welcome to the tenth annual division student · 2013. 4. 17. · the tenth annual dsac student...

Welcome to the Tenth Annual Division Student Advisory Council (DSAC) Research Symposium!

I know you will enjoy viewing the presentations of the interesting and exciting research of our talented GDBBS students. This year we will listen to 22 talks and view 68 posters. Judges will determine who among our presenters will receive special honor. The participation of the research community is vital to the success of this endeavor. This is an opportunity for the presenters to gain valuable experience and receive feedback on their work. All of us in the audience will learn new things that will help enrich our own research endeavors

Let me extend my heartfelt thanks to the 2012-13 DSAC members. This event is spearheaded and hosted by this group of motivated students. They are:

Julie Fritz, BCDB Jasmine Miller-Kleinhenz, CB Nita Sharma-Jegadeesh, GMB Gopi Mohan, IMP Elizabeth Ohneck, MMG

Erica Bizzell, MMG Sherry Adesina, MSP Sindhu Lakkur, NHS Katherine Reding, NS Amanda Pierce, PBEE

I am sure that you will join me in encouraging and supporting all of the students who have worked so hard to showcase their work today.

^x|à{ WA j|Ä~|ÇáÉÇ Keith D. Wilkinson, PhD Director, GDBBS Professor and Vice Chair, Biochemistry

THE TENTH ANNUAL DSAC STUDENT RESEARCH SYMPOSIUM [2]

The Tenth Annual DSAC Student Research Symposium Friday, January 11

Room 110, James B. Williams Medical Education Building

8:30 – 9:00 Breakfast - 1st Floor Lobby

Session I Animal Models 9:00 am

9:00-Abraham Mathai (NS) Cortical Inputs To The Subthalamic Nucleus Degenerate In Experimental Parkinsonism

9:15-Damon Lamb (NS)

Biophysical Modeling Of Leech Heart Motor Neurons

Session II DNA And Gene Expression 9:30 am

9:30-Nick Bauer (BCDB) Dynamic Localization Of Base Excision Repair Proteins Revealed By Quantitative Subcellular

Compartmentalization Analysis (Q-SCAN)

9:45-Callie Wigington (BCDB) The Novel Poly(A) RNA-Binding Protein, ZC3H14, Regulates A Key Subunit Of ATP Synthase In Breast

Cancer Cells

10:00-Brooke Weckselblatt (GMB) Large-Scale Sequence Analysis Of Translocation Breakpoint Junctions

10:15-Jennifer Gerfen (GMB)

Telomere Position Effect AT Human Chromosome Ends

10:30-10:45 Break

Session III Life Of A Cell 10:45 am

10:45-Brian Gaudette (IMP) Bcl-XL Protects From UPR-Associated Apoptosis During Plasma Cell Differentiation

11:00-Bree A. Szostek (MMG)

Determining The Role Of FlhD2C2, Negative Feedback, And The RCS Phosphorelay In The Regulation Of The Swarming Inhibitor disA In Proteus mirabilis.

11:15-Mary Puckett (MSP)

Interception Of Cell Death Signaling By The ASK1/IKK Signaling Axis


11:30-Rebecca Meyer (NS) GPR37 And GPR37l1 Are Receptors For The Neuroprotective And Glioprotective Factors Prosaptide And

Prosaposin

11:45-Matthew Randolph (BCDB) Satellite Cell Behavior In Pharyngeal Muscles

12:00 -1:30 pm Poster Sessions & Lunch is Served (Lobby Area of the SOM Bldg.)

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

Session IV Influence Of Experience And Environment 1:45 pm

1:45-Myra Woodworth-Hobbs (NHS) Docosahexaenoic Acid Antagonizes Palmitate-Induced Stimulation Of Protein Degradation In Myotubes

2:00-Kathryn Shepard (NS)

Noradrenergic Modulation Of Experience-Dependent Plasticity During Auditory Cortical Development

2:15-Teresa Madsen (NS) Delta Phase Modulation Of Gamma Oscillations In The BLA and mPFC During Fear Learning And

Expression In Freely Moving Rats

2:30-2:45 Break

Session V Microbe Virulence And Clearance 2:45 pm

2:45-Katherine Houser (IMP) Seasonal Trivalent Inactivated Influenza Vaccine Does Not Protect Against Newly Emerging Variants Of

Influenza A(H3N2)v Virus In Ferrets

3:00-Jill Seladi-Schulman (MMG) Effect Of Serial Passage On Influenza Virus Morphology And Growth

3:15-Joshua R. Shak (PBEE)

Streptococcus Pneumoniae Virulence Factor Pneumolysin Is Essential For Assembly Of Pneumococcal Biofilms

3:30-Tim Sampson (MMG)

A CRISPR-CAS System Mediates Bacterial Innate Immune Evasion and Virulence

3:45-4:00pm Break


Session VI Receptors And Signaling 4:00 pm

4:00-Sara Freeman (NS) The Primary Binding Site For Oxytocin In The Macaque Forebrain Is The Vasopresin 1A Receptor:

Evidence From In Vivo And In Vitro Localization Techniques

4:15-Andrew Bankston (BCDB) Molecular Mechanisms And Essential Roles Of CDK5 Activation In Oligodendroglia Differentiation And

Myelin Lesion Repair

4:30-Eric Armstrong (MSP) Determining The Molecular Mechanism Underlying Ligand-Dependent Activation Of FABP5

4:45-Kalynda Gonzales (NS)

GABAergic Regulation of Striatal Cholinergic Interneurons: Synaptic Inputs from Parvalbumin-containing Interneurons

5:00-5:15pm Break

Keynote Address

5:15-5:45 pm

Dr. Stephen T. Warren, PhD William Patterson Timmie Professor and Chair, Department of Human Genetics, Emory University School of Medicine also holds the Charles H. Candler Chair in Human Genetics and is Professor of

Biochemistry and Pediatrics at Emory.

6:00pm Reception and Awards Presentation

Images

Front Cover: Image Contest Winners 1st Place: Brooke Napier, MMG 2nd Place: Paul Evans, NS 3rd Place: Jill Seladi-Schulman, MMG

: Andrew Swanson, NS

Page 24: Jodi Godfrey, NS

Page 62: Jacob Billings, NS


Session I

Animal Models

9:00 am


Abraham Mathai, NS 9:00 am

CORTICAL INPUTS TO THE SUBTHALAMIC NUCLEUS DEGENERATE IN EXPERIMENTAL PARKINSONISM Abraham Mathai1,2, Yuxian Ma1,2, Jean-Francois Paré1,2, Susan Jenkins1,2, Thomas Wichmann1,2,3, Yoland Smith1,2,4 1Yerkes National Primate Research Center, 2Morris K. Udall Center of Excellence for Parkinson’s Disease Research, 3Department of Neurology, Emory University, Atlanta, GA.

Parkinson’s disease (PD) is a basal ganglia disorder associated with partial degeneration of the corticostriatal glutamatergic system, but it is unclear whether the integrity of the cortical inputs to the subthalamic nucleus (STN) is similarly affected. We addressed this issue through a comparative analysis of the synaptic microcircuitry of glutamatergic cortical afferents to the STN, and an assessment of the effects of electrical stimulation of the corticosubthalamic pathway on the activity of pallidal neurons (downstream from the STN) in normal and parkinsonian monkeys. Using immunostaining of the vesicular glutamate transporter type-1, a specific marker for cortical afferents, we found that the prevalence of cortical inputs to STN neurons, that terminate mainly on their distal dendrites, is reduced by up to 40% in parkinsonian animals. Electrophysiological recordings showed that the proportion of pallidal neurons that respond to electrical stimulation of the corticosubthalamic system is 35-50% lower in parkinsonian monkeys than in normal animals. Thus, parkinsonism is associated with a reduced extent and altered functional impact of the corticosubthalamic projection in nonhuman primates. It remains unclear whether the loss of cortical inputs to the striatum and STN is a primary pathological event or a compensatory phenomenon of PD in humans.

Damon Lamb, NS 9:15 am

BIOPHYSICAL MODELING OF LEECH HEART MOTOR NEURONS D. G. Lamb, R. L. Calabrese Emory University, Atlanta, GA

Motor neurons are often disregarded as mere followers - simply integrating their inputs and firing in a clearly defined manner, except where they play a direct role within a central pattern generator (CPG). The leech heart motor neurons appear to play a meaningful role in the production of their distinct intersegmental pattern, even though the dominant factor is the pattern of input from the CPG and the synaptic weights. We now leverage a multiobjective evolutionary algorithm (MOEA) to explore a parameter space and identify sets of 'acceptable' models. Our core model is a 7 compartment Hodgkin-Huxley style biophysical model of a mid-body motor neuron containing known voltage-gated and calcium-gated currents. This model is parameterized to allow the maximal conductances of active currents to vary. We use our unique set of individual animal input-output patterns, which each contain the complete temporal pattern (spike times) of the input onto ipsilateral midbody leech heart motor neurons, synaptic strength profile, and the output of the motor neurons recorded in a single individual. Acceptable models, defined as models which are within tolerable bounds around target values across all fitness functions, achieved traces which strikingly similar to intracellular recordings in the living system.


Session II

DNA And Gene Expression

9:30 am


Nicholas C. Bauer, BCDB 9:30 am

DYNAMIC LOCALIZATION OF BASE EXCISION REPAIR PROTEINS REVEALEDBY QUANTITATIVE SUBCELLULAR COMPARTMENTALIZATION ANALYSIS (Q-SCAN) Nicholas C. Bauer1,, Anita H. Corbett1,2, Paul W. Doetsch1,2,3,4 1Department of Biochemistry, 2Winship Cancer Institute, 3Department of Radiation Oncology, 4Department of Hematology and Medical Oncology, Emory University School of Medicine, Atlanta, GA.

The highly-conserved base excision repair (BER) pathway is responsible for repairing the most common forms of DNA damage including oxidation, deamination, and base hydrolysis. Defects in various components of 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. Saccharomyces cerevisiae Ntg1, a DNA N-glycosylase/AP lyase that initiates the repair of a wide variety of oxidative base damage products, dynamically localizes to both the nucleus and mitochondria in response to compartmental DNA damage, demonstrating that localization is an important component of its regulation. Other BER proteins (Ung1, Ogg1, Apn1) also localize to both genome-containing organelles. 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. Our results validate this method as a reliable and specific quantification of dynamic localization of BER proteins.

Callie Wigington (BCDB) 9:45 am

THE NOVEL POLY(A) RNA-BINDING PROTEIN, ZC3H14, REGULATES A KEY SUBUNIT OF ATP SYNTHASE IN BREAST CANCER CELLS CP Wigington1, PM Vertino2, AH Corbett1 Department of Biochemistry1 and Winship Cancer Institute2; Emory University, Atlanta, GA.

Altered gene expression is one critical point of dysregulation in cancer. Key post-transcriptional regulatory events are mediated by RNA-binding proteins, one class of which is comprised of poly(A) binding proteins (Pabs). A novel CCCH zinc finger Pab, ZC3H14, has been linked to the estrogen receptor status of breast tumors. Although the function of ZC3H14 is unknown, the budding yeast counterpart, Nab2, modulates polyadenylation and mRNA export from the nucleus, functions consistent with a role for ZC3H14 in post-transcriptional regulation. In a genome-wide study seeking target transcripts of ZC3H14, we identified the ATP Synthase Fo Subunit C (ATP5G1) as significantly decreased upon ZC3H14 knockdown in a breast cancer cell line. ATP Synthase is the central enzyme in oxidative phosphorylation and is responsible for the majority of ATP production in mammalian cells. Our data suggests that ZC3H14 specifically affects the steady-state mRNA level of ATP5G1 and not other ATP Synthase subunits, suggesting specificity consistent with recent structural studies of RNA recognition. We rescue the decrease in ATP5G1 by re-expression of ZC3H14. As ATP5G1 is a rate-limiting component in Fo Subunit C assembly and ultimately ATP Synthase activity, we hypothesize that ZC3H14 plays a role in regulating cellular energy levels in cancer cells.


Brooke Weckselblatt, GMB 10:00 am

LARGE-SCALE SEQUENCE ANALYSIS OF TRANSLOCATION BREAKPOINT JUNCTIONS Brooke Weckselblatt, Katie Rudd Department of Human Genetics, Emory University School of Medicine

Translocation is one of the most common structural chromosome abnormalities observed in humans. Germline unbalanced translocations result in partial monosomy and partial trisomy of many genes, leading to intellectual disability and birth defects. We have analyzed chromosome breakpoints from 55 human subjects with unique unbalanced translocations using a combination of array comparative genome hybridization (CGH), targeted sequence capture, and next-generation sequencing. This group of 55 rearrangements also includes translocations between three, four, or five nonhomologous chromosomes. Breakpoints with complex junctions involving multiple chromosomes may be indicative of chromosome shattering, a recently described process that plays a role in germline and somatic chromosome rearrangements. Sequence analysis of breakpoint junctions allows inference of the type of DNA repair at translocation breakpoints and can reveal a more complex rearrangement structure than expected from array CGH alone. Consistent with recent publications that demonstrate the variability in breakpoints, we have sequenced 21 junctions that have either no microhomology, a few basepairs of microhomology, hundreds of bases of homologous sequence, or insertions. Sequence analysis of the remaining translocation junctions is ongoing. Results from this large-scale study of patient breakpoint sequences will pave the way to more complete models for chromosome rearrangement by translocation.

Jennifer Gerfen, GMB 10:15 am

TELOMERE POSITION EFFECT AT HUMAN CHROMOSOME ENDS Jennifer Gerfen, Heather Mason-Suares, M. Katharine Rudd Department of Human Genetics, Emory University School of Medicine, Atlanta, GA

Silencing of gene expression due to proximity to a heterochromatic telomere is a well known epigenetic process in model organisms. This telomere position effect (TPE) may be measured at artificial chromosome ends with a reporter gene inserted adjacent to a newly truncated telomere. However, investigating TPE at normal human chromosome ends is difficult due to hundreds of kilobases (kb) of repetitive DNA adjacent to the telomere. We take advantage of human cell lines derived from patients with subtelomeric deletions to measure TPE in naturally occurring repositioned telomeres. We studied TPE in seven cell lines with terminal deletions using ChIP-seq and RNA-seq. In one cell line with an ~5.5-Megabase terminal deletion, we identified an ~400-kb region of histone H3 lysine 9 tri-methylation (H3K9me3) enrichment extending from the breakpoint. RNA-seq data from a gene within the H3K9me3-enriched region revealed allelic skewing consistent with gene silencing in cis with the deletion. A gene outside the H3K9me3-enriched region exhibits equal expression from both alleles. These data suggest that telomeric heterochromatin spreading can silence genes near terminal deletion breakpoints. Further study of patient-derived deletions is ongoing to evaluate human TPE on other chromosomes.


Session III

Life Of A Cell

10:45 am


Brian Gaudette, IMP 10:45 am

Bcl-xL PROTECTS FROM UPR-ASSOCIATED APOPTOSIS DURING PLASMA CELL DIFFERENTIATION. Brian T. Gaudette1,3, Neal N. Iwakoshi, Ph.D.2.3 and Lawrence H. Boise, Ph.D.1.3

1Departments of Hematology and Medical Oncology, Cell Biology and Winship Cancer Institute, 2Department of Surgery and Emory Transplant Center, Emory University, Atlanta, GA. Understanding factors that control plasma cell survival is important for the development of therapeutic approaches to diseases including multiple myeloma and autoimmune disorders. Plasma cell differentiation requires activation of the unfolded protein response, a set of signaling pathways in which prolonged activation leads to apoptosis in normal cells. Plasma cells activate this pathway indefinitely suggesting compensatory survival signals are activated during differentiation. We employed two models of plasma cell differentiation, the murine B cell lymphoma cell line, Bcl1 and primary murine B cells both of which can be induced to differentiate with cytokine and LPS. We showed in Bcl1 cells that LPS driven differentiation protects against ER stress-induced death due to tunicamycin treatment. We showed an increase in Bcl-xL expression in differentiating cells accompanied by a decrease in Mcl-1 or Bcl-2 in Bcl1 cells and primary B cells respectively. We used the Bcl-xL/Bcl-2 inhibitor, ABT-737 to show that LPS-mediated protection from tunicamycin-induced cell death was Bcl-xL-dependent. Importantly, we showed that during differentiation, these cells became Bcl-xL-dependent as ABT-737 sensitized cells to differentiation-induced death. Finally, we used co-immunoprecipitation to show that differentiation was accompanied by change in Bim binding from an Mcl-1/Bcl-2 pattern to a Bcl-xL dominated pattern in differentiating cells. Bree A. Szostek (MMG) 11:00 am

DETERMINING THE ROLE OF FlhD2C2, NEGATIVE FEEDBACK AND THE RCS PHOSPHORELAY IN THE REGULATION OF THE SWARMING INHIBITOR disA IN Proteus mirabilis. Bree A. Szostek1 and Philip N. Rather1,2 1Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA, 2Bacterial Pathogenesis Section, Atlanta VA Medical Center, Decatur, GA

Proteus mirabilis is a gram-negative bacillus that exists as a swimmer cell in culture and undergoes a differentiation process to form an elongated, hyperflagellated swarmer cell when in contact with an agar plate. We have further characterized the regulation of the previously identified inhibitor of swarming, disA. We have identified the transcriptional start site of disA as a Thymidine 70 base-pairs upstream of the open reading frame as well as the -10 promoter element. Furthermore, we found that FlhD2C2, the swarming master regulator, does not have a role in disA regulation and have data that suggests disA may be subject to feedback inhibition. Transposon mutagenesis identified two mutations that affected disA expession: one in cpsF, a capsular polysaccharide glycosyl transferase, decreased disA expression; and a second in umoB, an Rcs Phosphorelay regulator, increased disA expression. The involvement of the Rcs Phosphorelay in disA regulation was further investigated by constructing mutants in rcsC and rcsB, the relay’s sensor kinase and response regulator. Our data demonstrate that the regulatory role of UmoB on disA is dependent upon the Rcs Phosphorelay. Taken together, these data suggest at least two independent pathways are responsible for the regulation of the swarming inhibitor, disA.


Mary Puckett, MSP 11:15 am

INTERCEPTION OF CELL DEATH SIGNALING BY THE ASK1/IKK SIGNALING AXIS Mary Puckett2, Erinn Goldman1,2, Lisa Cockrell2, Fadlo Khuri1, and Haian Fu1,2 1Winship Cancer Institute, 2Department of Pharmacology, Emory University, Atlanta, GA

ASK1 is a key mediator of ROS signaling. Improper ASK1 regulation has been implicated in multiple diseases, including cancer. Conversely, nuclear factor kB (NfkB) protects cells against ROS in part through impairing JNK activation, a target of ASK1. However, the mechanism of this crosstalk remains unclear. This work has identified a direct, non-genetic method of cell death inhibition by a regulator of NfkB, the inhibitor of kB kinase (IKK).

