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.

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, Senior Research Specialist

Samantha Soncini, 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

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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DNA Replication and Repair

This research area focuses on the molecular mechanisms of DNA replication in microorganisms as well as the repair responses of mammalian cells following exposure to DNA damaging agents. Specific projects include mechanisms of rolling-circle replication of antibiotic resistance plasmids in bacteria, the role of protein kinases in DNA damage signal transduction, and relationship of DNA repair to cancer and aging.

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>

Cooper Lab

The primary goal of our laboratory is to understand how bacterial populations evolve and adapt to colonize hosts and cause disease. We are particularly focused on how bacterial populations form complex communities within biofilms and how cells perceive cues to attach or disperse. 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>

Levine Lab

To preserve the integrity of the genome, cells have developed various sophisticated mechanisms for repairing damaged DNA. The major DNA repair process that removes helixdistorting lesions from DNA, including UV-induced cyclobutane pyrimidine dimers (CPD) and 6,4 PhotoProducts (6,4-PP) is the nucleotide excision repair (NER) pathway. However, in eukaryotic cells, NER operates on chromatin-embedded DNA substrates and DNA folding with histone proteins into chromatin poses structural constraints likely to challenge detection and repair of DNA lesions. Only recently there has been an emphasis on the relationship of chromatin to NER. Learn more>

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Microbial Pathogenesis

Basic insights regarding the mechanisms of the host-pathogen relationship are essential to improvements in infectious disease prevention, vaccine development, and antimicrobial drug design. MMG faculty members are investigating the molecular biology and host immune responses to a diverse group of human pathogens responsible for tuberculosis, AIDS, yellow fever, dengue fever, tularemia, and Clostridia-associated food poisoning.

Apetrei Lab 

My laboratory is interested in the study of the HIV/ SIV diversity and pathogenesis. The AIDS pandemic is produced by two different viruses, HIV-1 and HIV-2. These two viruses resulted from cross-species transmissions of SIVs, the viruses that naturally infect nonhuman primate species (NHPs) in Africa.  Learn more>

Bina Lab 

Our research is centered on defining the molecular mechanisms used by bacteria to resist antibiotics and cause disease in humans. Our work currently focuses on two important gram negative human pathogens: Vibrio cholerae and Francisella tularensisLearn more>

Bomberger Lab

My research program is focused on understanding host-pathogen interactions, and more specifically, how each influences the other during an infection.  Emerging evidence reveals that pathogens have the ability to modulate the host response to infection, while at the same time, respond to host defense by altering their virulence and antibiotic resistance. Learn more>

Cooper Lab

The primary goal of our laboratory is to understand how bacterial populations evolve and adapt to colonize hosts and cause disease. We are particularly focused on how bacterial populations form complex communities within biofilms and how cells perceive cues to attach or disperse. Learn more>

Flynn Lab

My primary interest is in the interaction of pathogens with the host, with special emphasis on the immune mechanisms that protect against or exacerbate disease. Our focus is on Mycobacterium tuberculosis, the organism responsible for tuberculosis, which causes 2 million deaths per year worldwide. Learn more>

Klimstra Lab 

The goal has been to define the host and viral factors that determine the success or failure of the innate immune response to infection with arthropod-borne viruses. Learn more>

Lakdawala Lab

Our lab studies the molecular properties contributing to the epidemiological success of influenza A viruses to better predict future pandemics. There are two main areas of research in my lab 1) exploring the intracellular dynamics of influenza viral RNA assembly and 2) defining the viral properties necessary for efficient airborne transmission of influenza viruses. Learn more>

McClane Lab

Our research is focused on understanding bacterial pathogenesis, which remains a major medical problem in both developing and developed countries. Learn more>

Richardson Lab

The Richardson Lab is primarily focused on the effects of immunometabolism on infectious disease outcomes. Specifically, we study immunometabolism in the context of infections caused by the Gram-positive pathogen Staphylococcus aureus. 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>

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