic magnetic iron oxide nanoparticles (IONPs) carrying therapeutic agents that are clinically beneficial to TNBC patients such as doxorubicin (Dox). uPAR is an ideal target due to its high expression in malignant tumors including aggressive breast cancer tissues and tumor stromal cells that are enriched in TNBC tissues. Using a patient derived xenograft model of TNBC we investigated uPAR-targeted IONP-Dox effect on tumor growth inhibition. We also investigated the effect the nanoparticle treatment had on relevant biomarkers such as insulin-like growth factor 1 receptor (IGF-1R) and Ki67. Results of our study demonstrated that uPAR-targeted theranostic IONPs decrease expression of IGF-1R and Ki67.These results support further investigation of theranostic nanoparticles as an approach to overcome drug resistance in TNBC. Kevin Morris, BCDB Poster #32 DEFINING THE NEURONAL FUNCTION OF THE POLYADENOSINE RNA-BINDING PROTEIN ZC3H14 Kevin Morris1, Jennifer Rha1, Sara Leung1, Gary Bassell2, Kenneth Moberg2, and Anita Corbett1. Biochemistry, Cell and Developmental Biology Graduate Program, Emory University Laney Graduate School, Dept. of Biochemistry1, Dept. of Cell Biology2, Emory University School of Medicine, Atlanta, GA. RNA-binding proteins are tasked with the major responsibility of regulating the co- and posttranscriptional events of gene expression. Impairments to this process can result in human disease including intellectual disability (formerly termed mental retardation). Recently, our group identified inactivating mutations in the ZC3H14 gene, which encodes an evolutionarily conserved, RNA-binding protein, Zinc finger Cys3His domain-containing protein number 14 (ZC3H14) that lead to non-syndromic intellectual disability. ZC3H14 is an RNA-binding protein that binds with high affinity to polyadenosine RNA. Although ZC3H14 is ubiquitously expressed, ZC3H14 mutant patients only display brain specific phenotypes. While the steady-state localization of ZC3H14 is primarily nuclear, our recent studies reveal that a population of ZC3H14 is present in the axons of primary hippocampal neurons. In addition, ZC3H14 associates with 80S ribosomes. Notably, Drosophila studies of the ZC3H14 fly ortholog, reveal functional interactions with translation regulators. Therefore, we hypothesize that ZC3H14 works cooperatively with RNA regulatory factors to modulate translation. Currently, we are exploiting a recently created ZC3H14 knockout mouse to assess the requirement for ZC3H14 in translation (Aim 1) and to define the spectrum of ZC3H14-interacting RNA regulatory factors (Aim 2). Ultimately, the information gained from the proposed research could provide insight into the molecular role of ZC3H14 and how ZC3H14 affects proper higher order brain function.


Donald Noble, NS Poster #33 CONDITIONING SLOWED RESPIRATORY RATE FOR STRESS REDUCTION IN THE RAT Donald Noble1, William N. Goolsby2, Shawn Hochman1. 1Department of Physiology, 2Department of Cell Biology, Emory University, Atlanta, GA Meditation and yoga techniques involving slowed respiratory rate (SRR) have shown promise in treating a variety of cardiorespiratory and stress-related disorders. SRR exercises improve sympathetic-parasympathetic balance, inducing physical and emotional relaxation. We have developed an animal model of the relaxation response using strobe light as negative reinforcement to operantly condition SRR. During daily two-hour sessions, paired rats (“SRR” and “yoked”) were placed in dual chambers and respiration was continuously and noninvasively monitored using whole body plethysmography. Real-time respiratory rate (RR) was compared to a user-defined target and a closed-loop feedback control system provided time-varying light intensity cues. Only SRR rats had control over the stimulus, while yoked animals received response-independent reinforcement. SRR rats slowed their breathing by 16 breaths/minute over the course of 20 sessions and spent a greater percentage of each session under the target RR, whereas yoked controls did not. Results support the conclusion that feedback-based light intensity control can lead to a selective reduction in respiratory rate. SRR animals also showed a trend toward maintained RR reduction following conditioning, and decreased anxiety-like behavior in an open field. Further studies are aimed at verifying these preliminary findings and identifying mechanisms that link SRR to stress circuit deactivation.

Bridgette Peake, MSP Poster #34 GROWTH DIFFERENTIATION FACTOR 15 AND EPITHELIAL-MESENCHYMAL TRANSITION IN BREAST CANCER Bridgette F. Peake1, Siobhan M. Donnelly2 and Rita Nahta1,2,3,5,*

1Molecular & Systems Pharmacology Program, Graduate Division of Biological and Biomedical Sciences 2Departments of Pharmacology and 3Hematology & Medical Oncology, School of Medicine;; 4Biostatistics, School of Public Health; 5Winship Cancer Institute; Emory University, Atlanta, GA 30322 HER2 amplification occurs in approximately 25% of breast cancers. An additional 15% of breast cancers are called triple-negative (TNBC) due to the absence of estrogen and progesterone receptors and the lack of HER2 overexpression. We previously found that drug-resistant HER2-positive cells express increased levels of the cytokine growth differentiation factor 15 (GDF15). Similarly, TNBCs expressed relatively high levels of GDF15. Immunohistochemical analysis of a tumor tissue array representing almost 600 breast cancer patients indicated significant correlations between high GDF15 staining and ER-negativity, HER2-positivity, high tumor grade, and reduced patient survival. In vivo studies showed increased tumor growth and increased HER2 phosphorylation in GDF15-overexpressing xenografts compared to empty vector control xenografts. Furthermore, GDF15 overexpression induced epithelial-mesenchymal transition (EMT), as demonstrated by changes in cell morphology and increased expression of mesenchymal markers. Importantly, HER2-overexpressing GDF15 stable cell lines showed up-regulation of FOXM1 in comparison to empty vector control cells. We previously published that GDF15 induces MEK signaling, which is an established activator of FOXM1. We hypothesize that GDF15 may promote EMT and tumor growth via MEK-FOXM1 signaling. The current study supports further investigation into the role of GDF15-mediated EMT in both TNBC and HER2-overexpressing breast cancers.