Previous evidence indicates IKK phosphorylates ASK1 Ser967, impairing ASK1 activity. To further elucidate the relationship between these kinases, the interaction interface was identified and functional implications were studied. Both endogenous and overexpressed kinases interacted in cells by immunoprecipitation. Furthermore, their direct interaction was confirmed through time resolved fluorescence resonance energy transfer (TR-FRET). The ASK1/IKK interaction interface was dissected through iterative deletional mutagenesis. ASK1 activity was assessed by measuring H2O2 induced apoptosis, neurite outgrowth of ASK1-inducible cells, and phosphorylation of ASK1 Ser967 under several IKK states. IKK prevented stress-induced dephosphorylation and inhibited ASK1 mediated apoptosis.

These data suggest existence of an IKK/ASK1 signaling node that negatively regulates apoptosis in response to pro-survival conditions. Characterization of this interaction could lead to development of targeted agents for ASK1 and IKK-related diseases.

Rebecca Meyer, NS 11:30 am

GPR37 AND GPR37L1 ARE RECEPTORS FOR THE NEUROPROTECTIVE AND GLIOPROTECTIVE FACTORS PROSAPTIDE AND PROSAPOSIN Rebecca C. Meyer, Michelle M. Giddens & Randy A. Hall Pharmacology Department, Emory University, Atlanta, GA

Orphan G protein-coupled receptors (oGPCRs) have the potential to be novel therapeutic targets, and identification of endogenous ligands for orphan receptors is a vital step toward targeting these receptors. Here, we present evidence that the oGPCRs GPR37 and GPR37L1 can be activated by prosaptide, a neuroprotective peptide derived from the precursor protein prosaposin, which was identified as a neurotrophic and neuroprotective factor more than a decade ago.

We found that nanomolar concentrations of prosaptide bind to and activate GPR37 and GPR3L1 to strongly promote receptor internalization as well as ERK phosphorylation and reductions in cAMP levels in a pertussis toxin-sensitive manner. We also found that prosaptide induces ERK phosphorylation and cell survival in cortical astrocytes through stimulation of endogenous GPR37. Finally, we found that the endogenous ligand, prosaposin, is also able to stimulate signaling and cell survival through activation of these receptors. This discovery is of potential clinical interest since prosaptide and prosaposin have been shown to be neuroprotective in animal models of Parkinson’s disease and ischemia, as well as promoting nerve remyelination following injury. By identifying receptors for these neuroprotective factors, we open the door for novel therapeutic targets to treat these diseases.


Matthew Randolph, BCDB 11:45 am

SATELLITE CELL BEHAVIOR IN PHARYNGEAL MUSCLES Matthew Randolph1, Kristen Carroll1, Cherry Luo2, Justin Ho1, Alan Sokoloff2, Anita Corbett3 and Grace Pavlath1 Departments of 1Pharmacology, 2Physiology and 3Biochemistry, Emory University, Atlanta, GA

Pharyngeal muscles are essential for proper swallowing and prevention of food/water aspiration into the lungs yet the cellular and molecular mechanisms that control pharyngeal muscle growth and maintenance are unknown. Pharyngeal dysfunction results in swallowing difficulties, which is termed dysphagia. Dysphagia is observed in 20% of individuals over the age of 60 as well as in several muscular dystrophies. Given the critical importance of the pharyngeal muscles, surprisingly little is known about the role of adult muscle stem cells in pharyngeal muscle biology. Adult muscle stem cells, satellite cells, of the extraocular and masseter muscles differ in myogenic abilities from the commonly studied satellite cells of limb muscles, which suggests that other craniofacial muscles may also have unique myogenic properties. Our results demonstrate that pharyngeal satellite cells in mice undergo chronic proliferation, differentiation and fusion in the absence of induced injury. These phenotypes cannot be attributed to overt myofiber damage or myofiber type. These unusual myogenic properties may make pharyngeal muscles more vulnerable to disease and aging. By understanding the cellular and molecular regulatory controls that drive chronic myogenesis of pharyngeal satellite cells, we will gain valuable insight into the pathologic mechanisms that may underlie dysphagia in age and disease.


Session IV

Influence Of Experience And Environment

1:45 pm


Myra Woodworth-Hobbs, NHS 1:45 pm

DOCOSAHEXAENOIC ACID ANTAGONIZES PALMITATE-INDUCED STIMULATION OF PROTEIN DEGRADATION IN MYOTUBES Myra Woodworth-Hobbs, Matthew Hudson, Bin Zheng, Sara Zoromsky, Harold Franch, S. Russ Price Medicine/Nephrology, Emory University, Atlanta, GA

Muscle atrophy is a debilitating consequence of many chronic diseases, some of which are characterized by accumulation of lipids in skeletal muscle. High levels of saturated fatty acids (i.e. palmitic acid, PA) cause dysregulated protein metabolism in cultured myotubes, whereas the omega-3 fatty acid docosahexaenoic acid (DHA) has beneficial metabolic effects. We hypothesized that DHA prevents PA-induced muscle atrophy by counteracting its effects on protein degradation. To test our hypothesis, we studied C2C12 myotubes treated with 500uM PA and/or 100uM DHA. PA significantly increases the rate of protein degradation, and the response is reversed by co-treatment with DHA. The protein kinase Akt is a key regulator of protein balance and inhibits the FoxO3 transcription factor. PA reduces the ratio of phospho-to-total Akt and increases the mRNA and total and nuclear protein levels of FoxO3. It also increases the levels of mRNA encoding three FoxO3 targets, the MuRF-1 and atrogin-1/MAFbx E3 ubiquitin ligases and the autophagy mediator Bnip3. DHA attenuates the effects of PA on Akt, FoxO3, and all three atrogenes mRNAs. These data indicate that PA induces protein degradation by inducing the ubiquitin-proteasome and autophagic proteolytic systems and that DHA counters the catabolic effects of PA by improving Akt signaling.

Kathryn Shepard, NS 2:00 pm

NORADRENERGIC MODULATION OF EXPERIENCE-DEPENDENT PLASTICITY DURING AUDITORY CORTICAL DEVELOPMENT Kathryn N. Shepard , L. Cameron Liles1, David Weinshenker1 and Robert C. Liu2

1Department of Human Genetics, 2Department of Biology, Emory University, Atlanta, GA

During circumscribed “sensitive periods” in postnatal development, passive sensory experience can play a profound role in shaping neural responses in sensory cortex. For instance, the proportion of auditory cortex that is responsive to a particular frequency can be increased by exposure to that frequency during the sensitive period. Though it has been studied extensively, the mechanisms responsible for permitting plasticity during this time are not yet known. Here, we investigated whether norepinephrine (NE), a neuromodulator that has been linked to sensitive period plasticity in visual cortex, is required for plasticity during auditory cortical development. Mapping multi-unit electrophysiological recordings across auditory cortex, we compared auditory responses in the NE-deficient dopamine beta-hydroxylase knockout mouse (Dbh -/-) and its NE-competent counterpart (Dbh +/-). In colony-reared animals, best frequencies, tuning bandwidths, firing rates, and gross tonotopic organization were comparable between animal groups. However, in a subset of animals that were reared in the presence of pure tone repetitions, Dbh -/- mice failed to show the expansion in the cortical representation of the exposure frequency that normally follows this manipulation. If this preliminary result is borne out in a larger population, it would implicate NE as a critical modulator of sensory plasticity during development.


Teresa Madsen, NS 2:15 pm

DELTA PHASE MODULATION OF GAMMA OSCILLATIONS IN THE BLA AND mPFC DURING FEAR LEARNING AND EXPRESSION IN FREELY MOVING RATS Madsen TE2, Rainnie DG1,2 Psychiatry and Behavioral Sciences, Emory University, Atlanta, GA; 2 Behavioral Neuroscience and Psychiatric Disorders, Yerkes National Primate Research Center, Atlanta, GA In rats undergoing classical fear conditioning, we have previously demonstrated that a significant increase in power and coherence between local field potentials (LFPs) in the basolateral amygdala (BLA) and medial prefrontal cortex (mPFC) occurs in a tightly tuned delta (1-4 Hz) frequency band in response to either the unconditioned stimulus (foot shock) or the conditioned stimulus (tone). Moreover, spectral analysis further revealed a strong increase in gamma (45-65 Hz) power in LFPs in both the BLA and mPFC, especially immediately after the shock during fear acquisition and to a lesser extent at the end of the cue presentation during fear recall. In the hippocampus, critical memory functions have been associated with modulation of gamma amplitude by theta (4-10 Hz) phase. Furthermore, a 4 Hz phase modulation of gamma amplitude has recently been reported in the prefrontal cortex of rats performing a working memory task. Hence, we hypothesized that gamma may be similarly modulated by delta in the rat BLA and mPFC during fear learning and expression. Here, we report that the amplitude of the gamma oscillations is indeed dramatically modulated by the phase of the delta oscillations, specifically in the mPFC during and after the presentation of conditioned tones.


Session V

Microbe Virulence And Clearance

2:45 pm


Katherine Houser, IMP 2:45 pm

SEASONAL TRIVALENT INACTIVATED INFLUENZA VACCINE DOES NOT PROTECT AGAINST NEWLY EMERGING VARIANTS OF INFLUENZA A(H3N2)v VIRUS IN FERRETS Katherine V. Houser, Jacqueline M. Katz, Terrence M. Tumpey

Influenza Division, National Center for Immunization and Respiratory Diseases, Center for Disease Control and Prevention, Atlanta, GA

The increase in the number of human cases of influenza A H3N2 variant virus [A(H3N2)v] in the summer months of 2012 has raised public health concerns and highlights the need to assess whether contemporary seasonal influenza virus vaccination provides some measure of cross-protection against A(H3N2)v virus. In this study, we evaluated whether the 2011-2012 trivalent inactivated influenza vaccine (TIV) protected ferrets against viral challenge with A/Indiana/08/2011, an A(H3N2)v isolated in late 2011 that is antigenically and genetically similar to H3N2v viruses from the 2012 outbreak. Ferrets were vaccinated with the seasonal TIV and then challenged with either a homologous seasonal H3N2 virus (A/Perth/16/2009), or A(H3N2)v virus. Our data demonstrate that immunization with a seasonal TIV protected ferrets against homologous H3N2 virus challenge by significantly reducing upper respiratory tract virus replication, but provided minimal to no protection against A(H3N2)v challenge. The complete absence of cross-reactive hemagglutination-inhibition (HI) titers prior to challenge against A(H3N2)v is consistent with the poor cross-protection observed among TIV-immune animals. This data is supported by recent serological studies that demonstrated that contemporary TIV failed to substantially augment cross-reactive antibody to A(H3N2)v virus in the population. For optimal protection of the population an A(H3N2)v–specific vaccine is needed.

Jill Seladii-Schulman, MMG 3:00 pm

EFFECT OF SERIAL PASSAGE ON INFLUENZA VIRUS MORPHOLOGY AND GROWTH Jill Seladi-Schulman, John Steel, and Anice C. Lowen Emory University, Atlanta, GA

It has long been observed that influenza virus comes in two predominant morphologies: filamentous and spherical. The filamentous morphology is seen in low-passage isolates while the spherical morphology is seen in strains that have undergone repeated passage in laboratory substrates. The fact that the filamentous morphology is maintained in nature but not in the laboratory raises the question of whether the filamentous morphology confers a selective advantage in the host that is not necessary for growth in laboratory substrates. We have performed serial passage experiments to assess the effect of passage on both virion morphology and growth. Two filamentous strains, A/Netherlands/602/2009 (H1N1) and A/Georgia/M5081/2012 (H1N1), were passaged in eggs and MDCK cells, the laboratory substrates believed to select for spheres. Conversely, the spherical strain A/Puerto Rico/8/1934 (H1N1) was passaged a total of 12 times in guinea pigs. After passaging, the morphology and growth of the passaged isolates were compared to those of the wild-type. We found that, in laboratory substrates, a spherical morphology was not required for increased growth. However, in guinea pigs, the emergence of filamentous virions as well as the improved growth of the passaged strain suggested a functional role for filaments in the animal host.


Joshua R. Shak , PBEE 3:15 pm

STREPTOCOCCUS PNEUMONIAE VIRULENCE FACTOR PNEUMOLYSIN IS ESSENTIAL FOR ASSEMBLY OF PNEUMOCOCCAL BIOFILMS Joshua R. Shak1, Herbert P. Ludewick2, Fuminori Sakai2, Lesley McGee3, Keith P. Klugman2,3, Jorge E. Vidal2 1Molecules to Mankind Program, 2Hubert Department of Global Health, Emory's Rollins School of Public Health, 3Respiratory Diseases Branch, Division of Bacterial Disease, National Center for Immunization and Respiratory Diseases, Centers for Disease Control and Prevention Streptococcus pneumoniae (the pneumococcus) is an important bacterial pathogen, responsible for millions of deaths each year. The formation of multicellular biofilms is important for colonization of the nasopharynx and pathogenesis of pneumonia and otitis media. Pneumolysin (Ply) is the primary toxin of the pneumococcus and we have previously demonstrated that its expression is down-regulated in a quorum-sensing knockout strain that is deficient in biofilm formation. Therefore, we conducted a study to determine whether Ply has a direct role in pneumococcal biofilm formation. By staining biofilms with an anti-Ply fluorescent antibody, we demonstrated that the presence of Ply in biofilms peaks with maximal biofilm formation and that Ply is expressed in the earliest aggregates of biofilm cells. Confocal microscopy revealed Ply homogeneously distributed throughout pneumococcal biofilms, apparently linking chains of pneumococci. A pneumolysin-deficient mutant (D39ply) produced significantly less biofilms at early time points when compared to wild type D39. Using hemolytic activity assays we found that strains expressing Ply, but naturally deficient in hemolytic activity, were still capable of forming biofilms. Hemolytic activity of biofilm cells proportionally decreased with maturation of the biofilm structure. Altogether, these experiments demonstrate that a well-known toxin in S. pneumoniae has a previously unrecognized role in biofilm formation, distinct from its hemolytic function. Tim Sampson, MMG 3:30 pm

A CRISPR-CAS SYSTEM MEDIATES BACTERIAL INNATE IMMUNE EVASION AND VIRULENCE

Timothy R. Sampson and David S. Weiss

Division of Infectious Diseases, Department of Medicine, Emory University, Atlanta, GA

CRISPR (Clustered Regularly Interspaced Palindromic Repeats)-CAS (CRISPR-associated) systems are a recently described bacterial defense against invading foreign nucleic acids derived from bacteriophages or exogenous plasmids. These systems utilize an array of small CRISPR RNAs (crRNAs) consisting of repetitive sequences flanking unique spacers to recognize their targets, and conserved CAS proteins to mediate target degradation. It has been unclear if these systems have additional functions in bacterial physiology. Here, we demonstrate that the CAS protein Cas9 of Francisella novicida utilizes a unique, small, CRISPR-CAS-associated RNA(scaRNA) to mediate the repression of an endogenous transcript encoding a bacterial lipoprotein (BLP). As BLPs trigger a proinflammatory innate immune response aimed at combating pathogens, CRISPR-CAS mediated repression of BLP is critical for F. novicida to dampen the host inflammatory response and promote virulence. Since Cas9 proteins are highly enriched in pathogenic and commensal bacteria, our work suggests that CRISPR-CAS-mediated gene regulation may broadly contribute to the interaction of such bacteria with eukaryotic hosts.


Session VI

Receptors And Signaling

4:00 pm


Sara Freeman, NS 4:00 pm

THE PRIMARY BINDING SITE FOR OXYTOCIN IN THE MACAQUE FOREBRAIN IS THE VASOPRESIN 1A RECEPTOR: EVIDENCE FROM IN VIVO AND IN VITRO LOCALIZATION TECHNIQUES Sara Freeman1,2,4,5, Aaron Smith2,3,5, Kiyoshi Inoue2,4,5, Mark Goodman3,5, and Larry Young2,4,5 1Neuroscience PhD Program, 2Center for Translational Social Neuroscience, 3Department of Radiology, 4Division of Behavioral Neuroscience and Psychiatric Disorders, 5Yerkes National Primate Research Center. Emory University, Atlanta, GA

For decades, the prosocial behavioral effects of oxytocin (OT) have been studied in several animal species, including humans. However, the neuroanatomical distribution of the oxytocin receptor (OTR) in the human and non-human primate brain has not been elucidated. This is due to a lack of reliable OTR antibodies and to non-selective radioligands, which exhibit mixed affinities for OTR and vasopressin 1a receptor (V1aR) in primates. In order to determine where OTR is expressed in primate brain, we have been working on two detection methods: development of an OTR PET ligand for in vivo neuroimaging and a pharmacologically informed modification for receptor autoradiography in post-mortem primate brain sections. We report here the biological evaluation of novel candidate OTR PET ligands as well as in vivo neuroimaging results in rhesus and cynomolgus macaques. We also report the pharmacological characterization of the aforementioned radioligands for competitive binding autoradiography in primate tissue. Our results from neuroimaging, autoradiography, and in situ hybridization reveal a general lack of OTR density in the macaque brain and indicate that OT binds primarily to the V1aR in the macaque brain. These results support the interpretation that OT’s effects in macaque brain may be due to action at the V1aR. Andrew Bankston, BCDB 4:15 pm

MOLECULAR MECHANISMS AND ESSENTIAL ROLES OF CDK5 ACTIVATION IN OLIGODENDROGLIA DIFFERENTIATION AND MYELIN LESION REPAIR Andrew N. Bankston1, Wenqi Li1, Hui Zhang1, Li Ku1, Guanglu Liu1, Lixia Zhao1, and Yue Feng1 1 Department of Pharmacology, Emory University School of Medicine, Atlanta, GA

Cyclin-dependent kinase 5 (Cdk5) and its activators, p35 and p39, are key players that govern normal brain development and function. Dysregulation of the Cdk5 pathway results in neurodegeneration and cognitive impairment. Besides the well-demonstrated primary role of p35-activated Cdk5 in neurons, emerging evidence points to the essential function of Cdk5 in oligodendrocytes (OLs), the myelinating glia in the central nervous system (CNS). However, whether neurons and OLs employ similar or distinct mechanisms to regulate Cdk5 function remains elusive. We report here that in contrast to neurons that harbor both p35 and p39, OLs express abundant p39 but negligible p35. Furthermore, p39 is up-regulated in OLs during differentiation and myelinogenesis along with increased Cdk5 activity while p35 expression remains unaltered. Moreover, specific knockdown of p39 significantly attenuates Cdk5 activity and OL differentiation. Finally, p39, but not p35, is increased during normal myelin repair, and remyelination is severely impaired in p39-/- mice. Together, these results demonstrate that p39-dependent Cdk5 activation is essential in advancing OL differentiation during de novo myelinogenesis and myelin repair. Moreover, the differential regulation of Cdk5 activators in neurons and OLs allows cell-specific control of Cdk5 activity during the distinct developmental timings of these neural lineages, providing possible therapeutic targets for distinct brain disorders caused by Cdk5 dysregulation in neurons and glia.


