SCHEDULE OF EVENTS SATURDAY, APRIL 27 11:00-1:30PM POSTER SESSION VENDOR/RESEARCH SERVICES FAIR Main Lounge *lunch served 11:30-1:30PM 1:15-1:30PM VA PRESENTATION “The Impact of VA Research on Veterans’ North Room Health” Dr. Jane K. Battles, PhD Scientific Program Manager, Biomedical Laboratory and Clinical R&D Services Office of Research and Development Department of Veteran Affairs, Washington DC 1:30-3:15PM MINI-SYMPOSIA North Room, South Room & Iowa Theater *see Mini-Symposia Program for details 3:30-4:30PM KEYNOTE LECTURE “Catalyzing Translational Innovation” 2nd Floor Ballroom Dr. Christopher Austin, MD Director, National Center for Advancing Translational Sciences (NCATS) National Institutes of Health 4:30-6:00PM RECEPTION Main Lounge

KEY TO POSTER ENTRIES CCOM Carver College of Medicine COP College of Pharmacy ICTS Institute for Clinical & Translational Science VA Iowa City VA Medical Center

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MINI-SYMPOSIA SCHEDULE

NORTH ROOM SOUTH ROOM IOWA THEATER

1:30-1:45PM Wade Gutierrez “Epigenetic Combination Therapy Improves Survival and Promotes Tumor-Specific Immunomodulation in Soft Tissue Sarcoma”

Nitija Tiwari “Regulation of the S. aureus Survival and Pathogenicity by Redox Sensing Histidine Kinase SrrB”

Colleen Caldwell “Single molecule observation of the Rad52 recombination mediator mechanism”

1:45-2:00PM Benton Purnell “Breathing is Subject to Endogenous Circadian Regulation Which is Dependent on the Suprachiasmatic Nucleus”

Garima Dixit “The Role of ADAMs in FGFR2-mediated Trans-Activation of EGFR/MAPK Signaling and NOTCH Pathway in Endometrial Carcinomas”

Tingting Duan “An atypical mode of mitosis is involved in germline stem cell maintenance”

2:00-2:15PM Zeb Zacharias “Polyanhydride Nanoparticle-Based Vaccination Provides Robust Protection Against Influenza A Virus Infections”

Kaylia Duncan “Searching for the Missing Heritability for Orofacial Clefting in the Periderm Gene Regulatory Network”

Jordan Kohlmeyer “Targeting a Novel RABL6A-RB1 Pathway Suppresses MPNST Pathogenesis”

2:15-2:30PM BREAK

BREAK BREAK

2:30-2:45PM Kathryn Spitler, PhD “Tissue Specific Actions of Angptl4 During Long-Term High Fat Diet Feeding”

Satya Geesala, PhD “iRhom2 Protects II10-deificient Mice from Colitis by Regulating Regenerative Activities During Immune Responses”

Victoria Muller Ewald “When to seek and when to stop: how activity in the infralimbic cortex changes during the suppression of cocaine-seeking behavior”

2:45-3:00PM Antentor Hinton, PhD “OPA-1 Deficiency Increases Mitochondria-Endoplasmic Reticulum Contact Formation in Skeletal Muscle”

Megan Schmidt “Pre-Existing Neutralizing Antibodies Prevent CD8 T cell-mediated Immunopathology Following Respiratory Syncytial Virus Infection”

Rebecca Taugher, PhD “The amygdala differentially regulates defensive behaviors evoked by CO2”

3:00-3:15PM Adam Rauckhorst, PhD “Rapid Hypoxia-Induced Metabolomic Changes Parallel Delayed Postmortem Tissue Sampling”

Diogo Valadares “Myeloid and Lymphoid Immune Exhaustion Profile During Murine Visceral Leishmaniasis”

Daniel Infield, PhD “Atomic Mutagenesis of Voltage Sensing in Ion Channels”

