Principal Investigator

Joseph C. Glorioso, III, PhD
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Location

433 Bridgeside Point II
450 Technology Dr.
Pittsburgh, PA 15219

Research Description

Dr. Glorioso has established a 35-year history of research related to the basic biology and genetics of herpes simplex virus. His contributions to the field include defining antiviral immune responses to infection, the genetics of viral pathogenesis and latency, and mechanisms of viral infection. Furthermore, he has been a pioneer in the design and application of HSV gene vectors for the treatment of nervous system diseases such as peripheral neuropathies, chronic pain and brain tumors. He continues to be a worldwide leader in the HSV gene vector field through the creation of innovative gene vectors and the development of manufacturing methods for Phase I and Phase II human clinical trials for pain gene therapy. His enkephalin vector showed considerable promise in a Phase I human trial to treat cancer pain and phase II testing is underway. These trials were sponsored by Diamyd Medical AB in Stockholm. Dr. Glorioso is a Diamyd stockholder.

Herpes simplex virus infection of a tumor cell monolayer. Uninfected cells are outlined in blue, infected cells are red turning green following virus replication Dr. Glorioso’s most recent research has focused on (i) the design and application of HSV gene vectors for exploring the molecular events that occur in sensory afferents that are involved in the transition from acute to chronic pain, (ii) the development of retargeted oncolytic HSV vectors for specific infection and replication in human glioblastomas and applications to treatment of xenograft models of human brain human brain tumors, (iii) the creation of novel HSV vectors that cross the blood brain barrier by transcytosis followed targeted infection and gene expression in spiny neurons of the striatum; these vectors are being applied to treatment of animal models of Huntington’s disease and (iv) the use of HSV gene vectors for the creation of induced pluripotent stem (iPS) cells and the identification of transcriptional regulatory and signaling processes that participate in cellular reprogramming. 

Over the years, his laboratory has studied the structure/function domains of the HSV envelope glycoproteins that play a role in cognate receptor recognition and activation of the fusion/entry mechanism. HSV relies on 4 essential glycoproteins (gD, gB, gH, gL) to gain entry into cells. gD binding to its natural receptors, HVEM or nectin-1, results in signaling events required for activation of the gB-gH/gL membrane fusion apparatus. Virus envelope fusion with cell membranes releases the virus capsid into the cytoplasm whereupon it is trafficked to the nucleus, viral DNA is released into the nucleoplasm and the replication cycle is initiated. These studies have enabled the genetic manipulation of these attachment/entry function to create viruses with novel tropisms for neurons and tumor cells. For example, we have engineered HSV vectors for infection of stem cell markers, tumor markers and neurotrophic factor receptors. The safety of these vectors have been further enhanced by microRNA control of essential gene expression in normal brain.    

Glorioso, Vero CellsRetargeted, miR controlled vectors are also being exploited for selected gene expression in neurons of the peripheral and central nervous system. Of particular interest are a new generation of vectors that express neuron silencing by drug activated ion channels. The application of neuron silencing methods hold great promise for mapping neurons involved in cognitive processes and chronic pain signaling.

Members

Fang Han, Tsinghua Univ. Visiting Scholar
Mindgi Zhang, Senior Research Specialist

Ceren Tuzman, Postdoctoral Associate Sara Artusi, Postdoctoral Associate



Principal Investigator

Jennifer Bomberger, PhD
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Location