Eric Armstrong, MSP 4:30 pm

TERMINING THE MOLECULAR MECHANISM UNDERLYING LIGAND-DEPENDENT ACTIVATION OF FABP5 Eric H. Armstrong1, Eric A. Ortlund1 1Emory University, Atlanta, GA, USA

The fatty acid binding protein (FABP) family includes nine known members, each ~14-15kDa and found throughout the animal kingdom. Though traditionally thought to be involved in the solubilization/protection of their hydrophobic cargos, more recent studies have recast certain FABPs as specific mediators of vital signaling pathways. FABP5, like its family members, displays a promiscuous ligand binding profile, being able to form a complex with numerous long chain fatty acids of varying degrees of saturation. Interestingly, of those previously tested, only linoleic acid (LA) had been demonstrated as an “activator,” whose binding results in the protein’s nuclear translocation from the cytoplasm in cellular assays. Structural determination of both the apo and LA-bound protein has helped elucidate not only a tertiary nuclear localization signal (NLS) within FABP5’s alpha helical lid, but also residues within the protein’s α2 helix and portal loop that serve as the driving force behind NLS formation. Finally, as LA was able to adopt either of two possible conformations within the binding pocket, we hypothesize that those fatty acids more likely to assume a “U” versus a “bent” shape while bound will have a higher likelihood of inducing subsequent activation, thus implicating FABP5 specifically in polyunsaturated fatty acid signaling. Kalynda Gonzales, NS 4:45 pm

GABAERGIC REGULATION OF STRIATAL CHOLINERGIC INTERNEURONS: SYNAPTIC INPUTS FROM PARVALBUMIN-CONTAINING INTERNEURONS Gonzales KK1,2, Pare J-F, Wichmann T, and Smith Y. 1Yerkes National Primate Research Center, 2Deparment of Neurology, Emory University, Atlanta, GA

Anatomical, electrophysiological and neurochemical studies have shown that the activity of striatal cholinergic interneurons is tightly regulated by GABAergic inputs. Recent findings from our laboratory have shown that striatal cholinergic interneurons in monkeys, indeed, receive a prominent GABAergic innervation (Gonzales et al., 2012). Although about 24% of these terminals originate from axon collaterals of striatal projection neurons, the source(s) of the other GABAergic terminals that contribute to this innervation remain to be established. We, therefore, carried out a quantitative ultrastructural analysis of the innervation of striatal cholinergic interneurons by the GABAergic parvalbumin (PV)-containing interneurons. Striatal tissue was processed for pre-embedding double immunocytochemistry at the electron microscopic level using antibodies against PV and choline acetyltransferase (ChAT). PV-immunoreactive terminals that form symmetric synapses account for about 70% of the total population, while the remaining boutons form asymmetric synapses. Our double immunocytochemical studies revealed that almost 45% of PV-immunoreactive GABAergic terminals, which mostly arise from striatal interneurons, target ChAT-positive dendrites. In contrast, only 5% of glutamatergic PV-containing boutons, likely from the thalamus, terminate on ChAT-positive dendrites. Altogether, our data suggest that striatal cholinergic interneurons are under strong modulatory influences from intrinsic GABAergic axon collaterals of striatal projection neurons and PV-containing interneurons.


Poster Presenters Poster Presenter Program#1 Adesina, Sherry MSP #2 Allen, Megan BCDB #3 Alter, Shawn MSP #4 Barron, Daniel BCDB #5 Bienkowski, Rick GMB #6 Bizzell, Erica MMG #7 Bryant, Katherine L. NS #8 Burkett, James NS #9 Coakley, Kathryn NHS #10 Colucci, Jennifer BCDB #11 Cordova, James MSP #12 Donlin-Asp, Paul BCDB #13 Eufinger, Silvia NHS #14 Evans, Paul NS #15 Fagan, Crystal BCDB #16 Fan, Yanjie NS #18 Frediani, Jennifer NHS #17 Fritz-French, C. IMP #19 Gavile, Catherine IMP #20 Georgieva, Maria MMG #21 Giddens, Michelle NS #22 Harrell, Constance NS #23 Howery, Kristen MMG #24 Huang, Brenda GMB #25 Hudson, William MSP #26 Infield, Daniel MSP #27 Jackson, Sandra NHS #28 Kandler, Justin MMG #29 Keebaugh, Erin PBEE #30 King, Lanikea NS #31 Koenig, Seth NS #32 Kuiper, Emily BCDB #33 Lenzi, Gina MSP #34 Levine, Rebecca C.S. PBEE #35 Lohr, Kelly NS

Poster Presenter Program#36 Mandler, Mariana BCDB #37 Marcinkevage, Jessica NHS #38 Mariani, Laura NS #39 McGrath, Callie NS #40 Mohan, Gopi IMP #41 Moore, Carla NHS #42 Morreall, Jordan GMB #43 Murray, K.E. NS #44 Musille, Paul MSP #45 Myrick, Leila NS #46 Nanes, Benjamin BCDB #47 Newman, Laura BCDB #48 Petrova, Lucy GMB #49 Rha, Jennifer BCDB #50 Rose, Samuel NS #51 Rotterman, T.M. NS #52 Schureck, Marc BCDB #53 Shah, Shardule IMP #54 Shaikh, Nida NHS #55 Solyst, J.A. NS #56 Soucek, Sharon BCDB #57 Stahley, Sara BCDB #58 Swanson, Andrew NS #59 Vachon, Virgina MMG #60 Vinal, Kellie MMG #61 Wasson, Jadiel BCDB #62 Watkins, K.E. NS

#63 Williams-Newkirk, Amanda J PBEE

#64 Williams, Kathryn R. BCDB #65 Zakas, Philip MSP #66 Chau, Monica NS #67 Siegel, Karen NHS #68 Stout, Kristen MSP

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 NINTH ANNUAL DSAC STUDENT RESEARCH SYMPOSIUM [28]

Sherry Adesina, MSP Poster #1

MITOCHONDRIAL CATALASE EXPRESSION PROTECTS AGAINST HYPOXIA-INDUCED PULMONARY HYPERTENSION Sherry Eniola Adesina, C. Michael Hart, Roy L. Sutliff Pulmonary and Critical Care Medicine, Atlanta VAMC/Emory University, Decatur, GA Pulmonary hypertension (PH) is characterized by increased pulmonary vascular remodeling, resistance, and pressures. Reactive oxygen species (ROS) contribute to PH-associated vascular dysfunction. NADPH oxidases (Nox) and mitochondria are major sources of ROS in the vasculature. The crosstalk between these two ROS sources under hypoxic conditions is unknown.The antioxidant catalase attenuates H2O2. A mouse model with mitochondria-targeted catalase expression (mCAT) was used to assess the role of mitochondrial H2O2 in hypoxia-induced PH. We hypothesize that mCAT expression will reduce mitochondria-derived ROS and hypoxia-induced PH pathogenesis.Control and mCAT mice were exposed to normoxia (21% O2) or hypoxia (10% O2) for 3 weeks. PH development was assessed by measuring right ventricular systolic pressure (RVSP) and right ventricular hypertrophy (RVH). ROS levels and Nox mRNA expression were measured by Amplex red assay and qRT-PCR, respectively. Compared to control mice, mCAT mice developed less hypoxia-induced increases in RVSP, Nox mRNA, and lung H2O2 release. mCAT expression did not prevent hypoxia-induced increases in RVH. These studies suggest that attenuation of mitochondriaderived ROS may be protective against. Megan Allen, BCDB Poster #2

POTENTIAL ROLEFOR THE SELECTIVE RNA BINDING PROTEIN, HUD, IN REGULATING CDK5 SIGNALING Megan Allen, Li Ku, YueFeng Department of Pharmacology, Emory University, Atlanta, GA 30322 Cyclin dependent kinase 5 (Cdk5) governs neuronal development and synaptic functionby phosphorylatingdiverse proteins. Dysregulation of Cdk5 occurs in multiple neurodegenerative disorders.The abundance of two activating proteins, p35 and p39, controls Cdk5 activity.These distinct proteins are extremely labile, suggestingtight, dynamic regulation of Cdk5 activity.Unfortunately, regulation of p35 and p39 expression is understudied. Reports suggest regulation of p35 mRNA, while regulation of p39 has not been addressed. Whether p39 undergoes unique regulation is an interesting possibility. The neuronal RNA binding proteinHuD shares a functional spectrum with the Cdk5 pathwayand binds Au-Rich Elements (AREs) to stabilizes mRNA targets. Our sequence analysis identified AREs in the 3’UTRs of p35 and p39 mRNA.Here,our data suggests p39, but not p35, mRNA copurifies with HuD. Moreover,p39 mRNA and protein is elevated in the hippocampus of mice expressing transgenic HuD, while p35 protein remains unchanged. These data lead us to hypothesize thatHuD binds to and stabilizes p39 mRNAin an ARE-dependent manner, governing Cdk5 activator abundance and thus Cdk5 activity. These studies suggest a novel mechanism regulating Cdk5 signaling and provide clues to direct future studies investigating Cdk5 signaling in normal neurological function and disease.

The Tenth Annual DSAC Student Research Symposium [29]

Shawn Alter, MSP Poster #3

PROGRESSIVE NORADRENERGIC DEGENERATION IN MICE WITH REDUCED EXPRESSION OF THE VESICULAR MONOAMINE TRANSPORTER (VMAT2) Shawn P. Alter1,2,4

, Tonya N. Taylor1,2,5, Patti S. Sullivan6, David S. Goldstein6, Gary W. Miller1,2,3,4

1Center for Neurodegenerative Disease, 2Dept. of Environmental Health, 3 Dept of Neurology, and 4. Molecular and Systems Pharmacology Program, Emory University, Atlanta, GA, 5 Dept of Neuroscience, University of Minnesota, Minneapolis, MN, 6 Clinical Neurocardiology Section, National Institute of Health, Bethesda, MD Parkinson’s disease features the progressive loss of catecholamine neurons in the substantia nigra pars compacta (SNc) and locus ceruleus (LC). Here, we report parallel disruptions in the norepinephrine and dopamine systems of VMAT2-deficient mice. Unbiased stereological analysis reveals that VMAT2-deficient mice undergo progressive LC degeneration beginning at 12 months of age, preceding SNc degeneration at 18 months. LC degeneration is accompanied by relative reductions in norepinephrine transporter (NET) immunoreactivity throughout the brain including the hippocampus, the bed nucleus of the stria terminalis, and frontal cortex. 3H-nisoxetine binding reveals a 10% reduction in cortical NET. In addition to their neurodegenerative phenotype, VMAT2-deficient animals also exhibit alterations in catecholamine homeostasis. HPLC analysis revealed increased ratios of DHPG:NE and drastic increases in DOPAC:DA throughout the brain, indicative of increased cytosolic catecholamine catabolism. These aberrations are not limited to the central nervous system. VMAT2 deficient hearts show a 97% reduction in norepinephrine and a 10-fold increase in cardiac DHPG:NE, accompanied by a 40% downregulation of cardiac NET. These data describe an animal model of aberrant catecholamine metabolism and progressive catecholaminergic degeneration, demonstrating that reduced catecholamine storage in an in vivo model is sufficient to drive neurodegeneration in brain and peripheral regions afflicted in Parkinson’s disease. Daniel Barron, BCDB Poster #4

DYNEIN LIGHT CHAIN 1 (DLC1) IS REQUIRED FOR NORMAL GROWTH IN DROSOPHILA MELANOGASTER D. Barron and K. Moberg2*. Biochemistry, Cell, and Developmental Biology, Emory University, Atlanta, GA.

Carcinogenesis is fueled by the deregulation of oncogenes and tumor suppressor genes, which allows for excessive cell proliferation and tissue growth. The Dlc1 (dynein light chain 1) gene encodes a very well conserved protein that can promote cancerous phenotypes in cultured cells and is deregulated in human cancers. Our goal is to use Drosophila as a model system to elucidate the molecular pathway(s) through which Dlc1 influences cell biological process that underlie tumorgenesis.The Dlc1 protein was originally described for its accessory role in dynein motor complexes; however, it is now becoming clear that Dlc1 plays a variety of dynein-independent roles in the cell by acting as a ‘dimerization hub’ to promote the formation of higher-order protein complexes. Using Drosophila, we have identified a previously undescribed phenotype whereby ‘knocking down’ Dlc1 protein with an RNAi transgene reduces organ size. This phenotype offers a unique opportunity to genetically dissect the growth regulatory function of Dlc1. Furthermore, the levels of dMyc (the ortholog of the human oncogene c-Myc) and DIAP1 (a key anti-apoptotic protein) are reduced in Dlc1 deficient cells, thus linking Dlc1 to proteins that play conserved roles in cell growth and survival in human cells as well.


Rick Bienkowski, BCDB Poster #5

INTERACTIONS BETWEEN FNE AND THE POLYADENOSINE RNA BINDING PROTEIN DNAB2 IN A FLY MODEL OF HUMAN INTELLECTUAL DISABILITY Rick Bienkowski 1,2, Kenneth Moberg 1, and Anita Corbett 2 1 Department of Cell Biology, Emory University, Atlanta, GA, 2 Department of Biochemistry, Emory University, Atlanta, GA. Intellectual disability (ID) refers to a collection of diseases that are characterized by limited intellectual capacities and major constraints in adaptive behavior. We have found that inactivating mutations in the human gene encoding ZC3H14, a polyadenosine RNA binding protein, cause autosomal recessive non-syndromic ID. A Drosophila melanogaster model of this disease in which the orthologous gene, dNab2, has been removed, recapitulates key aspects of the human phenotype including impaired neural function. Published and unpublished data suggest that dNab2 may engage in physical, functional, and developmental interactions with found in neurons (fne). fne, an AU-rich element RNA binding protein, is conserved in yeast and humans. Previous work in yeast has shown Nab2 binds to the fne ortholog Pub1. Evidence from knockdown experiments in MCF7 cells suggests ZC3H14 regulates a common set of transcripts with HuR, a homolog of fne. In flies, mutants of both genes elicit similar developmental defects in the brain. Also, behavioral defects in flies knocked down for dNab2 in neurons are suppressed by co-depletion of fne. We have taken both a genetic and biochemical approach to test the hypothesis that dNab2 and fne regulate subset of mRNA transcripts that are essential for proper brain development and neuronal function. Erica Bizzell, MMG Poster #6

CHARACTERIZATION OF THE SERINE PROTEASE, RV2223C OF MYCOBACTERIUM TUBERCULOSIS, AND ITS SUBSTRATES 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 and replicate within immune cells. These include alteration of its complex cell wall during intracellular growth, and secretion of effectors that modulate immune responses and enhance pathogen survival. Our lab has identified a cell-wall associated serine protease, Hip1, which is involved in modification of the Mtb cell wall during infection, and which functions by dampening the normal immune response to Mtb infection. Encoded one gene downstream of hip1 is another serine protease, Rv2223c, with 52% identity to Hip1. Unlike Hip1, Rv2223c is predicted to be secreted extracellularly, and has been identified in the culture filtrate fraction of Mtb by mass spectrometry. Secreted proteases in other pathogenic bacteria have been shown to be involved in repression of cytokine production, degradation of surface-associated host proteins and ligands, and to even directly degrade cytokines. Therefore, we hypothesize that Rv2223c is a secreted protease that functions by cleaving Mtb substrates and/or host cell substrates to modulate host immune response to Mtb infection. To delineate the role of Rv2223c in Mtb pathogenesis, we are studying the effects of inactivating, deleting, and blocking secretion of Rv2223c on Mtb survival in macrophages and in vivo in mouse models. We are performing biochemical studies of Rv2223c enzymatic activity, and aim to understand its molecular function by identifying novel host and Mtb substrates of Rv2223c through pull-down assays and co-immunoprecipitation. These studies will provide insights into the mechanisms underlying secreted proteases in Mtb pathogenesis.


Katherine L. Bryant, NS Poster #7

ORGANIZATION OF THE CHIMPANZEE TEMPORAL LOBE: LOCALIZATION OF AUDITORY CORE AND THE AREA MI+ COMPLEX USING DIFFUSION TENSOR IMAGING K. L. Bryant1, D. A. Gutman2, 5, L. Li6, X. Zhang3, T. M. Preuss41Graduate Program in Neuroscience, 2Center for Comprehensive Informatics, 3Yerkes Imaging Center, 4Yerkes National Primate Research Center, Emory University, Atlanta, GA; 5Department of Biomedical Informatics, Emory School of Medicine, Atlanta, GA; 6Department of Biomedical Engineering, Emory University and Georgia Institute of Technology, Atlanta, GA

Understanding the evolutionary specializations of the human brain necessitates the study of our closest relatives, the chimpanzees. Noninvasive neuroimaging, especially DTI, has made direct, detailed comparisons of humans, chimpanzees, and other nonhuman primates possible. To evaluate the potential of DTI, we performed probabilistic tractography on in vivo chimpanzee scans to localize two major cortical landmarks: 1) the auditory core, and 2) the MT+ complex. Hypotheses: 1) the auditory core of the chimpanzee would be localized in Heschl’s gyrus; and 2) the MT+ complex would be localized in the posterior part of the superior temporal sulcus (STS). Region-of-interest (ROI) masks included the hypothesized locations of the two landmark regions and their known input regions on T1-weighted images with 0.8-mm isotropic resolution. FSL (www.fmrib.ox.ac.uk/fsl) was used to track connectivity in diffusion-weighted scans (1.8 mm isotropic resolution, 60 gradient directions, b values of 0, 1000s/mm2). MGN projections were localized in Heschl's gyrus, indicative of the auditory core. Tracts from IPS and V1/V2 converged in a small gyrus buried in the posterior STS, indicative of a macaque-like position of area MT+ rather than a human-like position. DTI and probabilistic tractography can contribute to the construction of a detailed map of chimpanzee cortical organization. James Burkett, NS Poster #8

NEUROBIOLOGY OF CONSOLING BEHAVIOR IN AN ANIMAL MODEL James Burkett1,2, Daniel Curry1, Melissa Reyes3, Larry Young1,2

1Graduate Program in Neuroscience, Emory University, 2Center for Translational Social Neuroscience, 3Department of Biology, Emory University.

In humans, consolation is a common expression of compassion, or sympathy with the suffering of another. Consoling behavior has now been observed in several animal species, including great apes, canids, corvids, and elephants. However, to date, no experimental animal model has been established that would allow for the discovery of causal neural mechanisms, since all existing studies have used observation of naturally occurring behavior. Here I present the first experimental data showing that male prairie voles (Microtus ochrogaster) express consoling behavior under laboratory conditions by directing affiliative grooming toward their female partners in a manner that is sensitive to the degree of stress the female partner has experienced. Stress and anxiety measures suggest that this behavior may serve to buffer the stress of the female. I will also show that promiscuous meadow voles do not display this stress-evoked increase in allogrooming under identical conditions. Finally, I will present additional evidence suggesting that oxytocin receptor activation in the brain is necessary for this stress-evoked increase in allogrooming. These experiments are the beginning of a foundation for an animal model that will inform us about the neurobiology of consolation.