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NORTH ROOM 1:30-1:45PM Wade Gutierrez, Graduate Student, MSTP/Cancer Biology Mentor: Dr. Rebecca Dodd, Internal Medicine “Epigenetic Combination Therapy Improves Survival and Promotes Tumor-Specific Immunomodulation in Soft Tissue Sarcoma” Wade Gutierrez, Amanda Scherer, Vickie Knepper-Adrian, Emily Laverty, Gavin McGivney, Victoria Stephens, Varun Monga, Rebecca Dodd Sarcomas are a diverse group of connective tissue tumors comprising approximately 1% of adult cancers and 15% of pediatric cancers. The profound heterogeneity of sarcomas presents a unique challenge when trying to understand and treat these malignancies. To date, few molecular targets have been identified for chemotherapeutic treatment. In collaboration with an ongoing Phase 1b clinical trial at the University of Iowa, we are investigating a novel combination therapy using standard-dose Gemcitabine (Gem), an antimetabolite, with low-dose Decitabine (DAC), a DNA methyltransferase inhibitor, for the treatment of soft-tissue sarcoma. To test the efficacy of Gem+DAC treatment and to elucidate its therapeutic mechanism, we are using a primary mouse model of undifferentiated pleomorphic sarcoma, the most common form of adult soft tissue sarcoma. Our data show that Gem+DAC combination treatment slows tumor growth and extends survival better than single-agent treatment alone. We have also found increased tumor-specific immune infiltration in Gem+DAC-treated mice. In addition to characterizing tumor immune cell infiltration, we are conducting a co-clinical trial examining biological correlates of treatment efficacy by analyzing immune cell profiles and cytokine signatures in longitudinal blood samples from Gem+DAC-treated mice and from patients from our Phase Ib clinical trial collaboration. 1:45-2:00PM Benton Purnell, Graduate Student, Neuroscience Mentor: Dr. Gordon Buchanan, Neurology “Breathing is Subject to Endogenous Circadian Regulation Which is Dependent on the Suprachiasmatic Nucleus” Benton Purnell, Gordon Buchanan Sudden unexpected death in epilepsy (SUDEP) is the leading cause of premature death in the 50 million people worldwide who live with epilepsy. It is not clear what conditions differentiate seizures which result in death from the countless others that do not; however, SUDEP typically happens consequent to respiratory arrest following a seizure during the night. Breathing is different at different times of day, but the mechanism for this is unknown. Day-night differences in breathing may alter a patient’s vulnerability to seizure induced respiratory arrest. We hypothesized that breathing is subject to endogenous circadian regulation by the suprachiasmatic nucleus (SCN). To test this hypothesis, we monitored breathing, both at baseline and in response to inhaled 7% CO2, using whole-body plethysmography at different times of day in normally entrained mice, at different circadian phases in free-running mice, and at different time points in mice which had received an SCN lesion or a sham surgery. Circadian rhythmicity in breathing was observed in constant conditions; however, these oscillations were abolished following SCN lesion, but not the sham surgery. The results of these experiments suggest that breathing is subject to endogenous circadian regulation which is dependent on the SCN.

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2:00-2:15PM Zeb Zacharias, Graduate Student, Pathology Mentor: Dr. Kevin Legge, Pathology “Polyanhydride Nanoparticle-Based Vaccination Provides Robust Protection Against Influenza A Virus Infections” Zeb Zacharias, Kathleen Ross, Emma Hornick, Jonathan Goodman, Balaji Narasimhan, Thomas Waldschmidt, Kevin Legge Influenza A virus (IAV) infections cause serious respiratory illness resulting in significant morbidity and mortality, worldwide. These negative impacts on human health, along with the associated economic burden, collectively stress the need to develop novel and efficacious IAV vaccination strategies. Recent studies indicate that in order to provide optimal protection against IAV infections, IAV vaccination strategies need to generate tissue-resident memory T and B cells at the local site of the infection (i.e. nasal mucosa and lung). Unfortunately, currently licensed IAV vaccines primarily induce systemic IAV-specific antibody responses and induce little to no lung-resident memory populations. Accordingly, our primary objective is to construct an IAV vaccine capable of inducing lung-resident T and B cells. Here, we demonstrate the efficacy of a biodegradable polyanhydride nanoparticle-based IAV vaccine (IAV-nanovax) to generate lung-resident, cross-protective immune responses. Our results illustrate that an intranasal inoculation with IAV-nanonax provides mice with protection against subsequent homologous and heterologous IAV challenges. During these challenges, IAV-nanovax vaccinated mice exhibited reduced viral titers within the lungs and also possessed lung-resident, IAV-specific CD4+ and CD8+ T cells. Furthermore, IAV-nanovax treated mice had lung-resident germinal center B cells responses and produced IAV-specific IgG and IgA responses. Altogether, our results establish the concept that polyanhydride nanoparticles have the capacity to induce tissue-resident immune responses that provide protection against homologous and heterologous IAV infections. 2:15-2:30PM BREAK 2:30-2:45PM Kathryn Spitler, PhD, Postdoctoral Research Scholar, Biochemistry Mentor: Dr. Brandon Davies, Biochemistry “Tissue Specific Actions of Angptl4 During Long-Term High Fat Diet Feeding” Kathryn Spitler, Emily Cushing, Shwetha Shetty, Brandon Davies Metabolic diseases, such as diabetes mellitus and obesity carry an increased risk for cardiovascular disease. A common feature of these diseases is elevated plasma triglyceride (TG) levels. Circulating TGs are hydrolyzed by lipoprotein lipase (LPL). An important regulator of LPL is angiopoietin-like 4 (Angptl4). Angptl4 is highly expressed by the liver and adipose tissues. Genetic loss of Angptl4 results in decreased plasma TG levels in both humans and mice. In mice, adipose-specific, but not liver-specific loss of Angptl4 leads to decreased fasting plasma TG levels. The aim of this study is to unravel the role of liver and adipose-specific deletion of Angptl4 during obesity. Liver specific or adipose-specific Angptl4cKO mice (Angptl4-LKO/AKO) were generated. At 8 weeks of age mice were fed a normal chow diet (NCD) or a high fat diet (60% kCal/fat; HFD) for 6 months. After 16 weeks and 6 months on NCD or HFD Angptl4-AKO mice had decreased fasting plasma TG levels compared to floxed controls; no differences were seen in fasting plasma TG levels in Angptl4-LKO mice. Angptl4-AKO mice gained more weight than their floxed controls while there was no difference in weight gain between the Angptl4-LKO mice versus floxed controls. Angptl4-AKO are protected from insulin resistance after 16 weeks of HFD but not after 24 weeks of HFD feeding. Angptl4-LKO mice are protected from insulin resistances at both 16 or 24 weeks on HFD. We have found that although there is no fasting TG phenotype, Angptl4-LKO mice are protected from HFD-induced insulin resistance.