445 Bridgeside Point II
450 Technology Dr.
Pittsburgh, PA 15219

Research Description

Viral-bacterial interactions in the lung.  A principal focus in the Bomberger lab is exploring the mechanistic basis by which viral infections promote the development of chronic bacterial infections in the lung, particularly in the setting of chronic lung diseases, like Cystic Fibrosis (CF), bronchiectasis and chronic obstructive pulmonary disease (COPD). Dr. Bomberger’s recent publications provide compelling evidence that respiratory viral infections, and the antiviral immune response to them, drive P. aeruginosa to transition from acute to chronic infection. Current studies in the lab are focused on elucidating molecular mechanisms that govern the innate immune induction of biofilm in the lung. Working to take her work from the bench to the bedside, Dr. Bomberger collaborates with Dr. Stella Lee in Otolaryngology at UPMC to examine viral-bacterial interactions in the sinuses of patients with chronic lung disease. The goal of these studies is to examine the adaptation and transmission of P. aeruginosa between the upper and lower respiratory tracts during viral exacerbations. These studies include novel genomic approaches, in collaboration with Dr. Vaughn Cooper in MMG, to capture the P. aeruginosa diversity in both anatomical sites and determine if this is altered with viral infection in a way that promotes disease progression. The long-term goal of this collaborative project is to translate observations at the bench to new therapeutic strategies to prevent chronic bacterial infections in patients with chronic rhinosinusitis, focusing on iron chelation therapy. 

P. aeruginosa targets pro-resolving lipid mediators in the lung. Approximately ninety-five percent of deaths in CF are attributed to P. aeruginosa infection and the resulting chronic inflammatory response in the airway. Lipoxins are bioactive eicosanoids that mediate resolution of inflammation and are dramatically reduced in the CF airway. Dr. Bomberger’s laboratory has demonstrated that a secreted P. aeruginosa protein Cif reduces lipoxins in the airway and thus, prevents the resolution of inflammation in the CF airways. These studies are the first to show bacterial inhibition of pro-resolving mediators, as well as describe a new bacterial virulence mechanism. In addition to Cif’s regulation of lipoxin in the airways, Dr. Bomberger’s laboratory is also examining antimicrobial functions for lipoxin in the airways, with the long-term goal of adapting stable lipoxin orthologs for therapeutic applications in chronic lung disease. 

Identification of P. aeruginosa biofilm inhibitors.  Dr. Bomberger’s laboratory utilizes in vitro live-cell imaging systems to culture P. aeruginosa biofilms in association with host airway epithelial cells. Due to a lack of small animal models that suitably model P. aeruginosa biofilm growth in vivo, this model allows the study of bacterial biofilm development in the lung. In collaboration with Drs. Ronald Montelaro (MMG, Center for Vaccine Research) and Mark Gladwin (PACCM) at the University of Pittsburgh, Dr. Bomberger’s laboratory is developing new antimicrobial therapies that disrupt P. aeruginosa biofilms growing in the lung. Dr. Bomberger’s laboratory recently reported that an engineered antimicrobial peptide (WLBU2) disrupts incredibly antibiotic resistant bacterial biofilms grown during a viral coinfection, while concurrently reducing viral burden. Dr. Bomberger’s group has also studied the antibiofilm properties of sodium nitrite by targeting bacterial respiration pathways. Truly translating these sodium nitrite studies at the bench to the bedside, the Cystic Fibrosis Center at the University of Pittsburgh is funded to conduct a clinical trial to examine the efficacy of sodium nitrite as a novel antimicrobial therapy for Cystic Fibrosis patients.

Members

Anna Zemke - Asistant Professor, Department of Pulmonary and Critical Care Medicine

Jordan Gaston - Research Technician

Brian Kocak - Research Assistant

Catherine Armbruster - Postdoctoral Associate



Principal Investigator

Fred L. Homa, PhD
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Location

539 Bridgeside Point II
450 Technology Dr.
Pittsburgh, PA 15219

Research Description

Research in our lab is focused on understanding the mechanism of herpesvirus capsid assembly and DNA packaging.  The herpesviruses comprise a large family of double stranded DNA viruses. Several of these viruses are important human pathogens and all remain with the host for life by residing in a latent or quiescent state where they avoid immune clearance. Primary or recurrent infections can be life threatening in immunocompromised patients, such as AIDS or transplant patients, where infection with human cytomegalovirus (HCMV) can result in retinitis, pneumonia, and gastrointestinal disease. Current treatments against herpes simplex viruses (HSV-1 and HSV-2) and the other human herpesviruses rely primarily on blocking viral DNA replication. Assembly of herpesvirus capsids involves highly specific interactions among at least five different proteins and seven additional proteins are involved in DNA packaging and cleavage. Most of the proteins involved in capsid assembly and DNA packaging are conserved suggesting that these mechanisms will also be similar for all herpesviruses.

