My laboratory is broadly focused on investigating microbe-microbe interactions, host-microbe interactions, and patient characteristics that influence disease severity. Our primary disease model is catheter-associated urinary tract infection (CAUTI), one of the most common healthcare-associated infections worldwide. Long-term use of indwelling catheters, which is common for management of certain conditions particularly in aging populations, results in continuous urine colonization by bacteria (bacteriuria) and can lead to infections of the bladder (cystitis), kidneys (pyelonephritis), and bloodstream (bacteremia). My lab uses a combination of basic science and patient-oriented research, with the long-term goals of 1) identifying the uropathogen(s) most likely to cause symptomatic infection and adverse outcomes in patients, 2) utilizing an experimental model of infection to identify key virulence factors of these organisms, and 3) developing inhibitors of the identified virulence factors to reduce colonization and infection severity in vulnerable patient populations. As bacteriuria and CAUTI are frequently polymicrobial, which can influence the progression of the infection and the efficacy of antibiotic therapy, a major emphasis of this work is on understanding the contribution of polymicrobial colonization and microbe-microbe interactions to infection progression. We recently identified the Gram-negative bacterium Proteus mirabilis as the most common cause of CAUTI in a cohort of nursing home residents, including cases of polymicrobial infection. P. mirabilis is well-known for its potent urease enzyme, which produces ammonia from the hydrolysis of urea in urine and leads to high urine pH, precipitation of polyvalent ions, and formation of urinary stones (urolithiasis). We recently determined that P. mirabilis urease activity is enhanced during co-culture with other common uropathogens in vitro and in vivo, contributing to increased tissue damage and bacteremia. Enhanced urease activity appears to be a broad phenomenon, occurring with numerous isolates of P. mirabilis during co-culture with most of the other common uropathogens, including Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa, and Klebsiella pneumoniae. Therefore, one project in the lab focuses on defining the underlying mechanism of enhanced urease activity and determining its potential as a therapeutic target. Another focus of the lab is utilizing genome-wide screens to uncover new genes that contribute to pathogenesis during both monomicrobial and polymicrobial infection. We are using transposon insertion-site sequencing (Tn-Seq) to identify the full potential arsenal of P. mirabilis fitness and virulence factors during experimental CAUTI, including core factors that contribute to pathogenicity under all infection conditions tested, and accessory factors that are only required under certain infection conditions.
Eukaryotic Pathogenesis; Microbial Pathogenesis; Microbiology; Molecular and Cellular Biology
Human African trypanosomiasis (commonly called Sleeping Sickness) is one of the global great neglected diseases, causing ~10,000 cases annually according to most recent estimates (2009). The related veterinary disease of livestock (Nagana) also has significant impact on human economic well being throughout sub-Saharan Africa wherever the insect vector (tsetse flies) are found. Both diseases are caused parasitic protozoa called trypanosomes (Trypanosoma brucei ssp.) Because trypanosomes are eukaryotic cells, organized similarly to every cell in our bodies, treatment of infection is not unlike cancer treatment in that chemotherapy against the parasite has harsh consequences for the patient. However, infection is invariably fatal without intervention, consequently new more specific drugs are desperately needed. In addition, because trypanosomes are an anciently divergent evolutionary lineage, they provide a unique model system for studying basic eukaryotic biology. My laboratory focuses on the cell biology of these protozoa, specifically on intracellular trafficking of lysosomal and cell surface proteins as key aspects of the host:parasite relationship. The trypanosome lifecycle alternates between the mammalian bloodstream and the tsetse midgut, and each stage has a unique protein surface coat that forms the first line of contact with the host. These coat proteins are anchored in membranes by glycosylphosphatidylinositol (GPI) anchors and are essential for survival in each stage. Consequently, correct protein targeting to the cell surface is critical to the success of the parasite. Also, endocytic and lysosomal functions are greatly up-regulated in the pathogenic bloodstream stage for both nutritional and defensive purposes. Using classic and current cell biological and biochemical approaches we work on four distinct areas: 1) GPI-dependent targeting of surface coat proteins; 2) machinery of secretory trafficking; 3) stage-specific lysosomal biogenesis and proteomics; and 4) role of sphingolipids in secretory transport. Our ultimate goal is to define aspects of trypanosomal secretory processes that may provide novel avenues to chemotherapeutic intervention.