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2:45-3:00PM Antentor Hinton, PhD, Postdoctoral Research Fellow, Internal Medicine Mentor: Dr. E. Dale Abel, Internal Medicine “OPA-1 Deficiency Increases Mitochondria-Endoplasmic Reticulum Contact Formation in Skeletal Muscle” Margaret Mungai, Youngdo Koo, Ben Kirk, Leo Kazma, Jacob Lam, Jessica Ponce, Rhonda Souvenir, Chad Grueter, Renata Pereira, E. Dale Abel, Defective mitochondria endoplasmic reticulum (ER) contact sites (MERCs) have been associated with insulin Resistant Type 2 Diabetes Mellitus (T2D). MERCs are hubs enriched with specific proteins and lipids believed to mediate inter-organellar communication such as calcium transfer, lipid transfer, and autophagosome formation. We have previously demonstrated that OPA-1 deficiency in skeletal muscle induces ER stress, which correlated with upregulation of Mitofusin-2 (MFN-2), a known tethering protein in MERCs. Therefore, we hypothesized that OPA1-induced mitochondrial stress results in narrowing of MERC distance through the upregulation of tethering proteins. To test this hypothesis, we surveyed for several MERC proteins in OPA1-deficient skeletal muscle and primary myotubes. Primary myotubes were generated by isolating satellite cells from OPA-1 floxed mice and subsequently deleting OPA1 by infecting the cells with adenoviral Cre recombinase. Ablation of OPA-1 in myotubes increased the following MERC proteins: MFN-1, MFN-2, and BIP. Similarly, loss of OPA-1 in skeletal muscle increased MFN-1, MFN-2, PACS-2, GRP75, BIP, and IP3R. Analysis of OPA-1 deficient cells by confocal imaging revealed an increase in MERCs, estimated by GRP-78 colocalization with MitoTracker after Z series reconstruction. TEM Analysis of OPA-1 deficient myotubes and soleus skeletal muscle confirmed an increase in the number of MERCs and a reduced distance between ER and Mitochondria. Collectively, these data suggest that loss of OPA-1 in muscle results in the induction of MERC proteins, which correlates with increased MERCS formation. Increased MERCs may represent a compensatory mechanism in response to mitochondrial stress that is precipitated by OPA1 deficiency. 3:00-3:15PM Adam Rauckhorst, PhD, Postdoctoral Research Fellow, Biochemistry Mentor: Dr. Eric Taylor, Biochemistry “Rapid Hypoxia-Induced Metabolomic Changes Parallel Delayed Postmortem Tissue Sampling” Adam Rauckhorst, Diego Scerbo, Eric Taylor, The metabolome is a biological system’s collection of enzymatic substrates and products. Metabolomic regulation is rapid, highly dynamic, and intrinsically linked to oxygen supply. Thus, loss of oxygen supply quickly propagates to broad metabolomic disruption. However, hypoxia-induced metabolomic changes elicited by tissue dissection are not well described. The implications of this absence for past and current metabolomic inquiries across the life sciences are far reaching. Here, we utilize GC- and LC-MS based metabolomics to examine the stability of the mouse liver metabolome during 10 minutes of acute hypoxia following tissue dissection. Striking metabolomic changes, detected by both principle component analysis and examination of individual metabolite levels, occurred within 30 seconds of tissue dissection. The magnitudes of these changes were amplified with time. Decreasing ATP levels coincided with markedly increasing AMP levels. Furthermore, glycolytic metabolites and lactate generally increased, whereas TCA cycle intermediate metabolites decreased. Levels of numerous additional metabolites including amino acids changed. These changes are consistent with increased anaerobic metabolism resulting from continued ATP demand during inhibited oxidative phosphorylation while attempting to maintain ATP levels via adenylate kinase activity. These results demonstrate the metabolome is exquisitely sensitive to hypoxia caused by loss of perfusion from dissection. This study illustrates the importance of freezing samples within seconds of dissection to minimize hypoxic insult and to obtain the most accurate portrait of the in vivo metabolome.

