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Manipulation of Sumoylation Processes by Epstein-Barr virus LMP1

Presenter: Gretchen Bentz
Other, Mercer University, Tallahassee, FL, 32306

In vivo Knock-down of the Herpes Simplex Virus Type 1 Latency-Associated Transcript Reduces Reactivation from Latency and Therapeutic Implications

Presenter: David Bloom
Other, University of Florida, Gainesville, FL, 32306

Trials and Errors in My Exploration of Viral Assembly

Presenter: Donald Caspar Ph.D.
Biological Sciences, Florida State University, Tallahassee, FL, 32306

Quantitative and qualitative changes of FoxP3+ Regulatory T cells in human acute viral infection.

Presenter: Yoon Seok Choi M.D., Ph.D.
Other, Chungnam National University College of Medicine, Daejon, FL, 32312
FoxP3+ regulatory T (Treg) cells play a major role in maintaining the immune homeostasis by preventing the activation of self-reactive T cells as well as in controlling a series of immune responses in viral infections. Treg cells, during chronic viral infections, have been shown to hamper virus-specific T cell immunity and thereby to allow persistence of viral infection. In contrast, roles of Treg cells in human acute viral infection remain largely unknown and functional changes of Treg cells in the context of immunopathology have not been re-ported so far. I investigated several functional changes of Treg cells and their potential implications in immunopathogenesis in acute hepatitis A (AHA).

The suppressive activity of total Treg cell pool in the peripheral blood was attenuated during AHA and inversely correlated with the serum alanine aminotransferase (ALT) level. Moreover, the frequency of circulating Treg cells was reduced during AHA without compartmentalization into the liver. Notably, the frequency of Treg cells in the peripheral blood directly correlated with the suppressive activity of the Treg population and inversely correlated with the serum ALT level.

Along with such quantitative contraction of Treg cell population, a remarkable proportion of CD4+CD25+Foxp3+ Treg cells from AHA patients produced TNF-α upon anti-CD3/CD28 stimulation, particularly during the acute phase. The Treg-specific demethylated region of TNF-α-producing Treg cells was largely demethylated, indicating that these cells are bona fide Treg cells. TNF-α-producing Treg cells from AHA patients exhibited Th17-like features including upregulation of RORγt, and their TNF-α production depended on RORγt. Furthermore, these cells exhibited a reduced suppressive function, and their frequency significantly correlated with serum ALT, indicating that the pathologic conversion of Treg cells to produce TNF-α contributes to liver injury in AHA.

Taken together, functional impairment of Treg cell population results from both quantitative defect of Treg cell pool and qualitative defect secondary to inflammatory conversion and is responsible for immune-mediated tissue injury in human acute viral infection model.

Dengue virus immune evasion: feeling the STING

Presenter: Ana Fernandez-Sesma
Other, Icahn School of Medicine at Mount Sinai, Tallahassee, FL, 32306

Controlling the message: viral manipulation of host gene expression during infection

Presenter: Britt Glaunsinger Ph.D.
Other, University of California Berkeley, Berkeley, CA, 32306

The Interplay between Noroviruses and Commensal Bacteria

Presenter: Stephanie Karst
Other, University of Florida, Gainesville, FL, 32306

Concentration of Soluble DPP4 in patients infected with MERS-CoV: is it correlated with the severity of MERS-CoV illness?

Presenter: Yeon-Sook Kim
Other, Chungnam National University College of Medicine, Daejon, FL, 32306

Background: Dipeptidyl peptidase 4 (DPP4) is known as the key functional receptor for entry of MERS-CoV into host cells. In humans, it is abundantly expressed on the epithelial and endothelial cells of most organs, and exists as a soluble form in the circulation. We intended to estimate the amount of soluble DPP4 (sDPP4) from the patients infected with MERS-CoV and correlate it with the severity of MERS-CoV illnesses.

Methods: We estimated the concentration of sDPP4 in sera and sputa of 14 MERS-CoV-infected patients using ELISA. The patients infected with MERS-CoV were classified into four groups (G 1: cases with only febrile illness, G 2: cases with mild pneumonia, G 3: cases with severe pneumonia requiring oxygen therapy, G4: fatal cases). We also checked the inhibitory effect of sDPP4 on MERS-CoV spike pseudotyped lentivirus and wild type virus.