Homa Lab ImageResearch in our lab is focused on understanding the mechanism of herpesvirus capsid assembly and DNA packaging. The structure of the HSV‑1 capsid was determined from three-dimensional image reconstruc­tions com­puted from cryo­-electron micrographs of capsids. The capsid shell is composed predominantly of four proteins, a major capsid protein, VP5, and three less abundant proteins, VP19C, VP23 and VP26. Herpesvirus DNA is incorporated into preassembled capsids through a ring-shaped portal present at a unique vertex. This process requires the action of six cleavage/packaging pro­teins that interact with the ­capsid either during capsid assembly or during DNA packaging. The terminase proteins (UL15, UL28, UL33) act as part of an ATP-dependent pump that drives DNA into the procapsid and cut the concatemeric DNA at specific sites yielding a capsid containing the intact genome. The final step in the process is “capsid completion” that results in the formation of a stable DNA-containing capsid. Of the seven HSV-1 pro­teins required for the cleavage/packaging reaction, only UL25 is required for maintaining the stable DNA-containing capsid; without UL25 the packaged DNA is lost resulting in “empty” A-capsids. The cleavage/packaging and capsid completion reactions can be viewed as separate steps in the overall process of generating a stable DNA-containing capsid.  The main goals of this project are to determine the function(s) of the individual cleavage/packaging proteins in this process in order to achieve a detailed understanding of the HSV DNA cleavage and packaging mechanism.

Ongoing studies are focused at defining the role of the UL25 protein in DNA packaging with regards to its functions in retention of viral DNA by binding to capsid vertices through its interaction with the UL17 protein.  Genetic and biochemical approaches are being be used to determine the role of UL28 in the assembly of a functional terminase complex and its interactions with UL15 and UL33.  These studies utilize genetic, biochemical and structural (cryoEM) approaches to understand how the protein complexes assemble and carry out the cleavage/packaging reaction.

In collaboration with Dr. James Conway’s lab (Department of Structural Biology) molecular genetics and cryo-electron microscopy (cryoEM) are being used to obtain high resolution models of the HSV capsid and the essential minor proteins that interact with the capsid during and following DNA packaging. The locations of most of these essential minor proteins are not known nor are details of their interactions with each other and the capsid. Capsids incorporating specifically labeled subunits will be visualized by cryoEM to identify the locations of subunits. The knowledge obtained from these studies enables not only a significantly better understanding of herpesvirus capsid structure, but also provides the means to reveal aspects of how the viral DNA packaging machinery interacts with the capsid during and after DNA packaging. In addition, the essential minor proteins offer novel and highly specific structural targets for the development of antivirals. It is anticipated that the studies in this proposal will not only enhance our understanding of the mechanisms of genome maturation and encapsidation and lead to the development of novel strategies for antiviral therapy.

LAB MEMBERS:

Mackenzie Kershner, Research Assistant

Seminars

MMG has a departmental seminar series that runs on Wednesday afternoons. All seminars begin at 12 noon in room 503 Bridgeside Point II unless otherwise noted. Please see below for upcoming seminar announcements.