My research focus is on the study of the interaction of inflammatory leukocytes and fibroblasts with tumor cells in human lung and ovarian tumor microenvironments by using an immunodeficient tumor xenograft model. This model includes the tumor, the tumor-associated stromal fibroblasts and the inflammatory cells, including lymphocytes. In my lab, I work with a research team of students and postdoctoral fellows to study the immune response of cancer patients to their tumors. Our data indicate that tumor-specific lymphocytes, once present in the tumor microenvironment, become hyporesponsive and fail to attack and kill tumor cells. This hyporesponsiveness is due to an arrest or checkpoint in the T cell receptor (TCR) signaling machinery. Our studies are designed to gain a better understanding of the molecular events that are responsible for signaling arrest. We also aim to determine ways to prevent, or even reverse the TCR signaling arrest, for example, by eliminating or blocking the lipid-mediated disruption of the TCR signaling cascade. Using the tumor xenograft models, we have structurally identified the immunoinhibitory factors present within the tumor ascites fluids and determined the mechanism by which they arrest the TCR signaling. We found that these cells fail to respond to activation signals due to the disruption of the TCR signaling cascade that occurs at, or just proximal to the activation of PLC-γ. An identical TCR signaling arrest also occurs in human T cells found in chronic inflammatory tissues. Using the xenograft models, we established that a local and sustained release of IL-12 into the tumor microenvironment activates the quiescent tumor-associated T cells to produce and secrete IFN-γ, which mobilizes an immune-mediated eradication of the tumor. We recently found that lipids present within the ascites fluids of human ovarian tumor mediate a reversible arrest in the TCR signaling pathway of ovarian tumor-associated T cells. We have now determined that extracellular microvesicles (exosomes) isolated from human ovarian tumors and tumor ascites induce a rapid and reversible arrest in the T cell signaling cascade. The T cell inhibition is causally linked to phosphatidylserine that is expressed on the outer leaflet of the exosome membrane. The target of this arrest is diacylglycerol, and the induced suppression is blocked or reversed by diacylglycerol kinase inhibitors. This suggests a likely mechanism by which the tumor-associated exosomes arrest both CD4+ and CD8+ T cell activation. The ability to eliminate or block/reverse that inhibitory activity of the exosomes represents a potentially viable therapeutic target for enhancing patients’ antitumor response and for preventing the loss of function of CAR-T cells upon entry into the tumor microenviroment. These findings will provide valuable information in designing new immunotherapeutic strategies for patients with advanced cancer.
Eukaryotic Pathogenesis; Immunology; Infectious Disease; Microbial Pathogenesis; Microbiology; Molecular Basis of Disease; Signal Transduction; Vision science
Toxoplasma gondii is an obligate intracellular parasite that has the unique ability of infecting most nucleated cells in almost all warm-blooded animals. It is one of the most widespread infections in the world: approximately 50 percent of the world‘s population is infected. Luckily, most infected people are asymptomatic; however, in AIDS patients and other immune-compromised individuals, Toxoplasma causes serious and life-threatening disease. Besides its own medical importance, we study Toxoplasma because it represents an ideal model system to study how other related pathogens cause disease. These include Plasmodium, which is the causative agent of malaria that is responsible for millions of deaths worldwide, and Cryptosporidium, which causes another important secondary infection in AIDS patients. Toxoplasma is a great model system because it can easily be grown in vitro, its genome has been sequenced and it can be genetically manipulated. My research team and I are focused on two different but related questions. First, we want to know how the parasite grows inside of its host cell. One of the important things Toxoplasma must do to grow is hijack host cell pathway and factors. We are using functional genomic assays such as microarrays and genome-wide RNA interference (RNAi) screens to identify these host factors. Identifying them is important because if the parasite cannot use these pathways, the parasite will not grow or cause disease. Thus, these pathways represent novel drug targets. As an example, we discovered that oxygen-regulated transcription factors in the host cell are necessary to support parasite growth. We are currently identifying how these transcription factors function and how the parasite adapts to the various oxygen environments it encounters during its lifecycle. Second, we want to know how Toxoplasma affects the central nervous system and how anti-Toxoplasma immune responses function in the central nervous system. These questions are important because Toxoplasma primarily causes disease in the brain and retina. Our work has revealed that when Toxoplasma actively grows in the brain (a condition known as toxoplasmic encephalitis), it causes a massive reorganization of inhibitory synapses. These changes inhibit GABAergic synaptic transmission and this inhibition is a major factor in the onset of seizures in infected individuals. A second line of research using an ocular infection model has focused on defining how immune responses in the central nervous system are generated by Toxoplasma and then resolved once the infection is under control.