Kathryn Coakley, NHS Poster #9

USING STANDARD PARAMETERS TO ESTIMATE BONE MINERAL DENSITY (BMD) IN PHENYLKETONURIA Kathryn E Coakley MS, RD1, Teresa D Douglas PhD2, Rani H Singh PhD, RD2 1Molecules to Mankind Program and Graduate Programs in Nutrition and Health Sciences and Human Genetics, Laney Graduate School, Emory University, Atlanta, GA, 2 Department of Human Genetics, Division of Medical Genetics, Emory University, Atlanta, GA Individuals with phenylketonuria (PKU) have poor bone health. Bone mineral density (BMD) measured DXA is expensive and involves radiation. Predicting BMD using standard-of-care measures could quickly identify those at-risk. Demographic (age, sex, BMI), clinical (medical-food prescription, serum phenylalanine and tyrosine), and dietary intake (energy, fat, protein, cholesterol, vitamin D, phenylalanine) parameters from baseline of a clinical trial in PKU patients were examined. Correlation coefficients adjusted for age, sex, and BMI were calculated between individual parameters and BMD. Models using predictors significantly correlated to BMD and interaction terms were considered. A final model was selected considering r-square and number of predictors. To validate, categories of actual and predicted BMD z-scores were compared: >-1SD from reference(normal), -2.5 to -1SD from reference(at-risk), >-2.5SD from reference(osteoporosis). Data were available for 52 individuals (mean age=17.3). 13 (25.0%) had at-risk BMD; 3 (5.8%) had BMD indicating osteoporosis. Significant correlations were found for age (0.58;p-value:<0.0001), BMI(0.56;p-value:<0.0001), phenylalanine:tyrosine(-0.28;p-value:0.04), and medical-food prescription(0.29;p-value:0.04). The final model included age, phenyalanine:tyrosine, and interaction between BMI and medical-food prescription (r-square:0.7490,p<0.0001). Predicted BMD correlated with actual BMD (0.87;p-value<0.0001). Predicted z-scores agreed with actual z-scores (weighted kappa=0.96;p-value:<0.0001). >30% of participants had at-risk BMD. Routinely collected parameters can predict BMD and z-scores in patients with PKU. Jennifer Colucci, BCDB Poster #10

EVOLUTION OF LIGAND SPECIFICITY IN STEROID HORMONE NUCLEAR RECEPTORS Jennifer Colucci1, Eric Ortlund1

1Emory University, Department of Biochemistry

Steroid receptors (SRs) are ligand-regulated transcription factors that play critical roles in inflammation, development, and cancer progression. Each member of the SR family of proteins is finely tuned to be activated by its cognate endogenous hormone with high specificity. This family arose from a single evolutionary precursor that was activated by an aromatized 3-hydroxysteroid (3-HS). Throughout a series of gene duplications, the reproduced genes lost the ability to respond to aromatized 3-HSs and gained the ability to respond to nonaromatic 3-ketosteroids (3-KSs). Currently, the molecular basis for this functional switch is not well understood. To understand the molecular basis behind this functional switch, the ancestral precursor to the modern-day nonaromatic 3-KS receptors, Ancestral Steroid Receptor 2 (SR2) was resurrected. We took a two-pronged approach to understanding ligand recognition and activation, using both biochemical and structural techniques. We show that SR2 binds and is activated by nonaromatic 3-KSs and binds, but is not activated by aromatic 3-HSs. Additionally, nonaromatic 3-KSs stabilize SR2 while aromatic 3-HSs decrease receptor stability. Using the crystal structure of the SR-progesterone complex, we re-created key historical substitutions to reverse the specificity of the receptor from nonaromatic 3-KSs to aromatic 3-HSs.


James Cordova, NHS Poster #11

VOLUMETRIC EVALUATION OF TUMOR RESECTION USING TUMOR-TARGETING DYE, 5-AMINOLEVULINIC ACID (ALA), IN PATIENTS WITH GLIOBLASTOMA MULTIFORME James S. Cordova1,2, Scott Hwang2, Chad Holder2, Hyunsuk Shim2, Constantinos G. Hadjipanayis3 1 Dept. of Radiology, Emory University School of Medicine, Atlanta, GA 30033 2 Medical Scientist Training Program, Emory University School of Medicine, GA, 30033 3 Dept. of Neurosurgery, Emory University School of Medicine, Atlanta, GA 30033

Accurately defining brain tumor margins during surgery allows for better discrimination of normal and tumor tissue leading to improved patient outcomes, but this is difficult intraoperatively due to the peculiar nature of tumor infiltration. To address this, phase II evaluations of the oral fluorophore, 5-aminolevulinic acid (ALA), to identify and remove grade-IV gliomas by fluorescence-guided surgical resection are on-going. The preferential uptake of ALA by high-grade gliomas relative to normal tissue and its flourescent properties allow for the accurate intraoperative discrimination of tumor and normal tissue. In these studies, gross tumor resection is completed via radical craniotomy in groups administered ALA or placebo preoperatively, followed by standard postoperative MRI (including T1-weighted images pre- and post- gadolinium) at 48 hours. To quantitatively access the improvement of ALA-guided surgery, we used the imaging informatics software VelocityAI (Velocity Medical Solutions, Atlanta, GA) to estimate the 3-D tumor volumes before and after resection. Residual contrast enhancement images were produced by systematically subtracting co-registered, post-resection, pre- and post-gadolinium T1-weighted images correcting for perioperative blood product accumulation. Pre- and post-resection tumor volumes were calculated using a signal and contrast-based semi-automated segmentation algorithm, and the volumes of the experimental group were compared with controls as percent resection. Paul Donlin-Asp, BCDB Poster #12

EXAMINING THE ROLE FOR THE SURVIVAL OF MOTOR NEURON PROTEIN IN AXONAL MRNA LOCALIZATION AND TRANSPORT IN SPINAL MOTOR NEURONS C.Fallini,P.Donlin-Asp,G.J. Bassell and W.Rossoll

Spinal muscular atrophy (SMA) is a neuromuscular disease with a specific degeneration of motorneurons. SMA results from a reduction in the survival of motor neuron (SMN) protein, with a role in the assembly of snRNPs. While defects in splicing occur in SMA mouse models, these defects are not unique to motorneurons, making it unclear what extent they play in the pathophysiology of SMA. We have demonstrated axonal colocalization of SMN and various mRNA binding proteins (mRBPs) known to play vital roles in mRNA transport. Additionally, a defect in axonal localization of β-actin mRNA in primary motorneurons from the SMA mouse model has been observed. These data suggest that SMN may play a role in mediating the assembly of actively transported mRNA granules. We investigated the effects of SMN deficiency on both the localization and transport of mRNPsin primary motor neurons. We have identified a significant reduction in the axonal levels of Gap-43 mRNA, and in the axonal localization ofmRNPs regulating Gap-43 mRNA in motor neurons with depleted SMN protein.Our results support a model where SMN plays a non-canonical role in the specific assembly, transport or both of mRNPs in the axon of motor neurons.


Silvia C. Eufinger, NHS Poster #13

THE ASSOCIATION BETWEEN HABITUAL DIETARY SODIUM AND SOLUBLE CELL ADHESION MOLECULES AMONG MIDDLE-AGED MALE TWINS Silvia C. Eufinger1, Jack Goldberg2, Thomas R. Ziegler1,5, William M McClellan3, Amita K. Manatunga4, Viola Vaccarino1,3,6 1Nutrition and Health Sciences Program, Division of Biological and Biomedical Sciences, Laney Graduate School, Emory University, Atlanta, GA, 2Vietnam Era Twin Registry, Seattle VA Epidemiologic Research and Information Center and the Department of Epidemiology, School of Public Health, University of Washington, Seattle, WA, 3Department of Epidemiology, Rollins School of Public Health, Emory University, Atlanta, GA, 4Department of Biostatistics and Bioinformatics, Rollins School of Public Health, Emory University, Atlanta, GA, 5Department of Medicine, Division of Endocrinology, Metabolism and Lipids, Emory University School of Medicine, Atlanta, GA, 6Department of Medicine, Division of Cardiology, Emory University School of Medicine, Atlanta, GA

We investigated the role of habitual dietary sodium in the expression of soluble cell adhesion molecules, which are markers of endothelial dysfunction that contribute to the development of atherosclerosis. Our study included 475 middle-aged male twins recruited from the Vietnam Era Twin Registry. Habitual daily sodium intake was assessed using the Willett food-frequency questionnaire. We measured sVCAM-1 and sICAM-1 concentration using commercially available ELISA kits. We used mixed-effect regression analysis to examine individual-level effects and robust regression analysis to examine within-pair differences. An increase in dietary sodium of 1,000 mg/d was associated with an adjusted 12.2% higher sVCAM-1 concentration (95% CI: 3.1, 21.2; P = 0.009). This association persisted within twin pairs (P = 0.03), yet differed by zygosity. Among dizygotic pairs, a 1,000 mg/d within-pair difference in dietary sodium was associated with an adjusted 17.3% higher sVCAM-1 concentration in the twin with higher dietary sodium than his co-twin (P = 0.005). No association, however, was found among monozygotic pairs. For sICAM-1, the adjusted individual-level effect did not reach statistical significance (P = 0.39). Habitual dietary sodium is directly associated with sVCAM-1 concentration, independent of traditional CVD risk factors. However, shared genetic factors may mediate this association. Paul Evans, NS Poster #14

EXPRESSION AND LOCALIZATION OF RGS14, A NATURAL SUPPRESSOR OF HIPPOCAMPAL SYNAPTIC PLASTICITY, IS DEVELOPMENTALLY REGULATED IN POSTNATAL MOUSE BRAIN Paul R. Evans1,2, Sarah E. Lee2, Yoland Smith1,3, and John R. Hepler1,2

1Neuroscience Training Program, 2Department of Pharmacology, and 3Yerkes National Primate Research Center, Emory University School of Medicine. Atlanta, GA 30322. RGS14 is a scaffolding protein that integrates G protein, calcium, and MAPkinase signaling pathways in hippocampus. Our recent findings show that RGS14 is a natural suppressor of CA2 synaptic plasticity and hippocampal-dependent learning and memory. However, the spatiotemporal pattern of RGS14 expression during postnatal mouse development is unknown. Here, we report that total brain RGS14 mRNA and protein are upregulated during early postnatal development, with low mRNA levels present at P0 and protein first detected at P7, with both reaching highest levels that are sustained in adult. Immunolabeling of mouse brain sections reveal that RGS14 is found not only in hippocampal CA2 but also other brain regions during early development. RGS14 is expressed in the anterior olfactory nucleus and piriform cortex, and in neocortical layers II/III and V and entorhinal cortex during postnatal development. RGS14 is detected in hippocampus beginning at P7 with strongest immunoreactivity in CA2 and fasciola cinerea. After P7, RGS14 protein increases in hippocampus to reach steady state in adulthood, but disappears in other brain regions. These results show that RGS14 mRNA and protein are upregulated throughout postnatal mouse development with a dynamic spatiotemporal localization that is restricted to hippocampus and piriform cortex in adult mouse brain.


Crystal Fagan, BCDB Poster #15

UNDERSTANDING THE MOLECULAR MECHANISMS RESPONSIBLE FOR TRANSLATIONAL FIDELITY Crystal Fagan1,2, Jack Dunkle1, Tatsuya Maehigashi1, Stacey Miles1, Aishwarya Devaraji3, Kurt Fredrick3 and Christine M. Dunham1 1Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322 2Biochemistry, cell, and Developmental Biology Graduate Program, Graduate Division of Biological and Biomedical Science, Emory University, Atlanta, Georgia 30322 3Department of Microbiology, Center for RNA Biology, Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210

Normal cellular function is dependent on the faithful replication of the genetic code. This requires that replication, transcription, and translation all proceed with high levels of fidelity. While DNA and RNA polymerases have evolved editing mechanisms that fix errors after chemical incorporation of the incorrect substrate, the ribosome is unable to remove and replace incorrect amino acids. Instead, there are multiple checkpoints during translation to ensure correct selection of the incoming tRNA. Additionally, it has been shown that the ribosome utilizes a different mechanism from other polymerases to deal with the incorrect incorporation. Rather than editing the incorrect amino acid, the ribosome senses errors and decreases translational fidelity to allow release factors to recognize a nonstop codon and thus terminate translation. Using X-ray crystallography, I have structurally characterized both scenarios; that is, 1) where the ribosome makes numerous mistakes resulting from 16S rRNA mutations and 2) where the incorrect tRNA is selected. The 16S rRNA mutation disrupts a conserved intersubunit bridge not previously appreciated in fidelity, while the selection of the incorrect tRNA leads to both altered mRNA position and disruption of fidelity checkpoints. By studying both distinct errors, we will obtain a more complete understanding of translational fidelity. Yanjie Fan, NS Poster #16

ACTIN CAPPING PROTEIN IN THE CONVERSION OF DENDRITIC PROTRUSIONS TO SPINES Yanjie Fan1, James Zheng1

Department of Cell Biology, Emory University

The formation of dendritic spines represents a key step in the functional connection of neurons. The actin cytoskeleton plays a crucial role in the formation of dendritic spines, but the underlying regulatory mechanisms remain to be elucidated. We reported previously that capping protein (CP) knockdown in cultured hippocampal neurons resulted in a marked decline of the density of mature spines, accompanied by an increase in filopodia-like dendritic protrusions. In this study, we found that CP underwent a dynamic change in its spatial distribution during synapse formation. CP accumulated in dendritic protrusions gradually along the course of spine formation and maturation, suggesting CP recruitment may be an essential step for the conversion of those protrusions to spines. Our subsequent analysis revealed that in the transition stage, the percentage of dendritic protrusions containing PSD95 was much higher in normal developing neurons than CP knockdown neurons. Moreover, time-lapse imaging showed that the extension and retraction of dendritic protrusions were much more dynamic after CP knockdown. Based on these results, we propose a model of dendritic spine development – CP and PSD95 accumulate to stabilize dendritic protrusions and enable the subsequent structural reorganization, leading to the formation of mature dendritic spines.


C. Fitz-French, IMP Poster #17

IN VITRO STUDIES OF INTERFERON-ALPHA NEUROTOXICITY C. Fritz-French1, G. Bassell1, & W. Tyor1,2. 1Emory University, Atlanta, GA. 2Atlanta VA Medical Center, Decatur, GA. Elevated levels of interferon-alpha (IFN) in the central nervous system (CNS) are linked to cognitive dysfunction in patients with inflammatory CNS diseases such as HIV-associated dementia. Previous studies showed that IFN treatment of neuronal cultures caused a dose dependent decrease in dendritic branching and length, that was prevented after pre-treatment with IFN neutralizing antibodies. Antagonists to NDMA receptor were also found to be partially protective against IFNα induced neurotoxicity. To begin to determine the mechanism of IFN induced neurotoxicity, the cell signaling pathway involving IFNα receptor (IFNAR) was evaluated. We demonstrate increased interferon stimulating gene 15 (ISG15) expression, an indicator for JAK-STAT cascade activation, after IFNα stimulation. Inhibiting IFNAR was found to partially protect neurons from IFNα neurotoxicity, but not as efficiently as neutralizing antibodies to IFNα. Our preliminary studies suggest that IFNα is in part acting through its receptor as well as NMDAR to reduce dendritic arborization in neurons. Further studies are needed to determine what other signaling pathways may be involved in IFNα induced neurotoxicity. Determining the mechanism of IFNα neurotoxicity could lead to therapies for cognitive dysfunction during neuroinflammation. Jennifer Frediani, NHS Poster #18

MACRONUTRIENT INTAKE AND BODY COMPOSITION OF PULMONARY TB PATIENTS IN TBILISI, GEORGIA Frediani JK1, Sanikidze E2, Gegechkori M2, Tukvadze N2, Kempker RR1, Hebbar G1, Blumberg HM1 Ziegler TR1 1Emory University, Atlanta, GA, USA; 2National Center for Tuberculosis and Lung Disease, Tbilisi, Georgia

Aim: To evaluate habitual intake of macronutrients and links with body composition in TB patients. Method: Dietary intake was obtained from patients at TB diagnosis using a tool specific to Georgian culture. Foods consumed during the previous 3 days were determined by one-on-one interviews. NDS-R software program was used and mean daily macronutrient intake determined. Body mass index (BMI; kg/m2) and body composition by bioelectrical impedance analysis were determined. Descriptive statistics, unpaired t-tests and linear regression methods were used. Results: In the 199 subjects (mean age 34 y; 64% male), BMI averaged 20.9±3.5 (SD) kg/m2; fat mass was 21.7±10.7%. Total energy intake averaged 54±21 kcal/kg/day (35% fat, 55% CHO and 12% protein). Protein intake averaged 1.6±0.7 g/kg/day. Underweight subjects (BMI ≤ 18.5 kg/m2; n=43) consumed more total energy (64±21 vs. 51±21 kcal/kg/d; p=0.0007), protein (1.9±0.7 vs. 1.5±0.7; p=0.0008) and fat (2.4±1.0 vs. 2.0±1.0 g/kg/d; p=0.0254) vs. those with BMI > 18.5 kg/m2 (n=149). BMI negatively correlated with calorie, fat and protein intake (p=0.001). Conclusions: TB patients in Tbilisi exhibited a low-normal BMI, and 22% were underweight. Overall, subjects consumed a high-kcal, high-protein diet from mixed nutrient food sources. Underweight patients consumed significantly higher amounts of macronutrients than non-underweight patients.


Catherine Gavile, IMP Poster #19

CD28 AND CD86 ARE NECESSARY FOR MYELOMA CELL SURVIVAL Catherine M. Gavile1, Jayakumar Nair2, Kelvin P. Lee2, Sagar Lonial1, Lawrence H. Boise1

1Departments of Hematology and Medical Oncology and Winship Cancer Institute, Emory University, Atlanta GA, 2Departments of Immunology and Medicine, Roswell Park Cancer Institute, Buffalo, New York 14263

CD28 and CD86 are better known for their key roles in T-cell costimulation, where interaction between CD28 on T-cells and CD86 on antigen-presenting cells leads to survival and proliferation of T-cells. Recent work has shown that the CD86-CD28 pathway also plays an important role in normal plasma cell generation and survival. Interestingly, high expression of CD28 and CD86 are poor prognostic markers for myeloma patients. In order to determine the role of each of these molecules in myeloma physiology, we knocked-down either CD28 or CD86 on the myeloma cell via lentivirus-mediated shRNAs. We found that knockdown of CD86 leads to apoptosis in 3 myeloma cell lines (RPMI8226, MM1.s, and KMS18). Four days after infection with lentivirus containing shCD86, 45.7±4.9 and 60.3±4.6 percent control apoptosis was observed in RPMI8226 and MM1.s respectively, while less death was observed in KMS18 (17.6±1.6). CD28-knockdown resulted in apoptosis as well (24.9±4.3 for RPMI8226, 26.8±4.1 for MM1s, 21.8±3.8 for KMS18, percent control apoptosis). Consistent with these findings, we were unable to establish a myeloma cell line with stable knockdown of either CD28 or CD86. These data suggest that CD28 and CD86 are essential to prevent apoptosis of myeloma cells in vitro. To confirm these findings we determined the effects of CTLA4-Ig on myeloma survival. CTLA4-Ig inhibits CD86-CD28 signaling by binding to CD86, blocking its interaction with CD28. We found that treatment of RPMI8226 and MM1.s cells with CTLA4-Ig caused apoptosis in the myeloma cells after 2 days. Thus like normal plasma cells, CD28 and CD86 are required for the survival of myeloma cells. Maria Georgieva, MMG Poster #20

COMPLEX INTERPLAY BETWEEN A MYCOBACTERIUM TUBERCULOSIS SERINE PROTEASE AND ITS PHYSIOLOGICAL SUBSTRATE Maria Georgieva1, Jacqueline Naffin-Olivos2, Ranjna Madan-Lala1, Erica Bizzell1, Nathan Goldfarb3, Benjamin M Dunn3, Gregory Petsko2 and Jyothi Rengarajan1, 4* 1Emory Vaccine Center, 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, Emory University, Atlanta, GA

While many studies have identified Mycobacterium tuberculosis (MTB) virulence factors that allow this bacterial pathogen to successfully evade host immunity, their molecular functions remain poorly understood. We have previously shown that a cell envelope localized serine protease, Hip1, is critical for MTB virulence in vivo, survival in macrophages and resistance to cell wall directed stresses. Moreover, Hip1 modulates innate immune responses to MTB infection by dampening TLR2 activation in macrophages and prevents robust macrophage activation. To delineate the molecular mechanisms behind Hip1 function, we have purified recombinant Hip1 and a Hip1 active site mutant and present data that provide key biochemical and molecular insights into Hip1 and its substrates. We show that Hip1 exhibits protease activity against synthetic substrates and have identified a physiological substrate, MTB protein GroEL2. We show that Hip1 directly cleaves purified GroEL2 in vitro and that cleavage is optimal at intraphagosomal pH conditions. GroEL2 cleavage is specifically inhibited by serine protease inhibitors and requires the Hip1 active site. We have mapped the cleavage site within the N-terminus of GroEL2 in vitro and show that this site is required for GroEL2 cleavage within MTB. Interestingly, using size exclusion chromatography, we have determined that GroEL2 is present in a dimeric state and is converted into monomers upon cleavage by Hip1. We propose that the balance between the dimeric and monomeric state of GroEL2 is important for its effector functions and are currently investigating the biological significance of Hip1-dependent cleavage of GroEL2 during MTB growth. These studies provide key insights into developing Hip1 inhibitors that can potentially be used as immunomodulatory therapeutics for tuberculosis in conjunction with antibiotics.