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SOUTH ROOM 1:30-1:45PM Nitija Tiwari, Graduate Student, Biochemistry Mentor: Dr. Ernie Fuentes, Biochemistry “Regulation of the S. aureus Survival and Pathogenicity by Redox Sensing Histidine Kinase SrrB” Nitija Tiwari, Zhen Xu, Young Joo Sun, Marisa Lopez-Redondo, Laura Miguel-Romero, Hassan Al-Tameemi, Alberto Marina, Jeffery Boyd, John McCormick, John Kirby, Patrick Schlievert, Ernesto Fuentes Staphylococcus aureus infections can lead to diseases that range from skin abscess to life threatening toxic shock syndrome. The SrrAB two-component system (TCS) is a global regulator of S. aureus virulence and critical for survival under the environmental conditions (hypoxia, oxidative and nitrosative stress) found at sites of infection. Despite the critical role of SrrAB in S. aureus pathogenicity, the signal(s) that SrrB histidine kinase senses and responds to, and the mechanism by which it is regulated remain unknown. The SrrB histidine kinase contains several domains, including an extracellular Cache domain and a cytoplasmic HAMP-PAS-DHpCA catalytic region. Here, we show that the PAS domain regulates both kinase and phosphatase enzyme activity of SrrB. Moreover, we show that heme binding to the PAS domain influences autophosphorylation and oligomerization. Small angle X-ray scattering analysis of the HAMP-PAS-DHpCA region indicates that the PAS and CA domains are flexibly linked and capable of domain-domain interactions, suggesting a mechanism by which the PAS domain might regulate enzyme activity. We also present the structure of the DHpCA catalytic domain. Importantly, this structure shows a cysteine disulfide bond in the CA domain that significantly affects autophosphorylation kinetics. Our in vivo data shows that the redox state of these cysteine residues affect S. aureus biofilm formation and Toxic Shock Syndrome Toxin -1 production. Together, our data are consistent with a model whereby the SrrB histidine kinase senses and responds the cellular redox environment through heme and a disulfide bond to support S. aureus survival and pathogenesis. 1:45-2:00PM Garima Dixit, Research Associate, Internal Medicine/Inflammation Program Mentor: Dr. Thorsten Maretzky, Internal Medicine “The Role of ADAMs in FGFR2-mediated Trans-Activation of EGFR/MAPK Signaling and NOTCH Pathway in Endometrial Carcinomas” Garima Dixit, Wenhui Zhou, Jesus Gonzalez Bosquet, Willow Schanz, Mikayla Biggs, Priya Issuree, Shujie Yang, Eric Devor, Kimberly Leslie, Thorsten Maretzky Endometrial cancer (EC) is the fourth most common cancer in women after breast, colorectal and lung cancer in the world. While only curable at an early stage through surgery and adjuvant radiotherapy, outcomes are poor for patients with metastatic or recurrent disease and the five-year survival rate for these patients is less than 15%. Somatic mutations of the receptor tyrosine kinase fibroblast growth factor receptor (Fgfr)2 isoform 2b occur in about 16% of endometrial carcinoma, giving the scope for identification of associated genomic events that contribute to the oncogenesis of this disease. Activation of FGFR2 mediates a disintegrin and metalloprotease (ADAM)17-dependent activation of the epidermal growth factor receptor (EGFR)/MAPK signaling pathway. Previous work has shown that crosstalk between EGFR signaling and Notch pathway can have an oncogenic role in several types of cancer. Since ADAMs are important mediators of EGFR and NOTCH pathway activation, we anticipated that somatic mutations in Fgfr2 might cause dysregulation of these two signal transduction pathways leading to oncogenic transformation in EC. Utilizing EC cells harboring FGFR2 mutations, we show that somatic mutations of FGFR2 were not constitutively active and required fibroblast growth factor (FGF)7 stimulation to display oncogenic potential. We demonstrate that FGFR2 mutations activate EGFR and NOTCH pathways upon FGF7 stimulation in a metalloprotease-dependent manner. Moreover, shedding of the EGFR-ligand and ADAM17-substrate HBEGF as well as of the ADAM10-substrate betacellulin was increased in FGF7/FGFR2 stimulated EC cells leading to downstream activation of EGFR and the NOTCH pathway. Inhibition of metalloprotease activity in these EC cells prevented