Maylen, Perez Diaz, NS Poster #35 THE HIGHLY SELECTIVE 5-HT2C AGONIST WAY-163909 ELIMINATES METHAMPHETAMINE SELF-ADMINISTRATION IN RHESUS MONKEYS Maylen Perez Diaz1, Kevin S. Murnane1, and Leonard L. Howell1,2 1Yerkes National Primate Research Center, Emory University, Atlanta, GA, USA2Department of Psychiatry and Behavioral Sciences, Emory University, Atlanta, GA, USA Growing evidence indicates that serotonin (5-HT) neurotransmission can modulate the neurochemical and behavioral effects of abused stimulants. Studies have shown that selective antagonists of the 5-HT2A receptor can attenuate cocaine reinstatement, but not self-administration (SA), in nonhuman primates. In contrast, 5-HT2C receptor agonists can partially block cocaine SA in rhesus monkeys. In the present study, we determined if a novel, highly selective 5-HT2C agonist could fully attenuate methamphetamine (METH) SA. Four rhesus macaques with a previous history of drug SA self-administered 0.01mg/kg/injection of METH on a fixed ratio (FR) 20 schedule of reinforcement. For all four subjects, WAY pretreatments decreased response rates during the SA sessions in a dose-dependent manner. In addition, METH SA was fully blocked in all subjects following pretreatment with 1.0mg/kg WAY. These data further inform previous studies by demonstrating that 5-HT2C receptor activation can suppress self-administration of METH as well as cocaine, suggesting effectiveness across psychostimulants in general. The ability of WAY to fully suppress SA may be due to its highly selective agonist activity at the 5-HT2C receptor compared to compounds that have been evaluated previously. The results of this study may inform our search for novel, effective pharmaceutical treatments for drug addiction. Riley Perszyk, MSP Poster #36 A NOVEL CLASS OF POSITIVE ALLOSTERIC NMDA RECEPTOR MODULATORS Riley E. Perszyk1, Brooke M. Katzman2, Dennis C. Liotta2, Stephen F. Traynelis1 1Department of Pharmacology, 2Department of Chemistry N-methyl-D-aspartate (NMDA) receptors are ionotropic ligand-gated ion channels that are activated by glutamate and glycine and mediate a slow component of excitatory synaptic currents. These receptors play important roles in synaptic plasticity, neuronal development, and a number of neurological disorders. The enhancement of NMDA receptor-mediated currents has been hypothesized to confer cognitive enhancement and could be potentially beneficial in schizophrenia. Testing these hypotheses has been limited due to a lack of potent and effective pharmacological compounds. During the development of a negative allosteric modulator series, identified by screening efforts, we discovered a series of substitutions on the molecule that converted these negative modulators into positive modulators. Using two-electrode voltage clamp recordings of NMDA receptors expressed in Xenopus oocytes, we found that potentiators in this class can enhance responses 150-250% with EC50 values ranging from 2 to 10μM. Co-application of positive and negative modulators suggests that the positive modulators can compete with the negative modulators. This raises the possibility that these modulators act at a single site. This class of compounds could be utilized to assess the validity of enhancing NMDA receptor activity as a potentially beneficial strategy in neuropsychiatric disorders and could lead to novel therapeutic strategies.


Katherine Reding, NS Poster #37 CHRONIC PSYCHOSOCIAL STRESS AND ESTRADIOL ALTER INTRINSIC FUNCTIONAL CONNECTIVITY IN RHESUS MACAQUES. Katherine Reding1, David S. Grayson2, Oscar Miranda-Dominguez3, Siddharth Ray3, Martin Styner4, Mark E. Wilson1, Donna Toufexis5, & Damien A. Fair3, Mar Sanchez1,6

1 Division of Developmental and Cognitive Neuroscience, Yerkes National Primate Research Center, Emory University, 2Center for Neuroscience, University of California - Davis, 3 Departments of Behavioral Neuroscience, Psychiatry, and Advanced Imaging Research Center, Oregon Health and Science University, 4Department of Psychiatry, University of North Carolina, Chapel Hill, 5Department of Psychology, University of Vermont, 6Department of Psychiatry & Behavioral Sciences, Emory University The ovarian hormone estradiol (E2) has activational effects on female rhesus macaque prosociaI, motivational, and sexual behavior that are attenuated during exposure to chronic psychosocial stress. However, the mechanisms by which this attenuation occurs are unknown. In humans, analysis of neural activation at rest using functional magnetic resonance imaging (fMRI) suggests that acute stress can increase intrinsic functional connectivity (iFC) between the amygdala and midline cortical structures, including the medial prefrontal cortex (mPFC). Studies on adult male rhesus macaques suggest that social subordination can lead to changes in iFC between the superior temporal sulcus (STS) and the PFC. Furthermore, increased iFC within amygdalar-cortical networks and between hemispheres is seen following exogenous E2 replacement therapy. Exposure to E2 in combination with chronic stress may exacerbate stress-related increases in functional connectivity. To investigate this hypothesis, we studied ovariectomized adult female rhesus macaques exposed to high levels of chronic social subordination stress, both with and without E2 replacement. Data show greater STS-mPFC iFC in Subordinate compared to Alpha females, which result from chronic exposure to psychosocial stress. Furthermore, greater STS-mPFC iFC in Subordinate females after E2 treatment suggests that E2 exacerbates the effects of subordination stress on iFC. Jennifer Rha, BCDB Poster #38 MUTATION OF A ZINC FINGER POLYADENOSINE RNA BINDING PROTEIN, ZC3H14, CAUSES AUTOSOMAL RECESSIVE INTELLECTUAL DISABILITY Jennifer Rha, Sara Leung, Gary Bassell, Anita Corbett. Emory University School of Medicine, Atlanta, GA. Patients with intellectual disability (ID) suffer from significantly subaverage intellectual function (IQ≤70), which impinges on quality of life. We have recently identified the first gene encoding a polyadenosine RNA binding protein, ZC3H14 (Zinc finger CysCysCysHis domain-containing protein 14), which is mutated in inherited nonsyndromic autosomal recessive intellectual disability (NS-ARID). This finding uncovers the molecular basis for disease in these patients and provides strong evidence that ZC3H14 is essential for proper brain function. Studies of ZC3H14 orthologs in budding yeast and Drosophila provide insight into the role of this protein in post-transcriptional regulation of gene expression and key evidence for its critical role in neurons. Despite these studies, functional characterization of ZC3H14 in vertebrates is crucial for understanding both normal brain function and the molecular mechanism underlying ID in these patients. We have developed a conditional ZC3H14 knockout mouse utilizing the Cre/loxP system to extend our studies to vertebrate ZC3H14 and address our hypothesis that ZC3H14 is required for proper expression of target mRNAs that are critical for neuronal function. These ZC3H14 knockout mice provide us with an optimal model to study the requirement for ZC3H14 in higher order brain function.