Principal Investigator

Martin C. Schmidt, PhD
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Location

543 Bridgeside Point II
450 Technology Dr.
Pittsburgh, PA 15219

Research Description

My lab studies the Snf1 kinase of yeast. The mammalian homologue of Snf1 is the AMP-activated protein kinase, an important therapeutic target for type II diabetes. Biochemical and genetic experiments have shown that Snf1 kinase is regulated by phosphorylation of the conserved threonine residue in the kinase activation loop. We have developed a phosphopeptide antibody that specifically recognizes the phosphorylated (active) form of Snf1 kinase. We have used the antibody to demonstrate that Snf1 is activated by three distinct upstream kinases called Sak1, Tos3 and Elm1. We now know that the Snf1-activating kinases are not themselves regulated by glucose. Instead, it is the DEphosphorylation of the Snf1 activation loop that responds to changes in glucose abundance. The yeast PP1 phosphatase is responsible for the dephosphorylation of Snf1 in response to changes in carbon source. We have shown that the PP1 phosphatase is active in low glucose toward most substrates. However, the Snf1 kinase becomes resistant to dephosphorylation. These data indicate that the active Snf1 kinase can adopt a phosphatase resistant structure. The phosphatase resistant structure is stabilized in vitro by binding low energy adenylate ligands such as AMP and ADP. In this way, the Snf1 kinase is a direct sensor of the cell’s energy status with low energy adenylate ligands stabilizing the active form of Snf1 which then promotes ATP synthesis and conservation. The long term goal of the lab is to identify all the components of the glucose signaling pathway in yeast and to understand how they interact in order to regulate gene expression and cellular metabolism. These studies will provide a better understanding of glucose-mediated regulation of cellular metabolism and have important implications for designing novel treatments for patients with diabetes.

Lab Members:

Dakshanyini Guddenahalli Chandrashekarappa, Research Specialist



Principal Investigator

Thomas E. Smithgall, PhD
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Location

533 Bridgeside Point II
450 Technology Dr.
Pittsburgh, PA 15219

Research Description

Smithgall, HIV Research in our laboratory is focused on non-receptor protein-tyrosine kinase structure, function, and inhibitor discovery. Specifically, we are interested in the Src, Abl and Fes kinase families, which were originally discovered in the context of avian transforming retrovirus many years ago. Since that time, normal human orthologs of these kinases have been identified and implicated in a wide variety of human diseases, ranging from cancer to HIV/AIDS. One important goal of our research program is to better understand the mechanisms of kinase regulation unique to each family. Structurally, these kinases are composed of a series of modular domains which assemble in unique ways to control kinase activity. For example, members of the Src family are composed of Src homology 2 (SH2) and SH3 domains, compact protein-protein interaction modules that work together to downregulate kinase activity. Our group has discovered that HIV encodes a protein that directly engages the SH3 domains of a subset of Src-family kinases, displacing SH3 from its regulatory position and causing kinase activation. Using high-throughput chemical library screening, we have identified selective inhibitors of this viral-host cell protein interaction that also interfere with HIV replication. Working with structural biologists, we are currently exploring the unique active conformations of Src kinases that result from interactions with HIV proteins. These studies will reveal high-resolution structural details essential to improving inhibitor potency and efficacy. Another example is Abl, best known in the context of Bcr-Abl, the chimeric oncogenic tyrosine kinase responsible for chronic myelogenous leukemia. Selective inhibitors of this kinase have been remarkably effective in the treatment of this form of cancer. Bcr-Abl inhibitors selectivity recognize and trap a unique, inactive kinase domain conformation. Like Src-family kinases, Bcr-Abl also has SH3 and SH2 modules important for kinase regulation. We are very interested in the discovery of small molecules that enhance this natural regulatory mechanism. The Fes-related kinases also share homology with Abl and Src in that they have SH2 and kinase domains. However, these kinases also possess a unique N-terminal region with coiled-coil homology domains. Coiled-coils are helical structures that hold proteins together, and are responsible the oligomeric nature of Fes in vivo. We have observed that the coiled-coils are also critical to downregulation of kinase activity. Unlike Src and Abl, no pharmacological inhibitors of c-Fes have been reported. To fill this void, we recently identified a variety of compounds with potent activity against c-Fes. Using these inhibitors, we demonstrated for the first time that Fes has an essential role in the differentiation of macrophages to osteoclasts, making it a possible drug target in osteoporosis, multiple myeloma, and tumor angiogenesis. More generally, our research program seeks to exploit the novel regulatory features of each of these kinase families to develop new classes of selective kinase inhibitors. Such compounds represent valuable probes to explore kinase function in normal cellular physiology and in disease.