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FGF7/FGFR2-mediated oncogenic cell transformation as well as EGFR/MAPK and NOTCH signaling as evident through 2D-soft agar colony formation assays. Additionally, RNAseq analysis of EC patient samples also supported this observation and indicated strong association of somatic mutations in Fgfr2 gene with NOTCH and EGFR/MAPK signaling components. These findings provide new and exciting insights into driving oncogenic pathways in endometrial malignancies, and warrant future clinical studies with FGFR inhibitors for EC and other FGFR2-associated carcinomas. 2:00-2:15PM Kaylia Duncan, Graduate Student, Molecular Medicine Mentor: Dr. Robert Cornell, Anatomy and Cell Biology “Searching for the Missing Heritability for Orofacial Clefting in the Periderm Gene Regulatory Network” Kaylia Duncan, Robert Cornell Orofacial cleft is a common structural birth defect with a strong genetic underpinning. However, most of the heritable risk for it has not been assigned to any gene. Two genes associated with orofacial cleft, IRF6 and GRHL3, encode transcription factors that promote differentiation of periderm, the most superficial layer of embryonic skin and oral epithelium. Additional members of the gene regulatory network driving periderm differentiation are candidates to harbor variants that increase risk for orofacial cleft. Using the tractable zebrafish as an experimental model, we are identifying the additional transcription factors, and the regulatory hierarchy, of this network. This work will facilitate the prioritization of variants found in patients with orofacial cleft. 2:15-2:30PM BREAK 2:30-2:45PM Ramasatyaveni Geesala, PhD, Postdoctoral Fellow, Internal Medicine Mentor: Dr. Thorsten Maretzky, Internal Medicine “iRhom2 Protects II10-deificient Mice from Colitis by Regulating Regenerative Activities During Immune Responses” Ramasatyaveni Geesala, Willow Schanz, Mikayla Biggs, Garima Dixit, Joseph Skurski, Prajwal Gurung, David Elliot, David Meyerholtz, Priya Issuree, Thorsten Maretzky Inflammatory bowel disease (IBD) arises from complex interactions of genetic, environmental, and microbial factors, which represents a heterogeneous group of idiopathic inflammatory conditions of the colon and small intestine, affecting about 0.3% of the population. The key pathogenic role of Tumor necrosis factor (TNF) in chronic intestinal inflammation made it a major therapeutic target for IBD. Anti-TNF agents markedly reduced the progression of disease which was limited to 30% of patients and also developed long term intolerance. We analyzed the pathophysiological role of iRhom2, a crucial regulator of the maturation and function of ADAM17 (TNF-a Convertase) in immune cells to identify a molecular target that involves in the activation of TNF-a. IRhom2-deficient (Rhbdf2-/-) mice were crossed and bred with interleukin 10-deficient (Il10-/-) mice to generate Rhbdf2-/-/Il10-/- mice and offspring were observed for signs of colitis for 48 weeks. Phenotypic changes in immune cells were assessed by flow cytometry and gene expression analysis. Homozygous Rhbdf2-/-/Il10-/- mice developed spontaneous colitis and exhibited severe weight loss compared to Il10-/- mice within the first 16 weeks of age. Immunophenotyping analysis depicted a Th1-driven inflammation in Rhbdf2-/-/Il10-/- mice which was well correlated with significant increase in the expression of Th1-derived cytokines IFN-? and IL-2. Our data provide critical insights into how iRhom2 maintains intestinal homeostasis and establish a role for iRhom2/ADAM17 interaction in chronically stimulated sites such as the intestinal surface that participate in the activation and regulation of the TNF/EGFR signaling axis and how this pathway contributes in achieving the critical balance of protective immunity and inflammation.