Samuel Rose, NS Poster #39 NEUROCHEMICAL AND BEHAVIORAL DYSFUNCTION IN A MOUSE MODEL OF DOPA-RESPONSIVE DYSTONIA Samuel J. Rose1,2, Xin Y. Yu2, H. A. Jinnah MD/PhD3,4,5, Ellen J. Hess PhD2,3

1 Neuroscience Graduate Program, Departments of 2 Pharmacology, 3 Neurology, 4 Human Genetics, and 5

Pediatrics

Dystonia is characterized by involuntary muscle contractions that cause debilitating twisting movements and postures. Although the pathogenesis of dystonia is not understood, reduced dopamine neurotransmission is consistently observed across many different forms of dystonia, including dopa-responsive dystonia (DRD). DRD is cause by mutations in genes integral to dopamine synthesis, including tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine synthesis. Therefore, to better understand the role of dopamine in dystonia, we have created a knockin mouse bearing the DRD-causing TH mutation 1160C>A; Q381K (DRD knockin mice). TH activity is reduced by ~90% in vivo in these mice, causing a dramatic reduction in dopamine concentration. Behaviorally, DRD knockin mice exhibit abnormal movements reminiscent of human dystonia. These movements are exacerbated by a D1-like dopamine receptor antagonist, suggestive of a mechanism for dystonia involving the D1 dopamine receptor. DRD knockin mice also exhibit abnormal circadian regulation of locomotor activity, where DRD knockin mice are more active than controls at the start of the dark cycle. However, by the start of the light cycle, DRD knockin mice exhibit catalepsy. These studies establish DRD knockin mice as a useful model for examining dopaminergic mechanisms underlying dystonia. Supported by PHS NS059645.

Chelsea Ruppersburg, BCDB Poster #40

A NOVEL ROLE FOR THE CHLORIDE CHANNEL ANO1 (TMEM16A) IN CILIOGENESIS: THE ANO1 NIMBUS

Chelsey Chandler Ruppersburg & H. Criss Hartzell. Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322 USA. Many cells possess a single, non-motile, primary cilium that is thought to be a sensory transducer akin to a cellular antenna. Although sensory transduction requires ion channels, relatively little is known about ion channels in the primary cilium. We show here for the first time the presence of a Cl- channel, the Ca2+-activated Cl- channel ANO1, in the primary cilium. Prior to cilia extension, ANO1 is organized into a torus-shaped structure along with other ciliary proteins including the small GTPases CDC42 and Arl13b, the exocyst complex component Sec6, and acetylated α-tubulin and γ-tubulin. This structure we call the “nimbus” forms an interface between the microtubule cytoskeleton of the nascent cilium and the surrounding cortical actin cytoskeleton. During ciliogenesis, the nimbus disassembles and ciliary components, including ANO1, move into the cilium. Blocking ANO1 currents pharmacologically or knockdown of the ANO1 channel disrupts ciliogenesis. Our data support a model where the nimbus provides a scaffold for staging of ciliary components and cilia assembly and that Cl- transport by ANO1 is required for the genesis or maintenance of cilia. Our data here may provide a mechanistic foundation for interpreting the roles of ANO1 in development and cancer.


Emily Rye, BCDB Poster #41

A NOVEL MECHANISM FOR THE GLUCOCORTICOID RECEPTOR IN INFLAMMATORY GENE REGULATION

Emily Rye1, Will Hudson1, Kendall Nettles2, Eric Ortlund1 1Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road, Atlanta, GA 303222Department of Cancer Biology, The Scripps Research Institute, Jupiter, Florida 33458 Inflammation is a multifaceted process required to heal injuries and fight infections. However, chronic inflammation underlies various debilitating diseases including asthma, rheumatoid arthritis, and heart disease. Glucocorticoids (GC) are the most widely prescribed anti-inflammatory drug. GCs bind and activate the glucocorticoid receptor (GR), inducing its translocation to the nucleus where it can stimulate or inhibit gene expression. To activate transcription, GR binds to glucocorticoid response elements to drive the transcription of metabolic and stress response genes. Conversely, GR can repress transcription by two distinct mechanisms, either DNA-dependent or independent. The first mechanism involves direct binding of GR to negative glucocorticoid response elements. The DNA-independent mechanism occurs when GR represses other pro-inflammatory transcription factors, most notably NF-κB and AP-1, through protein-protein interactions only. This mechanism, known as “tethering”, is currently the only accepted model for GR-mediated repression of inflammation. However, recent discoveries regarding DNA-dependent GR-mediated transrepression reveal that the mutations used to develop the tethering hypothesis are inadequate. In addition, fluorescence polarization binding assays and x-ray crystallographic structures show GR bound to inflammatory gene promoters at AP-1 binding sites, suggesting that GR-mediated repression of inflammatory genes could occur through an alternative mechanism, involving direct DNA binding. Alessandra Salgueiro, CB Poster #42

MECHANISTIC STUDIES OF VIMENTIN INTERMEDIATE FILAMENTS USING AN INDUCIBLE TRANSFECTION SYSTEM

Alessandra M. Salgueiro1, Alessandra M. Salgueiro, Lauren S. Havel1, Adam I. Marcus1. 1Winship Cancer Institute, Department of Hematology and Oncology, Emory University, Atlanta, GA 30322 Vimentin, an intermediate filament, is a marker of epithelial to mesenchymal transition (EMT), a mechanism in cancer metastasis. Gain of vimentin expression correlates with EMT, but little is known about the mechanisms by which vimentin function and dynamics contribute to EMT. We demonstrated that vimentin regulates focal adhesion kinase (FAK) to stabilize focal adhesions essential for motility. Other groups have shown that vimentin dynamics are mediated by serine phosphorylation sites in the head and tail domains. We created an inducible GFP:vimentin system to recapitulate gain of vimentin expression observed during EMT and use this model to determine how vimentin serine phosphorylation mediates invasion, and focal adhesion stability. HEK293 cells (vim +/+) were used to optimize the transfection system. 16HBECs (human bronchial epithelial cells) (vim null) were used for mechanistic studies of vimentin regulation of FAK and analysis of serine phosphorylation sites. The four serine sites studied were mutated to either unphosphorylatable alanine or phosphomimetic aspartic acid. Our studies in HEK293s demonstrate that wild type GFP:vimentin mimics the morphology of endogenous vimentin. Similar experiments in 16HBECs show different morphologies among GFP:vimentin phospho-mutants. These data demonstrate that mechanistic studies utilizing the phospho-mutants will provide better understanding of vimentin dynamics focal adhesion regulation.