Michelle M. Giddens, NS Poster #21

ACTIVATED MICROGLIA RELEASE THE NEUROTROPHIC FACTOR AND GPCR LIGAND PROSAPOSIN Michelle M. Giddens, Rebecca C. Meyer, Randy A. Hall Emory University, Rollins Research Center Rm 5172, 1510 Clifton Rd, Atlanta GA

Prosaposin is a neurotrophic factor that exerts protective actions on neurons and glia in the CNS. We have recently discovered that prosaposin is an agonist for the orphan GPCRs GPR37 and GPR37L1, however neither the mechanisms controlling its release nor the cell type(s) from which it is released in the CNS are known. Here we show that prosaposin is robustly secreted from cultured microglia, the primary immune cell of the brain. Interestingly, our studies have also revealed that microglial activation via the endotoxin lipopolysaccharide leads to substantially enhanced levels of prosaposin release. In contrast, stimulation of microglia with the active fragment of prosaposin, known as prosaptide, resulted in decreased levels of prosaposin release. This data suggests that endogenously expressed microglial GPR37 and GPR37L1 may serve as negative feedback autoreceptors to refine and constrain the prosaposin release from these cells. Since microglial activation in vivo occurs in response to injury or disease, our findings suggest a model in which injury can lead to prosaposin release from microglia in order to promote neuronal survival in the injured area. Understanding the mechanisms by which prosaposin is released by microglia could lead to novel treatments for stroke and neuroinflammatory conditions like multiple sclerosis. Constance Harrell, NS Poster #22

ADOLESCENT HIGH FRUCTOSE DIET INCREASES DEPRESSIVE AND ANXIETY-LIKE BEHAVIOR AND DECREASES HIPPOCAMPAL GLUCOSE TRANSPORTER 1 IN ADULT MALE RATS Constance S. Harrell1, Jillybeth Burgado2, Sean D. Kelly1, Gretchen N. Neigh1,3

1Department of Physiology, Emory University; 2Neuroscience and Behavioral Biology Program, Emory University; 3Department of Psychiatry and Behavioral Sciences, Emory University, Atlanta, GA.

Depression can be devastating during adolescence, altering physiology and behavior into adulthood. The prevalence of depression is doubled in diabetics, and both Type I and Type II diabetes’ rates are rising among youth. Moreover, adolescent stress is a risk factor for depression, and stress can promote metabolic dysfunction. PET studies have also demonstrated altered hippocampal glucose metabolism in depression. We hypothesize that primary changes in glucose transporters (GLUTs), particularly GLUT1, the blood-brain-barrier transporter, may partially underlie this effect. Here, we proposed that adolescent high fructose diet combined with chronic adolescent stress would a) induce an insulin-resistant state in adult male rats; b) alter affective behavior; and c) decrease hippocampal GLUT1 expression. Our experiments revealed that fructose-fed rats have elevated fasting plasma glucose and corticosterone relative to rats fed standard chow. Fructose-fed rats had a blunted corticosterone response in the forced swim test, irrespective of stress history. Behaviorally, fructose-fed animals spent less time in the open arms of the elevated plus maze and floated more in the forced swim test. Finally, RT-PCR and immunoblot both demonstrated decreases in hippocampal GLUT1 in fructose-fed rats. These studies show that adolescent high-fructose diet alters metabolism and changes affective behavior and hippocampal glucose transporter expression.


Kristen Howery, MMG Poster #23

QUORUM SENSING REGULATES STREPTOCOCCUS PNEUMONIAE BIOFILM FORMATION IN A BIOREACTOR WITH LIVING CULTURES OF HUMAN RESPIRATORY CELLS Kristen E. Howery1, Herbert P. Ludewick2, Porfirio Nava3, Keith P. Klugman2, and Jorge E. Vidal*2

1Department of Microbiology and Immunology, 2Hubert Department of Global Health, Rollins School of Public Health, and 3Department of Pathology and Laboratory Medicine, Emory University, Atlanta GA Streptococcus pneumoniae forms highly organized biofilms in the nasopharynx, ear epithelium, and lungs that are necessary for persistence, otitis media and pneumonia. In this work, we developed a bioreactor that mimics the microenvironment of the human respiratory epithelium to study production and regulation of S. pneumoniae biofilms (SPB). Biomass of SPB significantly increased when grown on human respiratory epithelial cells (HREC), compared to abiotic surfaces. Additionally, SPB made by invasive strains produced more biomass on lung cells compared to pharyngeal cells. Production of SPB was regulated by two quorum sensing systems, Com and LuxS/AI-2. While LuxS/AI-2 regulated biofilms on both HREC and abiotic surfaces, Com control was specific for those produced on HREC. SPB formed on immobilized HREC, and incubated under static conditions, were completely lysed 24 h post-inoculation. Biofilm lysis was regulated by quorum sensing, and the mechanism required the assemblage of a mature biofilm structure. Transcription of genes encoding the vaccine candidate pneumolysin (Ply) was upregulated in early and mature SPB, and structural studies further revealed that Ply links biofilm cells. Brenda Huang, GMB Poster #24

OLIGODENDROCYTE DYSFUNCTION IN HUNTINGTON’S DISEASE MICE Brenda Huang, Marta Anna Gaertig, Naureen Mitha, Xiao-Jiang Li, Shihua Li

Department of Human Genetics, Emory University, Atlanta, GA

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 movement disorder, as well as cognitive and behavioral deficits. 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. In HD patients, there is a decrease in the white matter volume of the brain that is apparent even before the onset of symptoms. Various mouse models of HD also have white matter abnormalities including defective myelination. These findings suggest 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 at 2 months of age and have locomotor and behavioral deficits, as well as body weight loss and reduced survival compared to control mice. These results suggest that the expression of mutant huntingtin in oligodendrocytes plays an important role in HD pathology.


William Hudson, MSP Poster #25

THE MECHANISM OF DIRECT GLUCOCORTICOID-MEDIATED TRANSREPRESSION William H Hudson, Christine Youn, Eric A Ortlund

Emory University, Department of Biochemistry, Atlanta, GA

Newly discovered negative glucocorticoid response elements (nGREs) mediate DNA-dependent transrepression by the glucocorticoid receptor (GR) across the genome. These nGREs control genes critical in metabolic, inflammatory, and circadian processes and may play a major role in side effects generated by glucocorticoids. nGREs differ dramatically in sequence and spacing from activating glucocorticoid response elements (GREs), and the mechanism driving GR binding to these elements is unknown. Using equilibrium binding studies, we demonstrate that GR binding to nGREs is mechanistically distinct from binding to GREs. Whereas GR binding to GREs exhibits strong cooperativity on two nearly identical binding sites, we demonstrate that nGREs contain two distinct binding sites to which GR binds with negative cooperativity. To uncover this mechanism of binding, we determined the structure of the GR DNA binding domain bound to an nGRE in the thymic stromal lymphopoetin (TSLP) promoter to a resolution of 1.9 Å. The structure of GR in complex with the TSLP nGRE reveals a dramatically different conformation than GR bound to activating GREs, preventing DNA-mediated dimerization. We propose that this novel GR conformation and negative cooperativity dictates monomeric binding at nGREs. Daniel Infield, MSP Poster #26

FLUORESCENCE STUDIES OF STRUCTURE AND CONFORMATIONAL CHANGE IN THE CYSTIC FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR Daniel T. Infield,1 Nael A. McCarty1,2 1Emory Medical School Department of Pediatrics, 2Children’s Healthcare of Atlanta

The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is a chloride channel expressed in several epithelial tissues. Inactivating mutations in CFTR cause the devastating disease Cystic Fibrosis. CFTR is unique in that it is the only member of the ABC Transporter family of ATP-powered pumps that functions as an ion channel. The conformational changes in CFTR’s transmembrane domains are a matter of debate; molecular dynamics simulations based on related transporters show dramatic inward to outward conformational transition that may or may not be conserved in CFTR’s gating cycle. To directly ask whether CFTR retains analogous movements accompanying channel opening, I have adopted the Voltage Clamp Fluorometry (VCF) method to CFTR, which allows simultaneous electrophysiological and optical recording of channels gating in cells. Thus far, we have successfully labeled one substituted cysteine residue in the first extracellular loop of CFTR (115C) and one near the top of the sixth transmembrane helix (R334C). In addition, we have successfully cloned GFP into the fourth extracellular loop of CFTR and verified the construct exhibits wild type CFTR behavior. With these constructs and a recently completed Voltage Clamp Fluorometry rig in hand, we are now conducting experiments measuring conformational changes in the pore of CFTR.


Sandra Jackson, NHS Poster #27

WHO IS MISED BY A1C SCREENING FOR DIABETES? Sandra L. Jackson, Darin E. Olson, Arun Mohan, Anne Tomolo, Diana Barb, Nadine Dubowitz, Phyllis Watson-Williams, Jonathan Ownby, Mary Rhee, Lawrence S. Phillips

Although there is increasing use of A1c screening to detect previously unrecognized diabetes, the characteristics of people with diabetes who are missed by A1c screening are unknown. To study this, we combined two datasets, one of veterans and another of the general population, where subjects had both OGTT and A1c assessed. We examined screening by ADA (A1c >= 6.5%) and VA (A1c >= 7.0%) criteria for diabetes. The 2,674 subjects were 64% male and 65% black, with average age 52 years and BMI 30. Of the 7% (n=183) who had previously unrecognized diabetes based on OGTT, A1c with ADA criteria missed 64% while VA criteria missed 87%. A1c screening was more likely to miss whites, possibly because of the association of black race with higher A1c, independent of glucose levels. Subjects missed by A1c screening had significantly lower fasting (both p<0.05) and 2hr OGTT glucose levels (both p<0.01), implying that they were earlier in their natural histories. Lower glucose levels could not be attributed to having lower age or BMI in multivariate analyses. A1c screening should be complemented by glucose measurements to detect patients who are earlier in their natural histories and who could benefit from preventive management. Justin Kandler, MMG Poster #28

THE MisR/MisS TWO-COMPONENT REGULATORY SYSTEM INFLUENCES EXPRESSION OF LptA AND MODULATES PHOSPHOETHANOLAMINE DECORATION OF LIPID A IN Neisseria gonorrhoeae Justin L. Kandler1,2, Jhuma Ganguly3,4, Russell Carlson3,4, 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; 3Department of Biochemistry and 4Complex Carbohydrate Research Center, University of Georgia, Athens, GA

Cationic antimicrobial peptides (CAMPs), a vital component of the human innate host defense, are constitutively produced by neutrophils and can be inducibly produced by epithelial cells. While many gonococci are killed by neutrophils during infection, a small subset of the gonococcal population can survive and perhaps even replicate within neutrophil phagolysosomes. We hypothesized that sub-lethal levels of CAMPs at epithelial surfaces might induce transcriptional changes in gonococci that increase their chances of neutrophil survival. Due to its well-established importance for CAMP resistance in meningococci, we asked if the MisR/MisS two component regulatory system (TCS) could impact gonococcal resistance to CAMPs. We found that a nonpolar insertional mutation in misR (which encodes the response regulator) decreased gonococcal resistance to polymyxin B (a model CAMP) and also decreased expression of lptA, which encodes a lipid A phosphoethanolamine (PEA) transferase essential for neisserial CAMP resistance. Furthermore, we found that PEA-modified lipid A species were less prevalent in the misR mutant than in the isogenic parent strain. We conclude that MisR transcriptionally controls PEA modification of lipid A in gonococci and that this mechanism is important for gonococcal resistance to CAMPs.


Erin Keebaugh, PBEE Poster #29

CHARACTERIZATION OF AN ANTI-WASP RESPONSE GENE IN DROSOPHILA Erin S. Keebaugh and Todd A. Schlenke Emory University Department of Biology

Parasitic wasps lay eggs in Drosophila larvae that hatch, consume larval tissues, and eclose from the fly pupal case, killing the fly in the process. In addition to eggs, wasps also inject venom proteins that serve to aid in wasp success. Drosophila larvae can respond to wasp infection by melanotic encapsulation, where fly hemocytes form a capsule around and kill the entrapped wasp egg. Microarray analysis of Drosophila larvae post-wasp infection identified several promising candidate anti-wasp genes including an extracellular C-type lectin, lectin-24A. Expression analysis of lectin-24A shows enriched expression in the larval fat body, regulation by the Imd/JNK pathway, and a wasp-specific regulatory response. Flies mutant for lectin-24A are less successful in encapsulating wasp eggs and surviving wasp attack. I have generated an anti-Lectin-24a antibody to determine whether Lectin-24a binds to wasp eggs or host tissues in infected flies. I have also generated a lectin-24a reporter construct to further dissect its temporal, spatial, and genetic regulation and to determine whether venoms of certain wasp species suppress induction of lectin-24a as part of their virulence mechanism. Lanikea King, NS Poster #30

A POLYMORPHIC CIS-ACTING REGULATORY ELEMENT SPECIFICALLY INFLUENCES OXYTOCIN RECEPTOR EXPRESSION IN THE PRAIRIE VOLE NUCLEUS ACCUMBENS K. Inoue, L. J. Young, L. B. King Center for Translational Neuroscience, Yerkes Primate Research Center, Department of Psychiatry, Emory University. Atlanta, GA.

Diversity in brain oxytocin receptor (Oxtr) expression is associated with species differences and individual variation in social behavior. Monogamous prairie voles have high densities of OXTR in the nucleus accumbens (NAcc) compared to non-monogamous species, and OXTR signaling in NAcc is necessary for monogamous behaviors. There is remarkable individual variation in OXTR density in the NAcc and higher levels of OXTR there have been shown to facilitate parental and bonding behaviors. To test the hypothesis that genetic polymorphisms contribute to this variation in Oxtr expression, we used pyrosequencing to survey for allelic expression imbalance (AEI) of Oxtr mRNA in the NAcc. AEI is indicative of regulatory elements contributing to variation in gene expression. PCR and pyrosequencing was targeted to SNP 2 (C/T) in the 3’UTR of Oxtr cDNA derived from NAcc of heterozygotes to measure relative abundance of the two alleles. T/C ratios were greater for cDNA than gDNA, confirming that Oxtr exhibits AEI in prairie vole NAcc. In a follow up experiment, we saw that T/T voles had higher overall density of OXTR than C/C voles. SNP2 is associated with a polymorphic regulatory element for Oxtr, and this element may confer higher sociality via increased NAcc OXTR.


Seth Koenig, NS Poster #31

POSTER ABSTRAC-PREDICTING VIEWING PATTERNS IN MACAQUES USING SALIENCE DETECTION AND A RANDOM WALK PROCESS Seth Koenig1, Elizabeth A. Buffalo2 1Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta GA 30332, USA, 2Yerkes National Primate Research Center and Department of Neurology, Emory University School of Medicine, Atlanta, GA Natural scenes are complex and composed of many objects. When macaques view natural scenes they choose to focus or fixate on certain objects but not others. Two prominent determinates of object fixation are thought to be social content (e.g. faces) and salience. In order to examine the extent to which macaques’ viewing patterns correlate with regions of high salience in natural scenes, we implemented a salience model to describe each scene. Salience was calculated using a linear combination of color, orientation, and intensity contrast on different spatial scales [1]. An analysis of salience values at each fixation location revealed that fixations occurred in regions with high salience significantly more often than what would be expected by chance (p < 0.05). These effects were strongest towards the beginning of each trial, suggesting that salient objects attract attention and guide the first few fixations in a natural scene. However, salience alone does not fully predict viewing behavior especially towards the end of each trial. Thus, we propose that a better model of viewing behavior might be to first detect the salient objects in the natural scene and then use the salience to create an environment for a biased, correlated random walk. Emily Kuiper , BCDB Poster #32

DISTINCT RNA STRUCTURES COORDINATE TARGET SITE MODIFICATION BY THE THIOSTREPTON-RESISTANCE METHYLTRANSFERASE Emily G. Kuiper1,2 and Graeme L. Conn1 1Department of Biochemistry, Emory University School of Medicine, Atlanta, GA, and 2Biochemistry, Cell and Developmental Biology (BCDB) Program.

Target site modification is one mechanism bacteria use to resist the effects of antibiotics. The antibiotic thiostrepton is produced by Streptomyces azureus, which protects itself from self-intoxication through methylation of adenosine 1067 (A1067) of 23S ribosomal RNA (rRNA) by the Thiostrepton-resistance methyltransferase (Tsr). This nucleotide is part of a short contiguous rRNA domain allowing us to investigate aspects of Tsr-RNA recognition using short model RNAs. To determine the RNA secondary structural features necessary for substrate recognition and catalysis, we have analyzed Tsr binding and methylation activity on RNAs of various sizes (15, 29 and 58 nts) and sequences. We show the 15 nt stem-loop containing A1067 is sufficient for binding but this RNA is not methylated. An adjacent RNA bulge loop, present in the 29 nt RNA, is necessary for catalysis and shortening the distance between the target loop and this bulge disrupts catalysis. These data support a model of substrate recognition where concomitant binding of the target loop and a distant site, potentially accompanied by conformation change within the protein, coordinate the specific recognition of nucleotide A1067 by Tsr.