Results: The concentration of sDPP4 in plasma was significantly low in patients infected with MERS-CoV (200 – 600 ng/mL), compared to that of normal controls, but it showed no significant difference depending on the groups. sDPP4 in sputa seemed to be lower than 300 ng/mL and significantly low (<100 ng/mL) in patients with only febrile illness and fatal cases. Viral entry was inhibited with 102 to 10 3 ?g/mL of sDPP4 by 10%.

Conclusions: This is the first report of sDPP4 in plasma and sputa of patients infected with MERS-CoV. The concentration of sDPP4 measured from the patients infected with MERS-CoV was low and tend to be lower in sputa than in plasma. We could not find any significant correlation between the concentration of sDPP4 and the severity of MERS-CoV illness. The exact function of sDPP4 in the pathogenesis of MERS-CoV infection needs to be elucidated with future study.

Mycobacterial components regulating macrophage apoptosis via targeting mitochondria

Presenter: Hwa-Jung Kim
Other, Chungnam National University College of Medicine, Daejon, FL, 32306
Macrophages are the primary target and an important reservoir of mycobacteria in the lungs, as indicated by the fact that virulent mycobacteria can survive and replicate within macrophages. Macrophage cell death is an important facet of host–mycobacteria interactions. Therefore, the properties of mycobacteria that are involved in modulating apoptosis have been extensively investigated. Most reports have indicated that the induction of apoptosis by Mycobacterium tuberculosis and M. avium is inversely proportional to bacterial virulence. However, apoptosis appears to act as a virulence mechanism for M. leprae and M. ulcerans. Therefore, induction of apoptosis can apparently serve different functions according to the mycobacterial strain and its infection course.

In general, apoptotic cell death is regarded an innate intracellular response to limit the multiplication of intracellular pathogens. Several apoptosis-inducing factors of M. tuberculosis have been identified: 19-kDa protein and lipoarabinomannan via TLR2, PE_PGRS33 with TNF-α inducing ability, 38-kDa protein with up-regulation of cell death receptor. In addition, we reported that M. tuberculosis HBHA may act as a strong pathogenic factor to cause apoptosis of professional phagocytes infected with M. tuberculosis. In fact, high intracellular burden of virulent M. tuberculosis induces apoptosis with rapid progression to necrosis as a mode of mycobacterial escape.

M. avium complex (MAC) and their sonic extracts induce a macrophage apoptosis. However, any components of MAC that are involved in inhibiting or triggering apoptosis are not identified. In recent, we found that that MAV2054 protein induced significant apoptosis in macrophages through production of reactive oxygen species (ROS), loss of mitochondrial membrane potential (MMP), and the release of cytochrome c, and MAV2054 was efficiently target to mitochondria of macrophages.

Mycobacterium abscessus (MAB) is a common respiratory pathogen. The rough (R) morphotype of MAB, lacking cell-surface glycopeptidolipids (GPLs), is associated with more severe and persistent infection than the smooth (S) type; however, the mechanisms underlying R-type virulence remain obscure. We found that the R type induced significantly more macrophage apoptosis than the S type, in which apoptosis is inhibited by the GPLs. The GPLs are targeted to mitochondria and interact with cyclophilin D, leading to preservation of the mitochondrial transmembrane potential and suppressed growth of R-type MAB in macrophages and mouse lung. Finally, GPLs enhanced the survival rate of mice infected with the R type. Our results provide insights into the cellular mechanism of R-type virulence and a potential strategy for the control of MAB infection through modulation of cell death.