Eduardo Sanabria-Figueroa, MSP Poster #43

TARGETING IGF-1R AND HER2 IN HER2-OVEREXPRESSING BREAST CANCER CELLS: ROLE IN CELL VIABILITY, GROWTH, INVASION, MORPHOLOGY, AND ELUCIDATION OF CROSSTALK MECHANISM

Eduardo Sanabria-Figueroa1, Siobhan M. Donnelly2, Elisavet Paplomata3, and Rita Nahta1,2,3,4. 1Molecular & Systems Pharmacology Program, Graduate Division of Biological and Biomedical Sciences; 2Department of Pharmacology, School of Medicine; 3Hematology & Medical Oncology, School of Medicine; 4Winship Cancer Institute; Emory University, Atlanta, GA, 30322. HER2 is overexpressed in ~25% of metastatic breast cancers in association with poor prognosis. Trastuzumab, a monoclonal antibody that targets the extracellular domain of HER2, is the leading therapy for this group of breast cancer patients. However, trastuzumab resistance often emerges, sometimes within a year of treatment, allowing disease progression. Previous studies have indicated that IGF-1R signaling contributes to trastuzumab resistance by crosstalk to HER2. Consistent with this idea, inhibition of IGF-1R restored trastuzumab sensitivity in resistant cells. Here, we show that IGF-1R co-localized with HER2 in HER2-positive cell lines. Inhibition of IGF-1R or inhibition of the intracellular kinase SRC reduced IGF-I-induced phosphorylation of HER2. We also found that combinatorial pharmacological inhibition of IGF-1R and HER2 decreased cell viability, anchorage-independent growth, and invasion of trastuzumab-resistant HER2-positive breast cancer cells. Our data also suggest that IGF-1R and HER2 co-inhibition might induce mesenchymal to epithelial transition. These results support the presence of IGF-1R/HER2 crosstalk and suggest a potential role for the intracellular kinase SRC in this crosstalk. Further studies will be performed to determine if co-targeting IGF-1R and HER2 will rescue the epithelial-mesenchymal transition and invasiveness of trastuzumab-resistant HER2-positive breast cancers.

Karl Schmidt, NS Poster #44

LOCUS COERULEUS NORADRENERGIC ACTIVATION REINFORCES OPERANT BEHAVIOR

Karl T. Schmidt1, 2, Chelsea A. Koller1, Elena M. Vazey3, Caroline E. Bass4, Ilana B. Witten5, Karl Deisseroth6, Gary Aston- Jones3, David Weinshenker1, 2

1Emory University, Department of Human Genetics 2 Emory University, Neuroscience Graduate Program 3Medical University of South Carolina, Department of Neurosciences, Department of Psychiatry and Behavioral Sciences 4SUNY Buffalo, Department of Pharmacology and Toxicology 5Princeton University, Department of Psychology 6Stanford University In order to probe the brain’s reward circuitry, early studies used electrodes to stimulate various regions contingent upon an operant response (e.g. lever press), a technique called intracranial self-stimulation (ICSS). In the 1970s, some studies found that electrical stimulation of the noradrenergic locus coeruleus (LC) and its axons comprising the dorsal noradrenergic bundle maintained operant behaviors, while others obtained the opposite result. Interpretation of the initial ICSS findings must be made cautiously as electrical stimulation is limited by significant caveats such as electrode placement in the small nucleus and the activation of non-noradrenergic sources including fibers of passage, innervating terminals from other brain regions, and nearby cells with various chemical phenotypes. To overcome these technical obstacles and conclusively determine whether noradrenergic activity can function as a reinforcer, we took an optogenetic approach. Using 2 different expression systems (a Cre-dependent AAV injected into tyrosine hydroxylase::Cre transgenic rats and a Phox2b-dependent PRSx8 lentivirus), we have expressed channelrhodopsin selectively in LC noradrenergic neurons. Response contingent activation of these neurons with light maintains behavior comparable to identical stimulation parameters of optogenetic activation of VTA dopamine neurons. These data provide new, convincing evidence implicating this noradrenergic nucleus as a component of the brain’s reward system.