Gina Lenzi, MSP Poster #33

BIOCHEMICAL AND STRUCTURAL BASIS FOR RELEASE OF STALLED RIBOSOMES BY A NOVEL TRANSLATION FACTOR, YAEJ Gina Lenzi1,2, Tatsuya Maehigashi2, Stacey Miles2 and Christine M. Dunham1,2 1Molecular and Systems Pharmacology Program, Emory University 2Biochemistry Dept, Emory University School of Medicine, Atlanta, GA Ribosomes are complex macromolecular machines responsible for catalyzing the synthesis of proteins in all organisms. Due to the essential role that ribosomes play in cell vitality, numerous mechanisms are in place to prevent the loss of fidelity during translation. Stalled ribosomes could potentially generate a mutant protein or amass on mRNAs to sequester them from translating other messages. Prokaryotes utilize a trans-translational mechanism controlled by transfer-messenger RNA (tmRNA) with small protein B (smpB) to recycle stalled ribosomes. However, deletion of tmRNA and smpB has no effect on growth despite recycling being an essential process. tmRNA-smpB requires an empty A site complex in order to alleviate a stalled ribosome so we hypothesize a second mechanism exists to liberate ribosomes stalled on truncated or nonstop mRNAs. This mechanism may rely on a small ribosome associated protein, YaeJ, which has been shown to hydrolyze peptidyl-tRNA. YaeJ contains the enzymatic domain of release factors (RF), but lacks the codon specificity domain that RFs use for stop codon recognition. My long-term goal is to determine how YaeJ attains specificity and activity on stalled ribosomes using biochemical and structural techniques. Here we present results from binding assays that compare YaeJ’s ability to recognize different ribosome complexes. Rebecca Levine, PBEE Poster #34

BLOOD-FEEDING PATTERNS OF CULEX MOSQUITOES IN ATLANTA, GA, 2010-2011 Rebecca C. S. Levine1, Daniel G. Mead2, Uriel D. Kitron1 1Emory University, Atlanta, GA, United States, 2University of Georgia, Athens, GA, United States

Since its introduction in 1999, West Nile Virus (WNV) has become the most important mosquito-borne disease in the USA. In order to determine feeding preferences of the primary WNV mosquito vector species in Atlanta, GA, we trapped Culex mosquitoes in various habitat types within Atlanta’s urban center during May-November, 2010-11. Those with blood in their abdomens were tested for WNV and analyzed to identify the vertebrate bloodmeal source using PCR and sequencing. Avian and mammalian (including human) bloodmeal host species were identified from 87% of the bloodfed mosquitoes. Bloodmeals were taken primarily (80.7%) from avian hosts alone, with a smaller number of meals (5.8%) taken from mammalian hosts alone. Approximately 13.5% of bloodmeals were mixed feeds taken from both avian and mammalian hosts. The avian bloodmeals represent several potential bridge and amplifying host species involved in the local transmission dynamics of WNV in Atlanta. We examine the host results in the context of each urban habitat type, with a particular focus on the observation that 65% of the feeds from mammalian hosts (alone or mixed) occurred in the most highly anthropogenically disturbed urban sites, and the implications of these results for potential spillover transmission of WNV in the urban setting.


Kelly Lohr, NS Poster #35

ALTERED NEUROCHEMICAL AND BEHAVIORAL PHENOTYPE OF MICE WITH OVEREXPRESSION OF THE VESICULAR MONOAMINE TRANSPORTER 2 (VMAT2; SLC18A2) Lohr, Kelly1, Bernstein, Alison I.1, Guillot, Thomas1, Stout, Kristen1, Heath, Ellen1, Wang, Minzheng1, Li, Yingjie1, Salahpour, Ali2, Miller, Gary W.1

1 Environmental Health , Emory University, Atlanta, GA, United States. 2 University of Toronto, Toronto, ON, Canada.

The vesicular monoamine transporter 2 (VMAT2) is responsible for packaging monoamines for rapid release at neuronal synapses. VMAT2 is also responsible for sequestering toxicants away from their sites of action in the cell. Despite previous evidence suggesting that decreased levels of VMAT2 can contribute to neuronal dysfunction, it was not known if an increase in VMAT2 protein would result in increased function in vivo. We generated C57BL6 BAC transgenic mice with increased VMAT2 expression to determine the effects of increased vesicular monoamine filling on associated neurochemical and behavioral outcomes. Our VMAT2-HI mice show robust VMAT2 overexpression, proper localization of VMAT2, increased whole brain synaptic vesicle [3H]-dopamine uptake, and a measurable behavioral phenotype. These data show the effects of elevated VMAT2 levels on neurochemistry and behavior, suggesting that increased vesicular dopamine storage mediated by increased VMAT2 levels can alter the neurochemical and behavioral output of the monoaminergic system. Finally, as VMAT2 acts as a neuroprotective mechanism in monoaminergic neurons, these VMAT2-HI mice serve as a valuable tool to investigate resistance to insult. Mariana Mandler, BCDB Poster #36

A NOVEL CYTOPLASMIC PATHWAY by QKI-hnRNP H/F GOVERNS ALTERNATIVE SPLICING IN CNS MYELINATION Mariana D. Mandler, Li Ku, Lixia Zhao, Yue Feng Department of Pharmacology, Emory University, Atlanta, GA

Alternative splicing (AS) allows for expression of multiple isoforms from single genes, which occurs in over 90% of the human transcriptome. Dysregulation of AS is observed in tumorigenesis and brain disorders. Particularly, key genes that control development of oligodendrocytes (OLs), the myelinating cells in the brain, are tightly regulated by AS. However, molecular mechanisms underlying AS in myelination and myelin-related diseases are poorly understood. In this study, we identified a novel pathway controlling AS in OLs. The selective RNA-binding protein QKI is essential for OL differentiation. Deficiency of QKI leads to severe defects in AS of OL transcripts in the quakingviable (qkv) hypomyelination mutant mice. Multiple QKI protein isoforms are expressed in OLs that display distinct cytoplasmic-nuclear distribution. Interestingly, the AS defects in qkv mutant can be rescued by expressing a cytoplasmic isoform of QKI. Moreover, we found that cytoplasmic QKI represses expression of the canonical splicing factors hnRNPF/H, leading to the developmentally programmed decline of hnRNPF/H during myelination. This novel pathway controls AS of a broad spectrum of transcripts that contain characteristic G-run motifs, which targets numerous genes implicated not only in normal brain development but also in the development of cancer.


Jessica Marcinkevage, NHS Poster #37

REDUCING SEDENTARY BEHAVIOR AND INCREASING PHYSICAL ACTIVITY DURING PREGNANCY: A FEASIBILITY STUDY Marcinkevage J1,2,3, Correa A4, Ramakrishnan U1, Sharma A1,5, Venkat Narayan KM1,6, Umpierrez GE6

1Program in Nutrition and Health Sciences, Department of Biological and Biomedical Sciences, Laney Graduate School, Emory University, Atlanta, GA, 2Oak Ridge Affiliated Universities, Oak Ridge, TN, 3Division of Birth Defects and Developmental Disabilities, Centers for Disease Control and Prevention, Atlanta, GA, 4Department of Medicine, University of Mississippi Medical Center, Jackson, MS, 5Department of Reproductive Health, Centers for Disease Control and Prevention, Atlanta, GA, 6Emory University School of Medicine, Atlanta, GA

Minority women are more likely than non-Hispanic white women to enter pregnancy overweight or obese, have higher gestational weight gain (GWG) and develop gestational diabetes (GDM). To help shape interventions to prevent excessive GWG and reduce rates of GDM, we are conducting a feasibility trial among overweight/obese black women receiving prenatal care at a large urban hospital. Women are randomly assigned to receive regular care or regular care plus a lifestyle intervention focused on increasing levels of moderate physical activity (PA). In the first trimester and at 24-28 weeks gestation we measure weight; glucose levels through 75g oral glucose tolerance test; and self-report PA. Preliminary analyses (n=55, 36(65%) with mid-pregnancy visit) show a decrease in median(range) total activity in both groups from first visit to mid-pregnancy: 340.8(86.5-744.6) to 286.2(76.3-553.3) MET-h/wk for intervention and 402.8(70.7-668.4) to 366.9(131.3-969.1) MET-h/wk for regular care. The median(range) GWG at mid-pregnancy is similar for both groups: 4.5(-0.9-9.8)kg for intervention vs 3.5(-5.1-13.1)kg for regular care. At mid-pregnancy, we see lower median fasting and 30min glucose values in intervention vs regular care participants (81.9 vs 86.9mg/dL and 113 vs 122mg/dL, respectively). These preliminary data demonstrate the feasibility of modifying lifestyle during early pregnancy in underserved black women. Laura Mariani, NS Poster #38

MOLECULAR MECHANISMS UNDERLYING HINDBRAIN MALFORMATIONS IN JOUBERT SYNDROME Laura E. Mariani, Tamara Caspary Department of Human Genetics, Emory University, Atlanta, GA

Joubert Syndrome (JS) is a complex congenital disorder involving a variety of neurological and non-neurological symptoms. JS is considered a ciliopathy because it can be caused by mutations in at least 18 different cilia-related genes, including ARL13B. Although the diagnostic hallmark of JS is the “molar tooth sign”—a hindbrain malformation involving misguidance of the axons that form the superior cerebellar peduncle—little is known about how the affected white matter tracts develop in normal individuals or in JS patients. Arl13b is a small GTPase that regulates cilia formation and Sonic hedgehog (Shh) signaling during neural development. We hypothesize that the neurons of the deep cerebellar nuclei, whose axons form the superior cerebellar peduncle, require Shh for proper guidance of their axons, and that ARL13B mutations cause hindbrain defects in JS by misregulating Shh signaling in these neurons. We are testing these hypotheses by deleting Arl13b in neurons known to use Shh as a guidance cue and observing the effects on axon pathfinding, as well as by deleting either Arl13b or the Shh effector, Smoothened, in deep cerebellar nuclei and observing the resultant hindbrain phenotypes.


Callie McGrath, NS Poster #39

ANTERIOR INSULA PREDICTS TREATMENT SPECIFIC REMISSION IN MAJOR DEPRESSION Callie L. McGrath1,2, Mary E. Kelley, PhD3, Paul E. Holtzheimer III, MD4, Boadie W. Dunlop, MD1, W. Edward Craighead, PhD1, Alexandre R. Franco, PhD5,6, R. Cameron Craddock, PhD7, Helen S. Mayberg, MD1,8 1 Department of Psychiatry and Behavioral Sciences, Emory University, 2Graduate Division of Biological and Biomedical Sciences, Emory University, 3Department of Biostatistics and Bioinformatics, Emory University, Atlanta, GA, USA, 4Geisel School of Medicine at Dartmouth, Lebanon, NH, USA, 5Department of Electrical Engineering, Pontifícia Universidade Católica do Rio Grande do Sul 6Brain Institute, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, Brazil, 7Center for the Developing Brain, Child Mind Institute, New York, NY, USA, 8Department of Neurology, Emory University, Atlanta, GA, USA

Currently, fewer than 40% of patients treated for a major depressive episode achieve remission with initial treatment. Identification of a biological marker that might improve these odds would have significant health and economic impact. Brain glucose metabolism was measured with positron emission tomography prior to treatment randomization of untreated major depressive disorder patients to 12 weeks of treatment with either escitalopram or manual-based cognitive behavioral therapy. Positive and negative predictors of remission (HDRS 17-item ≤ 7 at both weeks 10 and 12) were identified with a two-way ANOVA treatment (escitalopram, CBT) by outcome (remission, non-response) interaction. Of sixty-five protocol completers, 38 patients with clear outcomes and usable PET scans were included in the primary analysis. Six regions were identified with the right anterior insula showing the most robust discriminant properties across groups (Effect size =1.43). Lower insula metabolism was associated with remission to cognitive behavioral therapy and poor response to escitalopram, while higher insula metabolism was associated with remission to escitalopram and poor response to cognitive behavioral therapy. If verified with prospective testing, this insula biomarker provides the first objective biological stratifier to guide initial treatment selection for depression.

Gopi Mohan, IMP Poster #40

EBOLA VIRUS SECRETED GLYCOPROTEIN ALTERS THE HOST ANTIBODY RESPONSE VIA “ANTIGENIC SUBVERSION” Gopi S. Mohan, Ling Ye, Chinglai Yang, Richard W. Compans Department of Microbiology and Immunology, Emory University School of Medicine

In addition to its surface glycoprotein (GP1,2), Ebola virus (EBOV) directs the production of large quantities of a nonstructural secreted glycoprotein (sGP) whose function has been widely debated. The secretion of viral antigens has been studied in several viruses as a mechanism of host immune evasion, via competition for virus-specific antibodies. However, such activity has not been conclusively demonstrated for EBOV sGP. Here, we immunized mice with DNA constructs expressing GP1,2 and/or sGP and demonstrate that sGP can efficiently compete for anti-GP1,2 antibodies from mice immunized by sGP. However, sGP cannot compete for anti-GP1,2 antibodies from mice immunized solely against GP1,2. We term this phenomenon “antigenic subversion”, and propose a model whereby sGP induces the host immune response to focus on epitopes shared between sGP and GP1,2, thereby allowing sGP to bind and compete for anti-GP1,2 antibodies. This is distinct from previously proposed “decoy” mechanisms in which secreted antigens passively absorb virus-specific antibodies, since subversion requires exposure to sGP during generation of the anti-GP immune response. Antigenic subversion represents a novel virus escape strategy that likely helps EBOV survive in its host. Additionally, our findings suggest that a successful EBOV vaccine must elicit robust enough immunity to clear EBOV before it has a chance to establish infection and effect subversion.


Carla Moore, NHS Poster #41

DIETARY VARIETY IS ASSOCIATED WITH LARGER MEALS IN FEMALE RHESUS MONKEYS Carla J Moore1, 2, Vasiliki Michopoulos 3, Zachary P Johnson1, and Mark E Wilson1 1 Division of Developmental & Cognitive Neuroscience, Yerkes National Primate Research Center, Emory University, Atlanta GA; 2 Molecules to Mankind Program and Nutrition and Health Sciences Program, Division of Biological and Biomedical Sciences, Laney Graduate School, Emory University, Atlanta GA; 3

Department of Psychiatry and Behavioral Medicine, Emory University School of Medicine, Atlanta GA

The complex, interacting influences on energy balance prevent elucidation of the causal role of any single factor in the current obesity epidemic. However, greater variety in the food supply, particularly in the form of highly palatable, energy-dense foods, has likely made a contribution. This study was undertaken to test the hypothesis that greater dietary variety is associated with greater caloric intake within meals consumed by free-feeding female rhesus monkeys. Subjects were studied for two, two-week dietary phases. One phase consisted of a choice between a low caloric density diet (LCD) and a high caloric density diet (HCD). The other phase consisted of access to the LCD only. Animals consumed the most calories within meals comprised of both the LCD and HCD, which differed in caloric density, flavor, and texture. During the dietary choice condition, animals consumed fewer calories in the form of LCD-only meals. Interestingly, however, there was no statistically significant difference in the amount of calories consumed as HCD-only meals in the choice condition compared to meals in the LCD-only, no choice condition, suggesting consumption of a single food during a meal provides a constant sensory experience that may lead to more rapid habituation and subsequent meal cessation. Jordan Morreall, GMB Poster #42

A TRANSCRIPTION-FIRST VERSUS A REPLICATION-FIRST MODEL FOR THE APPEARANCE OF NEW MUTANTS DURING SELECTION Jordan Morreall1,2, Bernard Weiss3, Paul W. Doetsch1,4,5,6 1Department of Biochemistry, 2Graduate Program in Genetics and Molecular Biology, 3Department of Pathology, 4Emory Winship Cancer Institute, and 5Department of Radiation Oncology and 6Hematology and Medical Oncology, Emory University School of Medicine, Atlanta, GA

It has been a major tenet of evolutionary biology that mutation precedes selection. However, bacteria have been observed to evolve new mutations during selection under non-growth conditions, such as those within the human body, especially under antibiotic selection. One explanation for such “adaptive mutagenesis” is that an altered nucleotide may be transcribed to produce a mutant protein (“transcriptional mutagenesis”), which allows the cell to grow and undergo DNA replication, thereby establishing the mutation in the cell line (“retromutagenesis”). Transcriptional mutagenesis gives rise to mutations on the transcribed strand. We mutated lac amber (TAG) mutants of E. coli with nitrous acid and selected for Lac+ revertants. Because nitrous acid affects only A in an A/T base pair, DNA sequencing revealed the strand on which each mutation occurred. As predicted by the model, when the mutagen-treated cells were plated directly on selective media, the ratio of transcribed strand to non-transcribed strand mutations was 6-9:1; however, when plated after growth in rich media, the ratio was 1-2:1. The predominance of transcribed-strand mutations before selection suggests that transcriptional mutagenesis contributes to reversion to protrophy in our system via retromutagenesis. Therefore, transcriptional mutagenesis may give rise to other mutations such as those driving antibiotic resistance.


Karen Murray, NS Poster #43

VARIATIONS IN ESCITALOPRAM’S BEHAVIORAL EFFECTS ACROSS DIFFERENT MOUSE STRAINS FOR USE IN TARGETED MUTANT MOUSE STUDIES. K.E. Murray1, K.J. Ressler1,2, M.J. Owens1 1Department of Psychiatry and Behavioral Sciences, Emory University, 2HHMI

Escitalopram, the S-enantiomer of citalopram, is a selective serotonin reuptake inhibitor (SSRI) that binds to the serotonin transporter (SERT) to inhibit serotonin uptake. In addition to its classic SSRI mechanism it has been shown that a second escitalopram molecule is also able to bind allosterically to a distinct site on the SERT. The effects of this allosteric site are not fully understood. To study the allosteric site a line of allosteric null SERT knockin mice are being established. To identify its role in behavior, an appropriate behavior screen must be established that is sensitive to escitalopram and an allosteric inactive SSRI (fluoxetine) as well as having a dose-dependent SSRI response. The marble burying test (MB) was screened as such a test. Escitalopram and fluoxetine both decreased marbles buried in a dose-dependent manner. Consequently the MB test is an appropriate screen of SSRI behavior and will be useful in future studies to examine SSRI activity at the allosteric site using transgenic mice. Paul Musille, MSP Poster #44

STRUCTURAL MECHANISM FOR ACTIVATION OF THE ORPHAN NUCLEAR RECEPTOR LRH-1 BY ANTI-DIABETIC LIPID AGONISTS Paul M. Musille1, Manish Pathak1, Janelle R. Lauer2, William H. Hudson1, Patrick R. Griffin2 and Eric A. Ortlund1 1 Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 2Department of Molecular Therapeutics, The Scripps Research Institute, Jupiter, Florida The nuclear receptor Liver Receptor Homolog-1, LRH-1, plays an important role in controlling lipid and cholesterol homeostasis and is a potential target for treatment of diabetes and hepatic diseases. LRH-1 is known to bind phospholipids (PLs) but the role of PLs in controlling LRH-1 activation remains highly debated. Recently, the dietary PL, dilauroylphosphatidylcholine (DLPC, PC 12:0/12:0), was shown to activate LRH-1 in diabetic mice, lowering serum lipid levels and reducing blood glucose levels. DLPC selectively activates LRH-1 mediated transcription in luciferase assays, increases the ability of LRH-1 to interact with coactivators, and increases the production of LRH-1 target genes. Here we describe the x-ray crystal structure of both apo LRH-1 and the protein in complex with DLPC. This, in conjunction with biochemical, cellular, and state-of-the-art hydrogen-deuterium exchange methods, show that DLPC binding is a dynamic process that alters coregulator selectivity and that the lipid-free receptor interacts with the widely expressed corepressor SMRT. These results show that LRH-1 undergoes profound structural changes in response to ligand binding and confirms LRH-1’s viability as a therapeutic target for both agonist and antagonist design. This greatly enhances our understating of LRH-1 regulation and highlights its importance as a novel therapeutic target for controlling diabetes.