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2:45-3:00PM Megan Schmidt, Graduate Student, Immunology Mentor: Dr. Steven Varga, Microbiology and Immunology “Pre-Existing Neutralizing Antibodies Prevent CD8 T cell-mediated Immunopathology Following Respiratory Syncytial Virus Infection” Megan Schmidt, Steven Varga Despite being a leading cause of severe respiratory disease, there remains no licensed respiratory syncytial virus (RSV) vaccine. Neutralizing antibodies reduce the severity of RSV-associated disease but are not sufficient for preventing reinfection. In contrast, the role of memory CD8 T cells in providing protection against a secondary RSV infection is less established. We recently demonstrated that high magnitude memory CD8 T cells efficiently reduced lung viral titers following RSV infection but induced fatal immunopathology that was mediated by IFN-gamma. To evaluate the ability of RSV-specific neutralizing antibodies to prevent memory CD8 T cell-mediated immunopathology, mice with high magnitude memory CD8 T cell responses were treated with neutralizing antibodies prior to RSV challenge. Neutralizing antibody treatment significantly reduced morbidity and prevented mortality following RSV challenge compared to IgG-treated controls. Neutralizing antibody treatment restricted early virus replication, which resulted in a substantial reduction in memory CD8 T cell activation and IFN-gamma production. In contrast, therapeutic administration of neutralizing antibodies did not impact morbidity, mortality, or IFN-gamma levels despite significantly reducing lung viral titers. Therefore, only pre-existing neutralizing antibodies prevent memory CD8 T cell-mediated immunopathology following RSV infection. Overall, our results have important implications for the development of future RSV vaccines. 3:00-3:15PM Diogo Valadares, Research Assistant, Internal Medicine Mentor: Dr. Mary Wilson, Internal Medicine “Myeloid and Lymphoid Immune Exhaustion Profile During Murine Visceral Leishmaniasis” Diogo Valadares, Richard Davis, Yani Chen, Mary Wilson Leishmaniasis is a chronic progressive parasitic disease involving reticuloendothelial organs (liver, spleen, bone marrow). Leishmaniasis is usually controlled by TH1-type IFN-? producing CD T cells, but microbicidal responses are ineffective during disease progression. It is reported that T cells in humans and dogs with visceral leishmaniasis (VL) express markers of cell exhaustion (sometimes called immune checkpoint inhibitors). We hypothesized that myeloid cells provide counter-receptors for inhibitory receptors on T cells at the local sites of Leishmania infantum infection, inhibiting T cell effector functions. We addressed this hypothesis by examining inhibitory receptors PD1, LAG3, CTLA4 and TIM3 on lymphoid cells, and counter-receptors PDL1, MHCII, CD80 and galectin 9 on myeloid cells, using flow cytometry, throughout 5 weeks of infection. In both livers and spleens, significant increases in PD-L1-expressing myeloid cells and PD-1-expressing lymphoid cells were detected in infected mice compared to control uninfected mice inoculated with PBS. In livers of infected mice, an increased population of neutrophils with DC features (CD11c+MHCII+) expressing PD-L1 and IL-10 was also found. Also present was a tolerogenic DC subset expressing and reduced levels of MHCII, CD80 and CCR7 and higher levels of galectin 9 and PD-L1, than PBS controls. This DC subset also expressed higher levels of intracellular IL-10 compared to controls. In general, CD4 and CD8 T cells showed a similar profile of exhaustion, with CD8 cells demonstrating a deeper commitment to exhausted phenotype. Both liver and spleens of infected mice revealed a higher number of CD4 and CD8 T cells expressing LAG3, CTLA4 and PD-1 compared to uninfected control mice. In the infected group the majority of CD8 antigen experienced (atg exp) cells, in both liver and spleens, are IL-10 producing, demonstrating again their commitment to the regulatory phenotype. In the other hand, also in the infected group, CD4 atg exp cells are mostly producing IFN-g, revealing a transitioning phenotype or incomplete exhausted profile. CD4 atg exp PEPCK+ (Leishmania antigen complexed with MHCII tetramer) and PD1+ subsets in the infected group showed themselves to be driven to exhausted phenotype while the majority of cells, in both liver and spleen tissues, are IL-10 producer cells. In conclusion, pro-exhaustion and tolerogenic DCs, expressing a variety of immune-checkpoint ligands, play a role inducing T cell exhaustion during visceral leishmaniasis.