Marc Schureck, BCDB Poster #45 MECHANISM OF HIGB-MEDIATED RIBOSOME-DEPENDENT MRNA DEGRADATION Marc A. Schureck12, Tatsuya Maehigashi1, Jack Dunkle1, Stacey Miles1, Jhomar Marquez1, Ajchareeya Ruangprasert1, and Christine M. Dunham1 1Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322 2 Biochemistry, Cell and Developmental Biology Graduate Program, Emory University, Atlanta, GA 30322 Proteus vulgaris (P. vulgaris) is a major cause of urinary tract bacterial infections. P. vulgaris can grow on medical equipment, such as catheters, in an antibiotic-resistant state known as biofilms. In biofilms, bacteria are highly adhesive and are protected from antibiotics by a self-produced extracellular matrix of nucleic acid, protein and polysaccharides. My goal is to elucidate the molecular mechanism of the ribosome-dependent RNase family of toxins, which play an important, but unknown, role in biofilm formation. P. vulgaris Host inhibition of growth B (HigB) is a member of this family and cleaves ribosome-bound messenger RNA (mRNA). HigB is unique because it cleaves adenosine-rich mRNA sequences rather than a three-base mRNA codon, the unit by which translation factors and other better studied toxins, read mRNA. Towards the elucidation of this novel mRNA decoding mechanism, we solved the X-ray crystal structure of HigB bound to mRNA on the ribosome. This structure reveals that HigB binds mRNA in the A site of ribosome and highly conserved HigB amino acids and ribosomal RNA appear important for recognition of mRNA. Our biochemical studies of various adenosine-rich sequences cleaved by HigB have lead to our current model of how HigB reads mRNA codon-independently. Jill Seladi-Schulman, MMG Poster #46 SPHERICAL INFLUENZA VIRUSES HAVE A FITNESS ADVANTAGE IN EMBRYONATED EGGS, WHILE FILAMENT-PRODUCING STRAINS ARE SELECTED IN VIVO Jill Seladi-Schulman, John Steel, and Anice C. Lowen Emory University, Atlanta, GA, USA Influenza viruses assume two morphologies: filamentous and spherical. Filaments are observed in low-passage isolates while spheres are seen in strains grown extensively in laboratory substrates. Previous studies have shown that filamentous morphology is lost upon passage in eggs in favor of spherical morphology. The fact that filaments are maintained in nature but not in the laboratory raises the question of whether they provide an advantage within the infected host. We examined the effect of serial adaptation to eggs, MDCK cells, and guinea pigs on morphology. Two filamentous strains, A/Netherlands/602/2009(H1N1) and A/Georgia/M5081/2012(H1N1), were passaged ten times in eggs and MDCK cells, substrates believed to select for spheres. The spherical strain A/Puerto Rico/8/1934(H1N1) was passaged twelve times in guinea pigs, aiming to select for filamentous virions. We found that an exclusively spherical morphology is not required for improved growth in laboratory substrates. We did, however, identify two point mutations in the matrix protein of egg passage 10 isolates that confer a spherical morphology and increased growth in eggs, indicating that spherical viruses have a fitness advantage in eggs. Passage in guinea pigs resulted in the selection of filament-forming viruses. These findings suggest a functional role for filaments in the infected host.


Shardule Shah, IMP Poster #47 p38 IS A NEGATIVE REGULATOR OF THE BORTEZOMIB-INDUCED HEAT SHOCK RESPONSE IN MULTIPLE MYELOMA Shardule P. Shah1,2, Vikas A. Gupta2, MD, PhD, Shannon M. Matulis2, PhD, Ajay K. Nooka2, MD, MPH, Sagar Lonial2, MD, and Lawrence H. Boise1,2, PhD 1Laney Graduate School, Emory University, Atlanta, GA, 2Hematology and Medical Oncology and Winship Cancer Institute, Emory University, Atlanta, GA Studies characterizing HSP regulation upon proteasome inhibition (PI) in multiple myeloma are limited. The work displayed here sheds light on the regulation mechanism. We show that HSF1 silencing and the proteasome inhibitor, bortezomib (VelcadeTM), have an additive death effect in two myeloma cell lines, MM1.s and KMS18. HSF1 phosphorylation indicates activity and here we demonstrate PI-induced phosphorylation in those lines and two patient samples. There have been no published studies regarding kinases that phosphorylate HSF1 upon PI. To begin to identify them, we treated cells with JNK, MEK, and p38 inhibitors to detect changes in bortezomib-induced HSF1 phosphorylation. JNK and MEK inhibition yielded little change, but p38 inhibition ablated HSF1 constitutive phosphorylation and resulted in an increase in MM1.s and KMS18 inducible phosphorylation. HSF1 activation leads to increased expression of downstream HSP genes via binding of HSF1 to heat shock elements. We show that expression of HSP genes previously shown to be both induced upon PI and HSF1-regulated, were increased when bortezomib is combined with p38 inhibition. Therefore, p38 may be a direct negative regulator of the bortezomib-induced heat shock response. Ultimately, the results shown here demonstrate a regulatory mechanism of the bortezomib-induced heat shock response in multiple myeloma. Jonathon Sia, IMP Poster #48 MYCOBACTERIUM TUBERCULOSIS IMPAIRS DENDRITIC CELL FUNCTIONS THROUGH THE SERINE HYDROLASE HIP1 Ranjna Madan-Lala1, Jonathan Kevin Sia1,2, Rebecca King1, Toidi Adekambi1, Bali Pulendran1,3, and Jyothi Rengarajan1,4

1Emory Vaccine Center, Emory University, Atlanta, GA, USA2Immunology and Molecular Pathogenesis Graduate Program, Graduate Division of Biological and Biomedical Sciences, Emory University, Atlanta, GA, USA3Division of Pathology, Emory University School of Medicine, Atlanta, GA, USA4Division of Infectious Diseases, Department of Medicine, Emory University School of Medicine, Atlanta, GA, USA Mycobacterium tuberculosis (Mtb) is a highly successful human pathogen that primarily resides in host phagocytes and interferes with their functions. While multiple strategies used by Mtb to modulate macrophage responses have been discovered, interactions between Mtb and DCs are less well understood. DCs are the primary antigen presenting cells (APCs) of the immune system and play a central role in linking innate and adaptive immune responses against microbial pathogens. We show that Mtb impairs DC cytokine secretion and maturation through the cell envelope-associated serine hydrolase Hip1 via MyD88- and TLR2/9-dependent pathways. These impaired processes led to suboptimal T-cell responses, as Mtb infected DCs induced weaker Th1 and Th17 responses from antigen specific CD4 T-cells. Furthermore, we show that priming of antigen specific T-cells in-vivo is affected during the early stages of the immune response in a low dose aerosol mouse model of TB. Overall, these data show that Mtb impairs DC functions and modulates the nature of antigen-specific T cell responses, with important implications for vaccination strategies.