Leila Myrick, NS Poster #45

PATHOGENESIS OF NOVEL FMR1 MUTATIONS IN FRAGILE X SYNDROME Leila Myrick1, Mika Kinoshita1, Xuekun Li1, Stephen Warren1 1Department of Human Genetics, Emory University, Atlanta, GA

Fragile X Syndrome (FXS) is the most common cause of inherited intellectual disability and a leading known causes of autism. Most cases of FXS are caused by trinucleotide repeat expansion within the 5'UTR of the gene FMR1. To date, only one missense mutation has ever been reported in a patient (I304N-FMRP). Recently we identified 2 novel variants, G266E and R138Q, in patients with FXS-like symptoms who tested negative for repeat expansion. To determine if these variants are pathological, we used lentivirus to infect Fmr1 KO cells with G266E-FMRP or R138Q-FMRP. Interestingly, we found that G266E behaves like a functional null, similar to I304N, as it is unable to associate with polysomes or rescue AMPAR trafficking (two well-known functions of FMRP associated with regulating protein synthesis). Conversely, R138Q rescued all tested phenotypes except for synaptic overgrowth at the Drosophila neuromuscular junction (NMJ), indicating this variant does not impair global FMRP function but rather impairs presynaptic function specifically. These results provide information about how the different domains and structure of FMRP may be involved in FMRP’s normal functions, and also give important insights into the mechanisms of disease in these patients with non-traditional FMR1 mutations.

Benjamin Nanes, BCDB Poster #46

P120-CATENIN BINDING MASKS AN ENDOCYTIC SIGNAL CONSERVED IN CLASSICAL CADHERINS Benjamin A. Nanes,1,2 Christine Chiasson-MacKenzie,1 Anthony M. Lowery,3 Noboru Ishiyama,4 Victor Faundez,1 Mitsuhiko Ikura,4 Peter A. Vincent,3 and Andrew P. Kowalczyk1,5,6 1Department of Cell Biology, 2Graduate Program in Biochemistry, Cell, and Developmental Biology, 5Department of Dermatology, and 6Winship Cancer Institute, Emory University, Atlanta, GA; 3Center for Cardiovascular Sciences, Albany Medical College, Albany, NY; 4Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Toronto, Canada Classical cadherins mediate adhesive interactions that are essential for tissue architecture and integrity. Trafficking of cadherins to and from the plasma membrane is a critical cellular mechanism that dynamically modulates cadherin cell surface levels. Here, we demonstrate that the armadillo-family protein p120-catenin (p120) binds and physically occupies a previously unidentified endocytic motif in the cadherin cytoplasmic tail to prevent cadherin internalization. Mutagenesis of VE-cadherin revealed that a three amino-acid acidic motif (DEE) is critical for both p120 binding and cadherin endocytosis, and resulted in a cadherin that was both uncoupled from p120 and, paradoxically, stable at the cell surface. Mutation of three residues (GGG) directly adjacent to this motif also uncoupled p120 binding, but did not inhibit endocytosis. Using these mutants, we examined the effects of p120 binding and cadherin endocytosis on cell migration. Expression of wild-type and GGG-mutant VE-cadherin had no effect on endothelial cell migration in response to the angiogenic growth factor VEGF, while the endocytosis-defective DEE -mutant strongly suppressed migration in scratch wound assays. Collectively, these findings demonstrate that p120 masks a novel endocytic motif in the juxtamembrane domain of classical cadherins and that cadherin endocytosis is a critical feature of junction plasticity essential for cell migration.


Laura Newman, BCDB Poster #47

ARL2 LOCALIZES TO THE MITOCHONDRIAL MATRIX AND EXISTS IN A COMPLEX WITH ANT AND HSP60. Laura Newman1, Michael East1, Chengjing Zhou1, Samatha Mudigonda1, and Richard Kahn1. 1Department of Biochemistry, Emory University School of Medicine, USA Arl2, a member of the Arf family of GTPases, is highly conserved across eukaryotic evolution and vital for several model organisms. Though previously studied almost exclusively for its role in microtubule dynamics, we have discovered a role for Arl2 in mitochondrial function and energy metabolism. Knockdown of Arl2 by siRNA causes a 50% decrease in ATP levels, and we determined that this loss is in mitochondrial ATP. Additionally, loss of Arl2 activity causes mitochondrial fragmentation and a loss in mitochondrial motility. We have localized Arl2 to the mitochondrial matrix, along with an Arl2 effector (BART) and an Arl2 GAP (ELMOD2). Since ATP-generating machinery localizes to the mitochondrial inner membrane and matrix, Arl2 could directly regulate ATP production. Using blue native gels, we discoveredand partially characterized a ~500 kD band that is enriched in mitochondria and contains Arl2, the adenine nucleotide transporter (ANT), and HSP60.in one complex. From these data, we hypothesize that Arl2 regulates ATP generation, but we do not yet know whether complexed Arl2 or monomeric Arl2 regulates ATP. Future directions are to study Arl2 regulation by post-translational modifications, which might lend insight into how Arl2 regulates ATP levels. Lucy Petrova, GMB Poster #48

POTENTIAL ROLES OF TRANSCRIPTIONAL MUTAGENESIS IN ONCOGENE-INDUCED SENESCENCE AND TUMOR DEVELOPMENT Lucy Petrova1 and Paul W. Doetsch2, 3, 4, 5

1Program in Genetics and Molecular Biology, Departments of 2Biochemistry, 3Radiation Oncology, and 4Hematology and Medical Oncology and 5Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA

DNA damage, from endogenous and exogenous sources, poses a constant threat to genomic integrity. While the mutagenic potential of DNA damage during DNA replication is well known, a variety of mammalian cell types are largely quiescent, replicating their genomes infrequently, while still engaging in transcription. Certain base damages can also be bypassed by RNA polymerase, resulting in the misincorporation of the same bases opposite the lesion during transcription as in DNA replication, a process termed transcriptional mutagenesis (TM). 8-oxoguanine is a frequently occurring oxidative base damage product that is highly mutagenic at the level of both replication and transcription. When 8-oxoguanine-driven TM results in the production of oncogenic Ras transcripts, it can result in the activation of downstream Ras signaling effectors. However, it is unknown whether these biochemical changes can result in disease-relevant physiological phenotypes.

Under some circumstances mutant Ras can initiate the tumorigenic process, but in the absence of a cooperating mutation, it can drive a brief hyperproliferative phase associated with the accumulation of further, extensive DNA damage in the form of double strand breaks increases in reactive oxygen species and resultant base damage, culminating in the activation of the DNA damage response, epigenetic changes, and a stable growth arrest, termed oncogene-induced senescence, a potent tumorigenesis barrier. We are currently investigating whether TM driven by 8-oxoguanine, and other oxidative base damages, can induce such phenotypes.


Jennifer Rha, BCDB Poster #49

TWO MOUSE MODELS TO STUDY THE RNA BINDING PROTEIN, ZC3H14, IN THE BRAIN: INVESTIGATING HOW THE MUTATION OF A UBIQUITOUS PROTEIN LEADS TO NONSYNDROMIC INTELLECTUAL DISABILITY IN HUMANS Jennifer Rha1,4, Sara W. Leung1,4, ChangHui Pak2,4, Christina Gross3,4, Masoud Garshasbi5, Andreas W. Kuss5, Kenneth H. Moberg2,4, Gary J. Bassell3,4, Anita H. Corbett1,4 1Biochemistry; 2Genetics and Molecular Biology; 3Cell Biology; 4Emory University School of Medicine, Atlanta, GA USA; 5Max-Planck Institute for Molecular Genetics, Berlin, Germany

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 plan to exploit mice and mouse neuronal cell lines to extend our studies into vertebrate ZC3H14 and to address our hypothesis: ZC3H14 is required for proper post-transcriptional control of specific mRNA targets that are critical for neuronal function. Our long-term goals are to understand molecular mechanisms critical for normal brain function and how dysregulation of these processes leads to neuronal dysfunction and thus impaired brain function. Samuel Rose, NS Poster #50

CONDITIONAL GENETIC CELL-TYPE SPECIFIC ANALYSIS OF ATAXIA IN A KNOCK-IN MODEL OF EPISODIC ATAXIA TYPE-2 Samuel J. Rose1,2, Lisa H. Kreiner2, Edwin A. Melendez2, H. A. Jinnah3,4,5, Arn M.J.M van den Maagdenberg6,7, Ellen J. Hess2,3

1Neuroscience Graduate Program, Departments of 2Pharmacology, 3Neurology, 4Human Genetics, and 5Pediatrics – Emory University, Atlanta. Departments of 6Human Genetics and 7Neurology – Leiden University Medical Centre, The Netherlands.

Mutations in the CACNA1A gene, which encodes the pore forming subunit of the CaV2.1 (P/Q-type) Ca2+ channel, cause several neurological disorders including episodic ataxia type-2 (EA2). EA2 patients experience episodic attacks of ataxia. To better understand the pathogenesis of this disorder, we created a knock-in mouse model carrying a point mutation (4486T>G; F1406>C) that causes EA2 in humans. Mice carrying one copy of this mutation and a null Cacna1a allele (EA2/-) exhibit motor dysfunction. To determine the cell type(s) that cause this dysfunction, we are using Cre-lox genetics to confine the EA2/- genotype to cerebellar granule cells or Purkinje cells. Granule cell-specific EA2/- mice were generated by crossing math1CreER transgenic mice with mice carrying one copy of the EA2 knock-in and one copy of the floxed Cacna1a allele (EA2/Cacna1aflox mice). EA2/Cacna1aflox; math1CreER mice did not exhibit a phenotype on the accelerating rotarod, indicating that dysfunction of CaV2.1 in granule cells alone is insufficient to disrupt motor function. We are currently investigating whether dysfunction in Purkinje cells is sufficient to disrupt motor function using the same approach with the L7-Cre transgene. These studies more precisely delineate the contribution of different cerebellar cell types to the pathogenesis of EA2.


T. Rotterman , NS Poster #51

DISTRIBUTION OF VGLUT1 SYNAPSES ON THE DENDRITIC ARBORS OF CONTROL AND INJURED MOTONEURONS Rotterman T.M.1, Nardelli P.2, Cope T. C.2, Alvarez F. J.1

1Dept. of Physiology, Emory Univ., Atlanta, GA, 2Dept. Neuroscience, Cell Biol. & Physiol., Wright State Univ., Dayton, OH

Motoneurons receive proprioceptive Ia afferent synapses mediating the monosynaptic stretch reflex and are altered after peripheral nerve injury. The distribution of this synaptic input on dendritic arbors of control and injured motoneurons is still unclear. We electrophysiologically identified and filled with neurobiotin 6 medial gastrocnemius motoneurons in adult rats, 3 control and 3 that regenerated 1 year after tibial nerve transection and reattachment. Ia synapses on each motoneuron were identified by their VGLUT1 content. The motoneurons were reconstructed using Neurolucida. Individual dendrites were traced for up to 1600 µm, and they had total surface areas greater than 200,000 µm2 for both experimental and control motoneurons. Control motoneurons received approximately 1000 contacts, 50% of which were located within the first 250 µm from the soma. Within these first 250 µm, experimental motoneurons show a 75% reduction in overall contact number and 75% reduction in surface density. These data suggest that most of the VGLUT1 proprioceptive input on motoneurons target the more proximal dendrite locations. Because both stretch reflexes and proximal VGLUT1 synapses are reduced in parallel after peripheral nerve injury, we conclude that proximal proprioceptive VGLUT1 synapses are likely responsible for conveying stretch signals to motoneurons. Marc Schureck, BCDC Poster #52

MECHANISM OF HIGB-MEDIATED RIBOSOME-DEPENDENT MRNA DEGRADATION Marc A. Schureck 1,2, Tatsuya Maehigashi1, Jack Dunkle1, Jhomar Marquez1, Ajchareeya Ruangprasert1, Stacey Miles1, and Christine M. Dunham1,2

1Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322 2Biochemistry, Cell, and Developmental Biology Graduate Program, Emory University School of Medicine, Atlanta, GA 30322

Proteus vulgaris (P. vulgaris) is a major cause of urinary tract bacterial infections. This bacteria 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 DNA, protein, and polysaccharides. My goal is to elucidate the molecular mechanism of the ribosome-dependent RNase family of toxins that play an important but unknown role in biofilm formation. Host inhibition of growth B (HigB) is a member of this family and was first identified in P. vulgaris. HigB cleaves ribosome-bound messenger RNA (mRNA) however unlike other known toxin proteins, it cleaves adenosine-rich mRNA sequences rather than a three-base mRNA codon. Towards the elucidation of this novel mRNA decoding mechanism, we solved the X-ray crystal structure of HigB bound to the 70S ribosome. This structure reveals that HigB binds mRNA in the A site of ribosome and a highly conserved tyrosine appears important for recognition of one of the adenosine bases. Future biochemical and structural studies of various adenosine-rich sequences cleaved by HigB will help explain how HigB reads mRNA codon-independently.


Shardule Shah, IMP Poster #53

HEAT SHOCK FACTOR 1 (HSF1) INHIBITION SENSITIZES THE MULTIPLE MYELOMA CELL LINE MM.1S TO BORTEZOMIB-INDUCED APOPTOSIS Shardule P Shah and Lawrence H Boise, PhD Department of Hematology and Medical Oncology, Winship Cancer Institute, Emory University, Atlanta, GA, USA

Proteasome inhibition (PI) is a front-line therapy for multiple myeloma. However, myeloma cells upregulate genes in order to avoid PI-induced toxicity. Many of these genes are part of the heat shock response and correspondingly, the heat shock protein (HSP) family. Therefore, targeting the HSP family and its regulators is a potential myeloma therapy with the goal of maximizing apoptosis for the myeloma cell. In fact, combinations of PI with HSP90 are currently being tested in clinical trials. We found that the combination of the proteasome inhibitor, Bortezomib (Velcade®), with siRNA-mediated silencing of the heat shock response regulator, heat shock factor 1 (HSF1), resulted in enhanced apoptosis in MM.1S. To begin to define the effectors of the heat shock response important in controlling Bortezomib activity, we determined which heat shock response genes are induced by Bortezomib in an HSF1-dependent manner. Of the 84 heat shock genes probed for mRNA expression, 10 genes whose expression was induced by Bortezomib showed >50% reduction in HSF1-silenced cells. One of these genes, CRYAB (alpha-crystallin B chain), has been shown to inhibit caspase-3 in astrocytes, but has not previously been associated with multiple myeloma and could provide new insight into how myeloma cells evade PI-induced apoptosis. These results provide support for investigating the heat shock protein family and its regulation in response to PI to increase the efficacy of myeloma therapy. Nida Shaikh, NHS Poster #54

PREVALENCE OF DIABETES AND IMPAIRED FASTING GLUCOSE IN ADULTS IN SAUDI ARABIA Nida Shaikh1, Azadeh Zebatian2, Abdulelah Kutbi3, KM Venkat Narayan1,2 1Nutrition and Health Sciences, Graduate Division of Biological and Biomedical Sciences, Laney Graduate School, Emory University, 2Global Health and Epidemiology, Rollins School of Public Health, Emory University, 3Non-Communicable Disease Department, Ministry of Health, Riyadh, Saudi Arabia

Background: With urbanization and sedentary lifestyles, there are reports of increasing diabetes and impaired fasting glucose (IFG) in Saudi Arabia. However, there is limited information on the current prevalence of diabetes and IFG in Saudi Arabia. Objective: To determine the prevalence of diabetes and IFG in Saudi Arabia. Methods: Data from STEPwise nationally representative Saudi Arabia survey 2003-2005 for 4336 adults, aged 15-64 years were used to assess the prevalence of diabetes and IFG in Saudi Arabia. Using SAS (version 9.3), data were analyzed to classify individual as having diabetes, IFG, and normal (no diabetes). Results: Overall, prevalences’ of diabetes and IFG in Saudi Arabia were 22% and 13%, respectively. Prevalence of diagnosed diabetes and undiagnosed diabetes were 16% and 8%, respectively. Prevalence of diabetes was higher in older adults >45 years (37%), illiterate or less educated (31%), overweight and obese (25%), and the affluent (32%) compared to their counterparts. Prevalence of IFG was higher in obese (17%) and early-to-middle aged adults (16%), compared to their counterparts. Conclusion: Diabetes and IFG is a rising concern in Saudi Arabia, especially at younger ages. Early detection and prevention of increase in these conditions are paramount.


James Solyst, NS Poster #55

EFFECTS OF INTRANASAL OXYTOCIN ON SOCIAL ATTENTION AND MEMORY IN RHESUS MACAQUES J.A. Solyst1,2,3, E.A. Buffalo1,2,3,4,5 1Yerkes National Primate Research Center, Atlanta, GA, USA, 2Neuroscience Program, Emory University, Atlanta, GA, USA, 3Center for Translation Social Neuroscience, Atlanta, GA, USA, 4Center for Behavioral Neuroscience, Atlanta, GA, USA , 5Neurology, Emory University School of Medicine, Atlanta, GA, USA

Disruptions in attention and memory for socially relevant stimuli are distinctive features of several psychiatric disorders, particularly autism spectrum disorders (ASD). Eye tracking methodologies have proved to be a sensitive measure for assessing these disruptions, with the degree of attention towards faces and objects embedded in natural scenes strongly predicting subsequent diagnosis of autism in infants. By using similar tasks in rhesus monkeys, we can assess the effects of novel therapeutics on the same social behaviors that are altered in autism. One such therapeutic is oxytocin (OT), a neurohypophyseal peptide that is currently being tested in clinical trials as a treatment for social impairments in autism and schizophrenia. Here we tested the effects of OT on attention towards and memory for social and non-social items shown in visual scenes. Monkeys freely viewed static visual scenes comprised of novel objects and unfamiliar rhesus monkeys while their point of gaze was monitored with an infrared eye tracker. In alternating sessions, either saline or OT was administered through a pediatric nebulizer 40 minutes prior to scene presentation. In each session, ninety novel scenes were each shown twice, with thirty scenes repeated without change, and sixty scenes repeated with one item replaced by a new item to test the subject’s memory. Compared to saline, OT administration increased the proportion of time spent viewing monkeys with a visible face compared to monkeys without a visible face. By investigating OT’s effects on social attention and memory in rhesus monkeys we can begin to understand the mechanisms that may underlie it’s effects in humans.