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IOWA THEATER 1:30-1:45PM Colleen Caldwell, Graduate Student, Biochemistry Mentor: Dr. Maria Spies, Biochemistry “Single molecule observation of the Rad52 recombination mediator mechanism” Colleen Caldwell, Nilisha Pokhrel, Joseph Tibbs, Nina Jocic, Ali Tabei, Marc Wold, Edwin Antony, Maria Spies, Homologous recombination (HR) is essential to repair deleterious forms of double-strand DNA damage. While deficient HR is a cancer risk factor, cancers may also become dependent on excessive HR, making the process an attractive target for cancer therapy. Following a double strand break and resection of the 5’ end in HR, the 3’ overhang is coated by the ssDNA-binding protein, replication protein A (RPA). The high affinity of this interaction blocks formation of the Rad51 nucleoprotein filament – the active species in HR. Displacement of RPA requires a recombination mediator, BRCA2 in humans or Rad52 in Saccharomyces cerevisiae. The mechanism by which a recombination mediator promotes Rad51 nucleoprotein filament formation has remained elusive. RPA contains four DNA binding domains (DBDs), which are proposed to interact dynamically with ssDNA. We have used RPA with individually labeled DBDs and single molecule total internal reflection fluorescence microscopy (smTIRFM) to elucidate domain level dynamics of RPA on ssDNA. We observe four levels of fluorescence that are domain-dependent; suggesting microscopic dissociation events in the bound complex. Both DBD-A and D of RPA showed 4 states of fluorescence, though the behavior of the domains are distinct. Addition of Rad52 resulted in a loss of the highest state of DBD-D fluorescence, though had no effect of DBD-A. The effect of Rad52 on DBD-D dynamics, but not DBD-A, suggests the Rad52 acts to limit the dynamics of DBD-D specifically. The limitation of RPA DBD-D dynamics by Rad52 may allow for nucleation and filament formation of Rad51, thus promoting HR. 1:45-2:00PM Tingting Duan, Graduate Student, Biochemistry Mentor: Dr. Pamela Geyer, Biochemistry “An atypical mode of mitosis is involved in germline stem cell maintenance” Tingting Duan, Pamela Geyer Tissue homeostasis depends on the maintenance of a stable stem cell populations. Following each division, stem cells give rise to one daughter cell that maintains a stem cell identity and a second daughter that enters a differentiation pathway. Such distinct cell fates are often achieved through asymmetric distribution of cellular components during mitosis. Events controlling the asymmetric division are poorly understood. We are studying the Drosophila ovarian germline stem cells (GSCs) where nucleoplasmic regulators of nucleolar function are known to be asymmetrically distributed to different daughters. To understand mechanisms responsible for asymmetric division, we examined the sequential events during GSC mitosis, including nuclear lamina (NL) breakdown, chromosome condensation and spindle formation. Surprisingly, we found that GSCs undergo a semi-closed mitosis where the NL remains intact throughout the entire cell cycle. Nuclear pore complexes are remodeled before metaphase, allowing for the entrance of cytoplasmic components required for spindle formation. Interestingly, centrosomes are embedded in a cup-like structure formed by NL that nucleate the microtubule spindle formation inside and around the nucleus. The NL is essential for GSC mitosis, as loss of the NL component Otefin disrupts the mitotic lamina, causing ruffling and aggregation of Lamin around centrosomes. This distortion of mitotic NL eventually activates a GSC quality control checkpoint that leads to GSC death. Taken together, we propose that GSCs undergo a semi-closed mitosis that is required for asymmetric cell division.