Kristen Stout, MSP Poster #49 THE SYNAPTIC VESICLE GLYCOPROTEIN 2C (SV2C) MEDIATES DOPAMINE VESICULAR STORAGE AND RELEASE: IMPLICATIONS FOR PARKINSON’S DISEASE AND ADDICTION Kristen A. Stout1, Amy R. Dunn1, Kelly M. Lohr1, Yingjie Li1, Thomas S. Guillot1, and Gary W. Miller1,2 1Department of Environmental Health, 2Center for Neurodegenerative Disease, Emory University, Atlanta, GA Synaptic vesicle glycoproteins localize to synaptic vesicles and play a role in neurotransmission. Within this family, the synaptic vesicle glycoprotein 2C (SV2C) localizes to regions associated with dopamine signaling, such as the basal ganglia. Additionally, recent epidemiological data identified SV2C as mediating the neuroprotective effects of nicotine in Parkinson’s disease, with minor SV2C allele frequencies reversing nicotine neuroprotection. Thus, we hypothesize that SV2C contributes to dopamine storage and release. To test this hypothesis, we generated SV2C knockout (SV2C-KO) mice. These mice show decreased dopamine release in the dorsal striatum in comparison with WT littermate controls. Release within the nucleus accumbens was also measured. Vesicular storage capacity in SV2C-KO mice is also reduced compared with WT mice. These data suggest that SV2C plays an important role in the synaptic cycling of dopamine. Further investigation is necessary to determine the contribution of SV2C to both neurodegenerative disease and addiction. E. Shannon Torres, GMB Poster #50 UNDERSTANDING THE TRANSCRIPTIONAL REGULATORY MECHANISMS OF THE HISTONE VARIANT, H2A.Z, THROUGH ITS INTERACTIONS WITH CHROMATIN REMODELERS E. Shannon Torres1 and Roger B. Deal2 1Genetics and Molecular Biology Program in the Graduate Division of Biological and Biomedical Sciences,

Emory University, Atlanta, GA, 2Department of Biology, Emory University, Atlanta, GA

Differentiating cells acquire a stable transcriptional program necessary for proper development and homeostasis, which is mediated through epigenetic processes, such as incorporation of histone variants into nucleosomes. The histone H2A variant, H2A.Z, is involved in many genomic processes, including transcriptional regulation. However, the mechanism through which it regulates transcription is currently unclear. H2A.Z is necessary for transcription of the FLOWERING LOCUS C (FLC) gene in Arabidopsis, but in the absence of transcriptional repressors, such as Brahma, H2A.Z is no longer required for transcription of the gene. This project focuses on elucidating the mechanism by which Brahma antagonizes H2A.Z function to infer the role of H2A.Z in transcriptional regulation. We will determine whether Brahma antagonizes H2A.Z-mediated activation at FLC and similarly regulated loci through affecting nucleosome positioning or occupancy and whether H2A.Z localization across the gene is impacted without Brahma. Additionally, we are conducting a forward genetic suppressor screen to identify mutants that alleviate the need for H2A.Z incorporation into nucleosomes for FLC activation. Currently, three candidate lines have been identified as suppressor mutants. Future work to identify and characterize these genes will provide a deeper understanding of H2A.Z function and broaden our understanding of transcriptional regulation through chromatin remodeling.


MaKendra Umstead, CB Poster #51 CHARACTERIZING A NOVEL AURORA KINASE A PROTEIN-PROTEIN INTERACTION IN CANCER MaKendra Umstead1,2, Cau Pham2, Zenggang Li2, Andrei Ivanov2, Yuhong Du2, Haian Fu2,3 1Cancer Biology Graduate Program, 2Department of Pharmacology, 3Winship Cancer Institute Cancer development is driven by genomic changes that affect normal protein function and induce a malignant phenotype. Genomic data from The Cancer Genome Atlas (TCGA) provide opportunities to identifying the mechanism by which aberrantly regulated proteins, often clustered in protein signaling pathways, contribute to cancer. Therapeutic strategies can then be devised to target key protein hubs within the signaling network and perturb oncogenic signaling while preserving tumor suppressive functions. Proteins involved in many protein-protein interactions (PPIs) serve as signaling hubs. To identify these hubs, we conducted high-throughput TR-FRET and PCA screening using a library of genes identified by TCGA to be altered in cancer. One PPI hub revealed by the screen was Aurora Kinase A (AURKA). AURKA normally facilitates mitosis; however, its overexpression in cancer suggests additional functions that remain to be elucidated. For example, one novel binding partner of AURKA discovered by our screen was Ras. To investigate the AURKA/Ras interaction, validation was completed using pull-downs, the structural properties assessed by gene deletion analysis, and the impact of the interaction on cell signaling analyzed by western blotting for downstream signaling. Ultimately, understanding the role of AURKA in cellular signaling will provide new opportunities to develop targeted therapies for cancer. Virginia Vachon, MMG Poster #52 OPTIMAL ACTIVATION OF HUMAN 2’-5’ OLIGOADENYLATE SYNTHETASE-1 BY NON-CODING VIRAL AND CELLULAR RNAS IS DEPENDENT UPON 3’ POLY-URIDYLATION Virginia K. Vachon1,2, Brenda Calderon1 and Graeme L. Conn1 1Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road NE, Atlanta GA 30322, USA. 2Microbiology and Molecular Genetics (MMG) Program, Graduate Division of Biological and Biomedical Sciences, Emory University. The human 2’-5’ oligoadenylate synthetase-1 (hOAS1) is an important part of innate cellular immunity. Upon binding viral dsRNA, hOAS1 synthesizes 2’-5’ linked oligoadenylates that activate RNAseL, thereby halting replication through cleavage of viral mRNA or RNA genome. Both Adenovirus and Epstein Barr virus produce non-coding RNAs (ncRNA), VA RNAI and EBER-1respectively, that inhibit the anti-viral dsRNA binding protein PKR, ensuring continued translation of viral proteins. These Polymerase III (Pol III) RNA transcripts are produced at high levels in later stages of infection, and are critical for replication of these viruses. While both of these non-coding RNAs inhibit PKR’s anti-viral activity, they each activate rather than inhibit hOAS1. In addition to these viral RNAs, a cellular Pol III transcript, non-coding RNA 886 (nc886), has also been shown to inhibit PKR. Curiously, this cellular RNA is also a potent activator of hOAS1. We found that presence of a 3’ poly U tail in each of these contexts is critical for optimal activity against hOAS1. Activity of a model duplex RNA is also augmented by addition of a 3’ poly U tail, however a single U is sufficient for maximal enhancement. Work is currently underway to determine the structural basis for these observations.