Sharon Soucek, NS Poster #56

A ROLE FOR THE RNA BINDING PROTEIN, NAB2, IN QUALITY CONTROL Sharon Soucek1, Megan Bergkessel2, Deepti L. Bellur3, Christine Guthrie2, Jonathan P. Staley3, Anita H. Corbett1* 1Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA 2Department of Biochemistry and Biophysics, University of California, San Francisco, California 94158, USA.3Graduate Program in Cell and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA.

RNA binding proteins regulate the fate of RNA transcripts by modulating post-transcriptional events dictating cellular function. An essential zinc-finger nuclear polyadenosine RNA binding protein, Nab2, has been implicated in control of poly(A) tail length and mRNA export in budding yeast; however, a role for this protein in splicing has not been explored. To globally examine splicing of endogenous intron-containing transcripts, we performed a splicing microarray using nab2 mutant cells. A mild constitutive splicing defect was detected in these nab2 mutant cells. Consistent with a role for Nab2 in modulating splicing, we also detected defects in splicing in an in vitro splicing assay. To begin to understand which splicing step depends on Nab2, we tested for genetic interactions between NAB2 alleles and splicing factors required at different steps in splicing. We observed synthetic lethality with splicing factors required early in splicing as well as nuclear retention of unspliced transcripts. Alleles of NAB2 also genetically interact with the mRNA decay machinery, demonstrating that Nab2 acts as a nuclear watchdog to prevent accumulation and export of aberrant unspliced mRNAs. Our findings identify a role for Nab2 in regulating splicing and offer insights into how splicing and quality control are coupled in S. cerevisiae.


Sara Stahley, BCDB Poster #57

MEMBRANE RAFT MICRODOMAINS ARE PLATFORMS FOR DESMOSOME REGULATION Sara N. Stahley1, Masataka Saito1, Victor Faundez1, Michael Koval1,3 and Andrew P. Kowalczyk1,2

1Department of Cell Biology, 2Department of Dermatology and 3Division of Pulmonary, Allergy and Critical Care Medicine, Emory University School of Medicine, Atlanta, GA

Strong intercellular adhesion is critical for tissues that experience mechanical stress, such as the skin and heart. Desmosomes provide adhesive strength by anchoring desmosomal cadherins of neighboring cells to the intermediate filament cytoskeleton. Alterations in assembly and disassembly dynamics compromise desmosome function and cause disease states, such as the autoimmune blistering disease pemphigus vulgaris (PV). We have previously demonstrated that PV auto-antibodies (IgG) directed against the desmosomal cadherin desmoglein 3 (Dsg3) cause loss of adhesion by triggering membrane raft-mediated Dsg3 endocytosis. We hypothesized that rafts play a broader role in desmosome homeostasis by regulating the dynamics of desmosome assembly and disassembly. In human keratinocytes, Dsg3 is raft associated as determined by biochemical and super resolution microscopy methods. Interestingly, Dsg3 did not partition to rafts in cells that do not form desmosomes. Cholesterol depletion, which disrupts rafts, prevented desmosome assembly and adhesion, thus functionally linking rafts to desmosome formation. Additionally, PV IgG causedclustering andredistribution of Dsg3 into raft-containingendocytic carries. Furthermore, cholesterol depletion protected against PV IgG-induced desmosome disassembly. These findings demonstrate a requirement for functional rafts to facilitate Dsg3 assembly and disassembly dynamics, thus implicating raft manipulation as a promising therapy for desmosomal diseases such as PV. Andrew Swanson, NS Poster #58

INHIBITION OF RHO-KINASE PROMOTES GOAL-DIRECTED DECISION-MAKING Andrew M. Swanson1,2, Anthony J. Koleske3,4,5, Shannon L. Gourley1,2

1 Graduate Program in Neuroscience, Emory University, 2 Department of Pediatrics, Emory University , 3 Department of Molecular Biophysics and Biochemistry, Yale University, 4 Interdepartmental Neuroscience Program, Yale University, 5 Department of Neurobiology, Yale University

Goal-directed decision-making is based on the association between an action and a desired outcome. With repeated performance, behaviors that are initially goal-directed can assume stimulus-response, or ‘habitual,’ qualities. A range of insults, including drug and stressor exposure, can predispose both humans and animals to the formation of premature habits that occur at the expense of engagement in goal-directed action-outcome response strategies. A growing literature has identified mechanisms by which decision-making shifts from action-outcome-based to stimulus-response-based habit systems, but reversing habits has proven difficult. Using contingency degradation and outcome devaluation techniques, we show that systemic administration of fasudil, an inhibitor of Rho kinase (ROCK), a key regulator of the actin cytoskeleton, restores goal-directed decision-making in 'over-trained' mice that are otherwise engaged in stimulus-response habits. We provide evidence using wild type mice and mice deficient in Abl-related gene (Arg) that decreased deep-layer dendritic spine density selectively within the prelimbic prefrontal cortex is predictive of enhanced goal-directed decision-making. Moreover, local infusions of fasudil into the prelimbic cortex, but not the adjacent cingulate cortex, promote goal-directed decision-making. Together, these findings support the perspective that ROCK inhibition promotes goal-directed decision-making, possibly by enhancing the plasticity of deep-layer medial prefrontal cortical dendritic spines.


Virginia Vachon, MMG Poster #59

MECHANISMS OF 2’-5’ OLIGOADENYLATE SYNTHETASE REGULATION BY VIRAL NON-CODING RNA Virginia K. Vachon1,2, Turnee Malik1 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. Atlanta GA

The double-stranded (ds)RNA-activated protein kinase (PKR) and 2’-5’ oligoadenylate synthetase 1 (OAS1) are activated upon binding to dsRNA produced as a consequence of viral replication and act as a first line of defense in the human innate cellular anti-viral response. The small non-coding RNA transcripts VA RNAI and EBER1 are used by Adenovirus and Epstein-Barr virus, respectively, to subvert this system by inhibiting PKR, blocking its inactivation of cellular protein synthesis and ensuring continued production of viral proteins by host translational machinery. Counterintuitively, both RNAs activate rather than inhibit OAS1 leading to downstream RNase L activation and translational repression. However, in vivo the cellular RNase Dicer processes VA RNAI to a truncated fragment, transforming it into a pseudo-inhibitor of OAS1. The question of whether this processed RNA fails to activate OAS1 due to loss of a specific consensus activation sequence, or a particular structural feature is addressed here. Further, structural similarity between VA RNAI and EBER1 leads to the novel hypothesis that similar processing occurs for EBER1 that may affect its localization and activity. Our goal is to identify structural hallmarks responsible for the activities of these RNAs that define their roles in viral infection. Kellie Vinal, MMG Poster #60

MECHANISMS OF PATHOGEN-DERIVED 16S RRNA METHYLTRANSFERASE NPMA 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, Atlanta, GA

The 16S ribosomal RNA (rRNA) methyltransferase enzyme NpmA methylates nucleoside adenosine 1408 (A1408) in the bacterial ribosome aminoglycoside binding site, conferring resistance to these antibiotics. As the first identified pathogen-derived A1408 rRNA methyltransferase, it is crucial to characterize the mechanism by which NpmA recognizes its target and initiates enzymatic activity. Like other 16S rRNA methyltransferases, NpmA binds its cofactor S-adenosyl-L-methionine (SAM) in a structurally conserved SAM-binding fold. NpmA additionally possesses three unique extended structures, one at the N-terminus and two between the final three β-strands of the core SAM-binding fold, that we hypothesize should contain the major determinants of specific 30S target site recognition. Using data from 16S rRNA probing experiments, amino acid residues predicted to specifically contact the 30S subunit and play a role in target recognition were identified and mutated. The resulting recombinant enzymes were tested in kanamycin minimum inhibitory concentration (MIC) assays, and expressed and purified for analyses including in vitro methylation and 30S binding assays. Identifying key residues involved in NpmA recognition of its target will provide us with insights that will further our understanding of A1408 rRNA methyltransferase mechanisms of action and ultimately provide a platform for development of inhibitors to combat increasing antibiotic resistance.


Jadiel Wasson, BCDB Poster #61

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, Atlanta, GA

Epigenetic modificationsare 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 formthe totipotent zygote. This massive change in cell fate at fertilization implies there is a need to erase the gametic transcriptional program in order tore-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 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.

Kevin Watkins, NS Poster #62

THE EFFECT OF DEMONSTRATOR’S AGE ON SOCIAL LEARNING IN CAPTIVE CHIMPANZEES (PAN TROGLODYTES) K. E. Watkins1, L. A. Parr1,2,3, J. G. Herndon1, V. Horner4, T. M. Eppley4, and F. B. M. de Waal4

1Yerkes National Primate Research Center, Emory University, Atlanta, GA, 30329, USA; 2Center for Translational Social Neuroscience, Emory University; 3Division of Psychiatry and Behavioral Sciences, Emory University; 4Living Links Center, Yerkes National Primate Research Center, Emory University, Atlanta, GA

Research has demonstrated the capacity for cultural transmission in chimpanzees, but few studies have focused on directed social learning, where the identity and characteristics of the demonstrator affect the probability of social learning. Exploring individual factors influencing this choice to selectively imitate – such as age, sex, or dominance display – will inform the evolutionary history of directed social learning. In this study, we begin to isolate key differences between demonstrators and assess their influence on directed social learning. Chimpanzee subjects were shown video of unknown conspecific demonstrators solving a two-solution, grape-dispensing puzzle box. Demonstrators of different ages were presented solving the puzzle box in different ways (“push” vs. “slide”). One group (n=14) viewed video footage of the older demonstrator solving the puzzle box using the push technique and the younger demonstrator using the slide technique. Another group (n=15) viewed the same demonstrators, but the techniques used were reversed. The solution presented by the older demonstrator significantly affected the subject’s first attempt at solving the puzzle box. Our results indicate that age bias in social learning may be conserved across species. This approach has significant applications in ongoing research designed to assess the role of other social cues on social learning.


Amanda Jo Williams-Newkirk, PBEE Poster #63

CANDIDATUS MIDICHLORIA MITOCHONDRII IS PRESENT AND GENETICALLY VARIABLE IN THE LONE STAR TICK AMBLYOMMA AMERICANUM Amanda Jo Williams-Newkirk1, 2, Lori A. Rowe2, Tonya R. Mixson-Hayden2, and Gregory A. Dasch2 1Graduate Program in Population Biology, Ecology and Evolution, Emory University, 2US Centers for Disease Control and Prevention, Atlanta, GA, USA

The bacterium “Candidatus Midichloria mitochondrii” was detected for the first time in lone star ticks (Amblyomma americanum) from the eastern United States. We tested 177 individuals and 11 tick pools from seven sites in four states by pyrosequencing with barcoded 16S rDNA eubacterial primers targeting variable regions 5-3. Average infection prevalence was 0.15 across all surveyed populations (range 0-0.29) and only the site with the lowest sample size (n=5) was negative. Three genotypes differing by 2.58-4.06% in a 271 bp region of 16S rDNA were identified. Two variants co-occurred in sites in North Carolina and New York, but were not observed in the same tick at those sites. A third genotype was found only in Georgia. Phylogenetic analysis of this fragment indicated that the three variants are more closely related to “Candidatus Midichloria mitochondrii” genotypes from other tick species than to each other. This variation suggests that multiple independent introductions occurred in A. americanum which may provide insight into bacterial spread within its ecosystem. Whether the presence of this bacterium affects acquisition or maintenance of pathogens and symbionts in A. americanum or the biology and evolution of the tick itself is unknown.

Kathryn R. Williams, BCDB Poster #64

REPRESSION OF GAP-43 EXPRESSION BY THE MRNA BINDING PROTEIN HNRNP-Q1: INVESTIGATING A LOCALMECHANISM IN NEURONS K.R. Williams, L. Xing, G.J. Bassell

Department 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 recent 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 translationas 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.


Philip Zakas, MSP Poster #65

REDUCED ANTIGENICITY OF RECOMBINANT OVINE FACTOR VIII IN HEMOPHILIA A INHIBITOR PATIENT PLASMAS Philip M Zakas1, Shannon Meeks, MD2 and Christopher B Doering, PhD2 1Graduate Program in Molecular and Systems Pharmacology, Emory University, Atlanta, GA 2Aflac Cancer and Blood Disorders Center, Emory University/Children’s Healthcare of Atlanta, Atlanta, GA

Hemophilia A is an X-linked disorder caused by defects or deficiencies in coagulation factor VIII (fVIII). Approximately 20-30% of patients with severe hemophilia A develop antibodies against fVIII (inhibitors) following fVIII replacement therapy. Previously, plasma-derived porcine fVIII was an effective treatment option for inhibitor patients due to reduced antigenicity toward anti-human fVIII inhibitors. However, due to concerns regarding viral contamination, the plasma-derived porcine fVIII products were discontinued with no alternative option. Recently, a line of hemophilia A sheep was reestablished and the ovine fVIII (ofVIII) protein was characterized. We hypothesized that ofVIII confers differential antigenicity compared to human and/or porcine fVIII in congenital and acquired inhibitor patient plasmas. The reactivity of 28 plasma samples towards recombinant BDD human, porcine, and ovine fVIII were assessed by ELISA. Data revealed reduced reactivity towards ofVIII in 27 of 28 total samples. Furthermore, reactivity to ovine fVIII was significantly reduced compared to porcine fVIII (P = 0.025). Inhibitor analysis of samples revealed 22 patients with diminished titers against ofVIII. Furthermore, four acquired hemophilia A patients with compromising titers against hfVIII displayed titers near zero against ofVIII. These results suggest that recombinant fVIII molecules may be potential therapeutics for patients harboring inhibitors to human fVIII. Monica Chau, NS Poster #66

iPS CELL TRANSPLANTATION THERAPY INCREASES NEUROGENESIS AND ANGIOGENESIS IN NEONATAL RATS WITH ISCHEMIC STROKE Monica J. Chau1,2, Dongdong Chen2, Shan Ping Yu2, Ling Wei2 1Neuroscience Graduate Program. 2Department of Anesthesiology, Emory University School of Medicine, Atlanta, GA 30322

Stroke is a leading cause of death and disability. Neonatal stroke affects 26 of 100,000 live births each year, yet there are limited treatments available. Induced pluripotent stem (iPS) cells have provided a new source of stem cells that circumvents immune rejection and the ethical issues of using embryonic stem cells. However, the therapeutic benefits of iPS cell transplantation is relatively unknown. We hypothesized that iPS cell-derived neural progenitors provide cell replacement and beneficial growth factors to the stroke area. Seven days after stroke induction in the neonates, iPS progenitors were transplanted, received BrdU injections, and were sacrificed at 21d. iPS cell-derived neurons were patch-clamped to characterize activity. Stroke rats that received transplantation showed an increase of total BrdU-positive cells and an increase of BrdU/NeuN and BrdU/CollagenIV-labeled cells. Immunocytochemistry showed expression of trophic factors including VEGF, EPO, SDF-1, FGF, and GDNF in iPS neural progenitors. iPS cell-derived neurons exhibited neuronal functionality. The increased number of new neurons and vessels suggests increased neurogenesis and angiogenesis with iPS cell transplantation. Transplantation of iPS cells may contribute additional trophic support to increase endogenous progenitor migration to the infarct, cell survival, and endogenous neurogenesis. iPS cell transplantation is a possible therapeutic for neonatal stroke.


Karen R. Siegel, NHS Poster #67

SOCIETAL CORRELATES OF HYPERTENSION AND IMPLICATIONS FOR POLICY Karen R. Siegel1, Justin B. Echouffo-Tcheugui2, Mohammed K. Ali1, Neil K. Mehta1, K.M. Venkat Narayan1,3, 1Hubert Department of Global Health and Epidemiology, Rollins School of Public Health, Emory University, 2Johns Hopkins School of Medicine, 3School of Medicine, Emory University, Atlanta, GA

One-third of adults worldwide have hypertension. While we know that societal-level influences modify individual-level hypertension risk, the extent of their influence is unknown. Given important implications for identifying policy levers, we quantified relationships between societal factors and hypertension. We extracted recent estimates for country-level variables: hypertension prevalence; total caloric, animal fat, fruit and vegetable availability; salt consumption; physical inactivity markers (vehicles/capita and value-added from service sector); gross domestic product/capita (GDP); age-adjusted mortality rate. We used generalized linear models to investigate relationships between these factors and hypertension. Median global hypertension prevalence was 41.7% in 2008. Salt intake, measured as urinary sodium excretion or processed/packaged food volume, was not associated with hypertension (OR for mmol/day: 0.99, 95% CI 0.99-1.00, p=0.104 and OR for retail volume in kg/capita: 0.99 95%CI: 0.998-1.00, p=0.513, respectively), after adjusting for all other variables. Caloric availability (OR 1.03 for each kilocalorie/person/day, 95% CI 1.002-1.05, p=0.033) and animal fat availability (OR 1.04 for each percentage of animal fat per total calories/person/day, 95% CI 1.02-1.07, p=0.002) were significantly associated with hypertension. Although reduced salt intake has been prioritized for preventing hypertension, our results suggest that additional factors may have stronger ecological associations with hypertension. Interventions should consider these multiple factors.

Kristen Stout, MSP Poster #68

A CELLULAR MODEL TO ASSESS PLASMA MEMBRANE AND VESICULAR TRANSPORT OF NOREPINEPHRINE Kristen A Stout, Alison I Bernstein, Yingjie Li, Thomas S Guillot, Gary W Miller

Environmental exposure to neurotoxic compounds plays an important role in Parkinson’s disease (PD) development. Many neurotoxic compounds act as inhibitors of the vesicular monoamine transporter 2 (VMAT2). The primary role of VMAT2 is to sequester monoamines into vesicles, thereby protecting them from cytosolic metabolism. Inhibition of vesicular packaging results in increased neurotransmitter metabolism. Neurotransmiter metabolites can inhibit mitochondrial complex I, a likely cause of toxicity. Though the effect of neurotoxic chemicals in dopaminergic systems has been established, little has been done to characterize VMAT2 inhibition within norepinephrine neuron models. We are utilizing a monoamine-like fluorescent substrate (Molecular Devices) to investigate VMAT2 function in HEK cells stably expressing the human norepinephrine transporter (NET) and mCherry-tagged human VMAT2. The substrate is transported by NET. Desipramine, a competitive inhibitor of NET, potently blocks substrate transport (IC50 = 113.9 nM). Within the cell, the dye is sequestered into vesicles by VMAT2, evidenced by co-localization of fluorescence between the substrate and mCherry. Tetrabenazine, a VMAT2 inhibitor, inhibits co-localization in a dose dependent manner. Both co-localization and inhibition are detectable by microscopy. This cellular model allows us to assess both normal and altered transporter function in response to toxicants in a noradrenergic cellular model.


ACKNOWLEDGEMENTS

The GDBBS Division Student Advisory Council (DSAC) would first like to express our sincerest gratitude to all of the participants-students, faculty, post-docs, and alumni- who made this 10th 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.

welcome to the tenth annual division student · 2013. 4. 17. · the tenth annual dsac student...

Documents