Role of NADPH oxidases in T cell receptor Signaling

Presenter: Jaeyul Kwon
Other, Chungnam National University College of Medicine, Daejon, FL, 32306
Reactive oxygen species (ROS) are generated in pathological processes and also widely used in cell signaling and regulation of various physiological processes. Many of the responses to growth factors, hormones and inflammatory cytokines involve redox signaling, with hydrogen peroxide generally proposed as a second messenger. The Nox and dual oxidase (Duox) enzymes in the NADPH oxidases (Nox) family are membrane flavocytochromes that catalyze NADPH-dependent reduction of molecular oxygen to generate superoxide and/or hydrogen peroxide (H2O2). Although originally understood as an anti-bacterial mechanism employed by phagocytes, our studies revealed that ROS intentionally generated by the members of NADPH oxidase (Nox) family play specific signaling roles in T cell receptor (TCR)-stimulated T cells. We found that at least three Nox family members are identified to play important roles in TCR signaling. Early H2O2 generation induced by TCR stimulation comes from activated Duox1 downstream of inositol 1,4,5 triphosphate receptor 1; the later response comes from activated Nox2 downstream of the Fas receptor. Also another Nox isoform Nox4 was induced by T cell activation. In terms of cytokine production, Nox2-derived late ROS inhibits Th1 but augments Th2 cytokine production, whereas Duox1 augments both Th1 and Th2 cytokine production. Another Nox isoform Nox4 strongly inhibits Th17 differentiation and Th2 differentiation. Our data suggest there are separate signaling processes based on ROS generated from different Nox isoforms, producing different biological outcomes in stimulated T cells

Epigenetic Control of Gammaherpesvirus Latency and Targeted Therapy

Presenter: Paul Lieberman
Other, The Wistar Institute, Tallahassee, FL, 32306

Hijacking the host exosomal trafficking pathways by Epstein-Barr virus

Presenter: David Meckes Ph.D.
Biomedical Sciences, Florida State University, Tallahassee, FL, 32306
The Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) is released from latently infected tumor cells in small membrane-enclosed vesicles called exosomes. These LMP1-modified extracellular vesicles can be taken up by cells, resulting in the activation of cellular signal transduction pathways through paracrine or autocrine mechanisms. Accumulating evidence suggests that LMP1 is a major driver of exosome content and functions. For example, LMP1-modified exosomes have been shown to influence cell growth, migration, differentiation, and immune cell function. Even though the importance of LMP1-modified extracellular vesicles on the infected microenvironment is well recognized, very little is known about how this viral protein enters or manipulates the host exosome pathway. Our results indicate N-terminal domains of LMP1 direct exosomal packaging and vesicle production through CD63- and ceramide-dependent mechanisms. Additional LMP1 interactome studies identified a large number of proteins important in exosome formation and protein trafficking, including CD63, syntenin-1, ALIX, TSG101, Hrs, CHMPs, and sorting nexins. Taken together, these data suggest that LMP1 manipulates the host exosome pathway through modification of CD63/ceramide-containing microdomains and endocytic routes.

Zika virus activation of the centrosome

Presenter: Timothy Megraw Ph.D.
Biomedical Sciences, Florida State University, Tallahassee, FL, 32306

Structural insights into MAP kinase mediated cellular signaling

Presenter: Attila Remenyi
Other, Institute of Enzymology, Research Center for Natural Sciences, Hungarian Academy of Sciences, Budapest, FL, 32306
Cells receive myriads of signals from their environment and my group has been exploring how members of the so-called mitogen activated protein kinase (MAPK) family mechanistically control cellular responses to extracellular signals. We know that MAPKs are ubiquitous signaling enzymes involved in cellular life and death decisions. They exert their effect on other proteins by phosphorylating them on specific sites. These phosphorylation events then will cause their substrate proteins to change their function leading to specific changes in the cells'' behavior. In the past years, we discovered how MAPKs can form specific signaling complexes with their partner proteins. We are currently exploring how marker proteins of some pathological disorders as well as viral proteins may influence MAPK mediated signaling by interfering with the formation of key protein-protein complexes. Furthermore, structural characterization of MAPK involving signaling complexes also suggests new strategies on how to combat MAPK involving signaling diseases such cancer or inflammation.