Kellie Vinal, MMG Poster #53 PATHOGEN-DERIVED 16S RRNA METHYLTRANSFERASE NPMA: MECHANISMS OF TARGET RECOGNITION AND ENZYMATIC ACTIVITY Kellie Vinal1,2, Pooja Desai1, Natalia Zelinskaya1, John Wang1 and Graeme L. Conn1 1Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road NE, Atlanta GA 30322, USA. 2Microbiology and Molecular Genetics (MMG) Program, Graduate Division of Biological and Biomedical Sciences, Emory University. Antibiotic resistance remains a pervasive problem in the treatment of bacterial infections due to the continual evolution of resistance mechanisms in bacteria, as well as horizontal gene transfer between bacterial species. The 16S rRNA methyltransferase NpmA specifically methylates nucleoside adenosine 1408 (A1408) in the aminoglycoside binding site of the bacterial ribosome, conferring resistance to clinically relevant aminoglycosides such as tobramycin and gentamicin. As the first A1408 16S rRNA methyltransferase isolated from a human pathogen, it is critical to characterize the mechanism by which NpmA recognizes and methylates its substrate as a platform for inhibitor development to combat this resistance determinant. Based on data from rRNA probing experiments, residues predicted to interact with the 30S subunit were identified and mutated. Kanamycin minimum inhibitory concentration (MIC) and 30S binding assays with these mutants were used to pinpoint critical residues and characterize their role in conferring resistance. Additionally, the crystal structure of the 30S subunit-NpmA complex was recently solved by our collaborators. This new structural insight, in conjunction with functional data from the mutagenesis studies, identified key residues for 30S binding and A1408 target base flipping, and allowed us to develop a molecular model for the mechanism NpmA uses to methylate its substrate. Jadiel Wasson, BCDB Poster #54 THE FUNCTION OF LSD1 IN EPIGENETIC REPROGRAMMING AT FERTILIZATION Jadiel A. Wasson1, Ashley L. King1, and David J. Katz1

1Department of Cell Biology, Emory University Epigenetic modifications are implicated in the maintenance and regulation of transcriptional programs. One modification, H3K4me2 (dimethylation of lysine 4 on histone 3) is suggested to function as a type of transcriptional memory by marking genes that were previously transcribed, establishing the cell’s transcriptional program, and facilitating the transmission of this transcriptional program through cell division. However, during cell fate change, H3K4me2 must be removed. At fertilization, the highly differentiated sperm and egg, which have specific transcriptional programs, fuse and subsequently form the totipotent zygote. This massive change in cell fate at fertilization implies there is a need to erase the gametic transcriptional program in order to re-establish totipotency. The histone demethylase LSD1 specifically erases H3K4me2 and is maternally deposited into the zygote at fertilization. We hypothesized that LSD1 plays a maternal role in epigenetic reprogramming at fertilization. To test our hypothesis, we have generated two conditional deletion mouse models to specifically delete LSD1 in mouse oocytes. We find that maternal deletion of LSD1 leads to a 1-2 cell arrest of progeny and a mis-expression of the oocyte transcriptional program. In addition, by using a hypomorphic Cre allele, we find maternal loss of LSD1 leads to long range epigenetic defects in progeny.


David Weir, CB Poster #55 PROCASPASE-3 REGULATES FIBRONECTIN SECRETION AND INFLUENCES ADHESION, MIGRATION AND SURVIVAL INDEPENDENT OF CATALYTIC FUNCTION Matthew Brentnall1,2, David B. Weir1, Anthony Rongvaux3, Adam I. Marcus1, and Lawrence H. Boise1 1Departments of Hematology and Medical Oncology and Cell Biology, Winship Cancer Institute of Emory University, Atlanta, GA, 2Sheila and David Fuente Graduate Program in Cancer Biology, University of Miami Miller School of Medicine, Miami, FL, 3Department of Immunobiology, Yale School of Medicine, New Haven, CT Apoptosis has profound effects on development and homeostasis. Inactivation of apoptosis can lead to developmental disorders, carcinogenesis, autoimmunity, neurodegenerative disorders and defects in wound healing. During apoptosis, caspase-3 (the main effector caspase) is activated downstream of Bax activation, mitochondrial outer membrane permeablization and cytochrome c release. Previous work has shown that caspase-3-deficient mouse embryonic fibroblasts are resistant to mitochondria-mediated cell death and exhibit delayed Bax activation, mitochondrial outer membrane permeablization and cytochrome c release. There are two ways to interpret this finding: caspase-3 is part of a feedback mechanism on mitochondria or, alternatively, that caspase-3 has an independent non-apoptotic function that has an influence upstream of mitochondrial events in apoptosis. Our laboratory has recently demonstrated that procaspase-3 functions as a regulator of cell adhesion, migration and survival. This function is distinct and independent from the function of activated caspase-3 in apoptosis. We plan to exploit mouse embryonic fibroblast cell lines to test our overall hypothesis that fibronectin secretion is required for procaspase-3 regulation of cell adhesion, migration and survival. Our long-term goal is to develop a comprehensive understanding of the molecular mechanisms regulating cell adhesion, migration and survival, which will yield novel insight into the processes of carcinogenesis and metastasis. Amanda Wendt, NHS Poster #56 UNDERSTANDING HEALTHCARE WORKERS’ PERCEPTIONS OF IRON AND FOLIC ACID SUPPLEMENTATION IN BIHAR, INDIA Amanda Wendt1, Melissa Young2, Amy Webb Girard1,2, Reynaldo Martorell1,2 1Nutrition and Health Sciences, Laney Graduate School, Emory University, GA 30322 2Hubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, GA 30322 High anemia prevalence and low iron and folic acid (IFA) consumption among pregnant women in Bihar is an important public health issue. Surveys find 30% of Bihari women indicate receiving IFA and 23% report IFA counseling during their last pregnancy. To address this, we conducted twenty in-depth interviews and focus group discussions with three key types of frontline healthcare workers (FLWs) to explore their knowledge of anemia and IFA counseling. FLWs attributed anemia to factors including poor diet, close child spacing, high parity, and lack of rest, which they perceived to stem from poverty, lack of awareness, low priority of women’s status, and family tension. Many felt anemia was not a significant issue, estimating its prevalence as 0-20% (compared to survey estimates of 60%). FLWs tended to restrict IFA consumption to the 4th-8th month. In conclusion, FLWs had high knowledge of anemia causes but did not view it as a common issue. Further, their views of proper consumption windows may prevent added health benefits of IFA consumption during the 9th month and lactation. Efforts to allow IFA intake over a longer timeframe may increase opportunities for women to receive IFA and be a key step to increasing IFA consumption.