Members:

Heather Rust, Postdoctoral Associate                            

Haibin Shi, Research Assistant Professor

Ryan Staudt, Graduate Student

Li Chen, Research Specialist

Julie Reese, Research Specialist

Manish Aryal, Graduate Student

Shoucheng Du, Research Assistant Professor 

Molecular Basis of Cancer

Several MMG research groups are investigating the molecular etiology of cancer induced by tumor viruses as well as the alterations in signaling pathways associated with oncogenic transformation. Specific projects are focused on the role of microRNAs in HPV-associated cervical cancer, the KSHV and MCV human tumor viruses, and protein-tyrosine kinases as molecular targets for cancer therapy.

Bernstein Lab 

Repair of DNA damage is crucial to prevent accumulation of mutations that can cause human disease, such as cancer. Our lab studies how double-strand breaks in the DNA, one of the most lethal types of DNA lesions, are repaired. Learn more>

Khan Lab 

We are involved in three main areas of research. The first one deals with the role of microRNAs in human papillomavirus-associated cervical and oral cancers as well as role of miRNAs in aging. The second area deals with the cellular functions and mechanism of action of the PcrA helicase which is specifically found in Gram-positive bacteria. The third area of our interest deals with a molecular analysis of the role of the RepX protein in the replication and segregation of the anthrax toxin-encoding pXO1 plasmid in Bacillus anthracis. Learn more>

Moore Lab

We study 1) Kaposi’s sarcoma-associated herpesvirus (KSHV), the viral cause of Kaposi’s sarcoma, 2)  Merkel  cell  polyomavirus  (MCV), the viral cause of Merkel cell carcinoma and 3) methods to search for undiscovered human tumor viruses.  Learn more>

Shair Lab

The Shair lab studies the molecular mechanisms of cancer induced by this latent virus with the purpose of defining how these mechanisms contribute to the oncogenic and metastatic properties of EBV-associated diseases. Learn more>

Smithgall Lab

This laboratory research is focused on non-receptor protein-tyrosine kinase structure, function, and inhibitor discovery. Interest lies specifically in the Src, Abl and Fes kinase families, which were originally discovered in the context of avian transforming retrovirus many years ago. Learn more>

Thomas Lab

Our research program focuses on signaling pathways that integrate membrane traffic with the regulation of homeostasis and the onset of disease. These studies were grounded by our identification of the proprotein convertase furin, which is the first member of a family of secretory pathway-localized endoproteases that catalyze the activation of bioactive proteins and peptide hormones. Learn more>

Xiao-Qu Lab

Our primary research interests include the study of signaling transduction pathways in immunity and tumorigenesis, particularly NF-kB, as well as the molecular mechanisms underlying the type-1 human T cell leukemia virus (HTLV-I) mediated T cell transformation for disease prevention and therapeutic purposes. Learn more>

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Early Development, Epigenetics, and Stem Cell Biology

MMG investigators in this group are interested in the molecular mechanisms controlling embryonic stem (ES) cell as well as tissue-specific stem cell growth and differentiation, the early stages of embryogenesis, and the application of these findings to the regenerative medicine. Specific projects include genetic and epigenetic mechanisms that regulate ES cell differentiation, genomic imprinting in ES cell biology, the impact of aging on stem cells and tissue regeneration, and kinase signaling pathways in Drosophila development.

Glorioso Lab

Dr. Glorioso’s most recent research has focused on (i) the design and application of HSV gene vectors for exploring the molecular events that occur in sensory afferents that are involved in the transition from acute to chronic pain. Learn more>

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