New insights into Epstein-Barr virus LMP1 oncogenesis

Presenter: Kathy Shair Ph.D.
Other, University of Pittsburgh, Tallahassee, FL, 32306
Epstein Barr virus (EBV) is an important ubiquitous human pathogen associated with several lymphoid and epithelial cell malignancies. Although EBV immortalizes B-lymphocytes, the historical association of latent EBV infection in nasopharyngeal carcinoma (NPC) - an epithelial cell cancer that arises in the immunocompetent, is more complex as infection alone is insufficient to immortalize epithelial cells. This is consistent with the hypothesis that multiple risk factors are involved including genetic and viral pre-disposition. The EBV-encoded latent membrane protein 1 (LMP1) is a pleiotropic signaling molecule associated with oncogenic growth properties in NPC. Data from transgenic mice and studies with genotoxins support that LMP1 co-operates with another latent protein (LMP2A) to affect DNA damage response signaling, genomic instability and progression to carcinoma. Additionally, the sequence variants of LMP1 and pleiotropic signaling properties localized to conserved domains are a unique mapping tool for probing the poorly defined cellular mechanisms of cancer cell repopulation in metastasis. These new insights into LMP1-induced biologies utilize molecular virology to illuminate cancer mechanisms.

The Structures of Adeno-Associated Virus and It''s Binding Partners

Presenter: Scott Stagg Ph.D.
Chemistry and Biochemistry, Florida State University, Tallahassee, FL, 32306

Stem Cell Models and Zika Virus Infection

Presenter: Hengli Tang Ph.D.
Biological Sciences, Florida State University, Tallahassee, FL, 32306
Stem cell-derived progeny cells can be used to study virus infections, in many cases providing alternative cell culture models for viruses that had been difficult to culture in vitro. My lab has previously used hepatocyte-like cells produced by hepatic differentiation of human pluripotent stem cells (hPSCs) to study the infection of hepatitis C virus and dengue virus (DENV). More recently, and as a first step to demonstrate Zika virus (ZIKV) causality for microcephaly, we demonstrate dthat ZIKV targets human cortical neural progenitor cells (hNPCs) derived from induced pluripotent stem cells (iPSCs). Infection by ZIKV, but not by DENV, induced a profound perturbation of cell cycle progression and caspase activation in these important cells for cortex neuron differentiation. In addition to stem cell-derived NPCs, we are also using hPSC-derived macrophages to model interaction between human macrophage and ZIKV or DENV. Our results indicate the distinct cellular responses to these two closely-related flaviviruses by human macrophages may underlie the different pathogenic outcomes of the Zika and dengue diseases.

Resolving the structure and function of noncoding RNAs from a lymphomagenic gammaherpesvirus

Presenter: Scott Tibbetts
Other, University of Florida, Gainesville, FL, 32306

Transcriptional reprogramming of primary effusion lymphoma cells during KSHV reactivation

Presenter: Zsolt Toth
Other, University of Florida College of Dentistry, Gainesville, FL, 32610
Kaposi’s sarcoma-associated herpesvirus (KSHV) establishes lifelong infection in humans, which can lead to the development of cancers such as Kaposi’s sarcoma and primary effusion lymphoma (PEL) in immunocompromised people. While the majority of PEL cells are latently infected by KSHV, the virus can be reactivated, which contributes to not only virus production but also the expression of a number of lytic viral proteins that can facilitate the proliferation of PEL. The reactivation of the virus starts with the expression of the replication and transcription activator (RTA) encoded by KSHV. RTA is required and sufficient for inducing lytic reactivation of KSHV from latently infected B cell lymphoma cells. Although several viral and host genes regulated by RTA have been identified, a systematic and unbiased identification of RTA target genes, which can play a role in the lytic reactivation and replication of KSHV is still missing. To uncover the viral and host target genes, which are directly regulated by RTA, we performed RTA ChIP-seq and a time course RNA-seq analysis using reactivated PEL cells. The results of our genomic analysis and the identified RTA-regulated host pathways will be discussed.

Structural Basis of AIM2 inflammasome Activation and Regulation

Presenter: Qian Yin Ph.D.
Biological Sciences, Florida State University, Tallahassee, FL, 32306

In situ structures of viral genome and genome transcription

Presenter: Hong Zhou
Other, UCLA, Los Angeles, CA, 32306

Innate immune sensing of cytosolic DNA and viral evasion strategies

Presenter: Fanxiu Zhu
Biological Sciences, Florida State University, Tallahassee, FL, 32306