Scott Wilkinson, CB Poster #57

LKB1 REGULATES CELL POLARITY AND MOTILITY IN A FARNESYLATION-DEPENDENT MANNER Scott Wilkinson1, Adam I. Marcus2 1Graduate Program in Cancer Biology, Emory University, Atlanta, GA, 2Hematology and Medical Oncology, Emory University, Atlanta, GA

The tumor suppressor LKB1 is a serine/threonine kinase that serves as a master regulator of cell polarity, and is mutated in 30% of non-small cell lung cancer tumors. LKB1 contains a C-terminal (CTD) farnesylation motif to allow for plasma membrane insertion and interaction with other membrane-bound proteins. Using confocal microscopy, we show that the LKB1 CTD regulates cell polarity and LKB1:actin association at the leading edge in a farnesylation-dependent manner. We first created GFP constructs of both LKB1 wildtype and CTD, as well as C430S farnesylation motif mutants of each. These constructs were transfected into LKB1-null cells. Upon confluency, a scratch-wound assay was performed and 6 hours later cells were fixed and permeabilized. We probed with a GM130 antibody (Golgi marker, used to identify polarization) and stained with phalloidin to visualize actin filaments. Using confocal microscopy, we identified LKB1 farnesylation as critical for regulating cell polarity and leading edge actin association. Importantly, this is the first report of kinase farnesylation in maintaining cell polarity. These data show that LKB1 farnesylation is critical for regulating cell polarity and motility through its actin association, potentially leading to novel insights into mechanisms of LKB1 function in regulating cell polarity and motility. Kathryn Williams, BCDB Poster #58

REPRESSION OF GAP-43 EXPRESSION BY THE MRNA BINDING PROTEIN HNRNP-Q1: INVESTIGATING A LOCAL MECHANISM IN NEURONS K.R. Williams1, L. Xing1, G.J. Bassell1

1Department of Cell Biology, Emory University School of Medicine, Atlanta, GA mRNA binding proteins function to post-transcriptionally regulate gene expression, a key process that is required for proper neuronal development. The mRNA binding protein hnRNP-Q1 is an ideal candidate regulator of mRNA transport and/or local translation given that it is a component of transport mRNP granules and regulates mRNA translation. We have identified GAP-43 mRNA as a new target of hnRNP-Q1. GAP-43 protein is highly enriched in axonal growth cones where it functions to regulate actin dynamics and ultimately axonal growth and/or guidance. Additionally, GAP-43 mRNA is localized to developing axons and growth cones. Therefore, we hypothesize that hnRNP-Q1 regulates the axonal localization and/or local translation of GAP-43 mRNA as a mechanism to enrich GAP-43 protein and affect axon growth. Using RNAi methods to knockdown hnRNP-Q1 in Neuro2a cells, our results suggest that hnRNP-Q1 suppresses the expression of GAP-43 protein but does not affect steady state GAP-43 mRNA expression. Work in progress is to knockdown hnRNP-Q1 in primary cortical neurons to assess possible impairments in GAP-43 mRNA localization and local translation as well as axonal phenotypes that may result from impaired localization of GAP-43 mRNA. hnRNP-Q1-mediated regulation of GAP-43 may be a key mechanism for regulating neuronal development.


Myra Woodworth-Hobbs, NHS Poster #59

DOCOSAHEXAENOIC ACID COUNTERACTS PALMITATE-INDUCED PROTEOLYTIC SIGNALING IN MYOTUBES Myra E. Woodworth-Hobbs1,2¸ Matthew B. Hudson2, Jill A. Rahnert2, Bin Zheng2, Harold Franch2, S. Russ Price2,3. 1Nutrition and Health Sciences, Graduate Division of Biological and Biomedical Sciences, 2Medicine/Nephrology, Emory University, Atlanta, GA; 3Atlanta VAMC, Decatur, GA Dyslipidemia is a comorbidity common to illnesses like diabetes which contributes to debilitating muscle atrophy. The saturated fatty acid palmitate (PA) induces insulin resistance and disrupts protein metabolism in cultured myotubes, whereas the omega-3 fatty acid docosahexaenoic acid (DHA) has beneficial metabolic effects. We evaluated the effects of PA and DHA on atrophy-related signaling in skeletal muscle, with the hypothesis that DHA prevents PA-induced myotube atrophy by counteracting the stimulatory effects of PA on protein degradation. C2C12 myotubes were treated with 500uM PA and/or 100uM DHA for up to 28h. PA increased the rate of protein degradation, while co-treatment with DHA completely prevented the response. Akt inhibits the FoxO3 transcription factors which regulate “atrogene” expression. PA reduced the activation state of Akt as well as increased the level of nuclear FoxO3 protein while decreasing cytosolic FoxO3. PA also increased the mRNA levels of two FoxO3 atrogene targets, the E3 ubiquitin ligase atrogin-1/MAFbx and the autophagy mediator Bnip3. DHA reversed the effects of PA on Akt, FoxO3, and both atrogenes. These data indicate that PA induces myotube atrophy by inducing components of the ubiquitin-proteasome and autophagic proteolytic systems and that DHA counters the catabolic effects of PA by improving Akt signaling.


ACKNOWLEDGEMENTS

The GDBBS Division Student Advisory Council (DSAC) would first like to express our sincerest gratitude to all of the participants-students, faculty, and post-docs- who made this 11th Annual DSAC Student Research Symposium possible.

Our deepest gratitude goes to the Graduate Student Council (GSC) for funding the event. In addition we want to thank Monica Taylor for her continued efforts to help make the symposium a success. We would also like to thank the GDBBS office and program staff for their assistance. Lastly we thank the GDBBS Director, Keith Wilkinson for continuing to support our efforts to do more than sponsor mixers.

Again, thank you, and we look forward to seeing you next year as we convene again to make the symposium a signature event for all of us in GDBBS.

Documents

The 11th Annual DSAC Student Research Symposium Friday ...biomed.emory.edu/news-events/documents/2014-DSAC... · Session II: Immunity to Infection 10:30 am 10:30 Maria Georgieva (MMG)