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2:00-2:15PM Jordan Kohlmeyer, Graduate Student, Molecular Medicine Mentor: Dr. Dawn Quelle, Pharmacology “Targeting a Novel RABL6A-RB1 Pathway Suppresses MPNST Pathogenesis” Jordan Kohlmeyer, Courtney Kaemmer, Casey Pulliam, Allison Moreno Samayoa, Chandra Maharjan, Vickie Knepper-Adrian, Nancy Ratner, Munir Tanas, Rebecca Dodd, Dawn Quelle Malignant peripheral nerve sheath tumors (MPNSTs) are deadly sarcomas that lack effective therapies. Greater insight into MPNST pathogenesis is needed to develop new, more targeted treatments. In most MPNSTs, the retinoblastoma (RB1) tumor suppressor is disabled by hyperactivation of cyclin dependent kinases (CDK), commonly through loss of CDK inhibitors such as p27(Kip1). We discovered dramatic upregulation of RABL6A, a newly recognized inhibitor of RB1, in human MPNSTs compared to precursor neurofibromas. Levels of p27 and RABL6A were inversely correlated in the tumors. Silencing RABL6A in MPNST cell lines caused cellular death and G1 phase arrest that coincided with p27 upregulation, CDK downregulation and RB1 activation. The growth suppressive effects of RABL6A loss, and its regulation of RB1, were rescued by p27 depletion. These findings establish a critical role for RABL6A-p27-RB1 signaling in MPNST pathogenesis. Indeed, a CDK4/6 inhibitor (palbociclib) that reactivates RB1 killed MPNST cells in vitro in a RABL6A-dependent manner and strongly suppressed MPNST growth in vivo. Excitingly, treatment with palbociclib promotes tumor cell redifferentiation as measured by restored S100 positivity. This work identifies RABL6A as a powerful driver of MPNST proliferation and survival that acts, at least in part, through p27-RB1 inactivation. Additionally, our findings suggest that RB1 reactivation with FDA-approved CDK4/6 inhibitors may be an effective option for MPNST therapy. 2:15-2:30PM BREAK 2:30-2:45PM Victoria Muller Ewald, Graduate Student, Neuroscience Mentor: Dr. Ryan LaLumiere, Psychological and Brain Sciences “When to seek and when to stop: how activity in the infralimbic cortex changes during the suppression of cocaine-seeking behavior” Victoria Muller Ewald, Ryan LaLumiere Similarities in function exist between the human ventromedial prefrontal cortex and the rodent infralimbic cortex (IL); most importantly, both regions are involved in response inhibition. Prior work in the field of neurobiology of addiction has implicated the IL in the consolidation of extinction learning following cocaine self-administration. However, most studies investigating this region employ manipulations of the IL, and little work has observed IL activity during the extinction of cocaine seeking. To determine how this brain region codes for response inhibition, we used in vivo electrophysiology to record from the IL of behaving rats as they underwent cocaine self-administration and extinction. Male Sprague-Dawley rats underwent surgery for implantation of an intravenous catheter and a fixed electrode array aimed at the IL, followed by cocaine self-administration. Behavioral training occurred inside an operant chamber where a lever press resulted in an intrajugular infusion of cocaine. During extinction training, lever presses were inconsequential. Findings indicate that specific subpopulations of IL neurons change their firing rates and action potential discharge patterns throughout extinction training in a matter that increases infralimbic output to downstream brain regions. Together with work using brain-based manipulations, these findings suggest how IL activity is able to inhibit cocaine-seeking behaviors during extinction training.

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2:45-3:00PM Rebecca Taugher, PhD, Postdoctoral Research Fellow, Psychiatry Mentor: Dr. John Wemmie, Psychiatry “The amygdala differentially regulates defensive behaviors evoked by CO2” Rebecca Taugher, Brian Dlouhy, Collin Kreple, Ali Ghobbeh, Yimo Wang, John Wemmie, CO2 induces panic attacks in patients with panic disorder, though the neural basis of this response remains unclear. In mice, the amygdala appears to promote CO2-evoked responses; however, CO2 still provoked panic attacks and fear in patients with amygdala lesions due to Urbach-Wiethe disease. To clarify the role of the amygdala, we produced lesions in mice paralleling the human lesions. Compared to sham controls, we found that amygdala-lesioned mice froze less to 10% CO2, although they also began to jump frenetically. Jumping was not seen in control mice at 10% CO2, although it was observed at 20% CO2, suggesting jumping is a normal response to more extreme CO2 concentrations. This effect of amygdala lesions was specific to CO2 since amygdala-lesioned mice did not jump in response to a predator odor or to an auditory conditioned stimulus. Jumping evoked by 10% CO2 in amygdala-lesioned mice was eliminated by co-lesioning the dorsal periaqueductal gray a structure previously implicated in panic and escape. Together, these observations suggest a dual role for the amygdala in the CO2 response: promoting CO2-induced freezing, and opposing CO2-induced jumping, and this jumping behavior seems to parallel the exaggerated CO2 responses in Urbach-Wiethe patients. 3:00-3:15PM Daniel Infield, PhD, Postdoctoral Research Fellow, Molecular Physiology and Biophysics Mentor: Dr. Christopher Ahern, Molecular Physiology and Biophysics “Atomic Mutagenesis of Voltage Sensing in Ion Channels” Daniel Infield Ion channels serve as signal decoders in cells. Despite decades of intensive structural and functional work, many important regulatory mechanisms for voltage-gated channels remain poorly understood. We have used a chemical biology approach to circumvent traditional limitations in the design and manipulation of ion channels to attain enhanced insights into the structural role of a specialized alpha helix that confers voltage sensitivity on these highly clinically relevant proteins.