Research in my laboratory focuses on age-associated changes in innate immune responses that render the elderly more susceptible to infections. As the number of individuals above 65 years old is projected to reach 2 billion by 2050, infections in this population poses a serious health and economic burden. A major area of our work is on infections caused by Streptococcus pneumoniae (pneumococcus) that despite the availability of vaccines, remain the leading cause of community-acquired pneumonia in the elderly. Immunosenescence, the age-related decline in immune-cell function, and inflammaging, the age-related increase in basal inflammation, may both contribute to the increased susceptibility of the elderly to life-threatening S. pneumoniae infections such as pneumonia, bacteremia and meningitis. Of particular interest are polymorphonuclear leukocytes (PMN) or neutrophil responses. PMNs are innate immune cells that are key determinants of disease following infection because their initial presence is required to control bacterial numbers, but their persistence in the lungs is detrimental to the host. PMN responses are dysregulated in aging; however, the pathways driving this are not well elucidated. We found that in young hosts, resistance to infection, PMN antibacterial function as well as pulmonary recruitment and resolution following pneumococcal pneumonia is controlled by the extracellular adenosine (EAD) pathway. EAD is produced by the sequential action of two exonucleosidases, CD39 and CD73, and can signal via four known adenosine receptors, that can be pro- or anti-inflammatory. Interestingly, we found that pneumococci can modulate host inflammatory responses by targeting the expression of EAD pathway components. We are using a variety of approaches including in vitro modeling of PMN responses from human donors, mouse models of infection as well as genetic manipulation of bacteria to elucidate the following: 1) How the EAD pathway shapes PMN responses during infection; 2) The role of the EAD pathway in age-driven immune dysregulation; 3) The role of PMNs and the EAD pathway in mounting protective memory responses following vaccination in young and aged hosts; 4) The S. pneumoniae virulence factors required to manipulate the EAD pathway. Elucidating what drives the dysregulated immune responses during aging has the potential of using novel therapies to combat infections in the elderly.
Bacterial Pathogenesis; Infectious Disease; Microbial Pathogenesis; Microbiology
My research interests focus on bacterial pathogenesis, emphasizing bacterial biofilms, antimicrobial therapies and vaccine antigens. One major area of my research lab is otitis media (OM) or middle ear disease. Approximately 80 percent of children experience one episode of OM while others have recurrent infections. Chronic OM infection causes hearing impairment leading to developmental problems as these children reach school age. My laboratory has concentrated on two major causes of OM, Moraxella catarrhalis and Streptococcus pneumoniae. Our recent work suggests that M. catarrhalis colonization predisposes patients to colonization with S. pneumoniae in polymicrobial biofilms. The goals of this work are to define biofilm-associated factors and to identify signals that induce bacteria to transition from asymptomatic colonizers to pathogenic organisms leading to OM. Our second major research focus is the identification of novel antimicrobial therapies. Chronic OM is likely a biofilm-associated disease and biofilms are highly antibiotic resistant. Antibiotic resistance is a major problem worldwide and new drug development is both time consuming and extremely expensive. We have demonstrated that photodynamic therapy (PDT), an FDA-approved cancer treatment, is also bactericidal against the three major otopathogens. Thus, the goal of this research is to adapt PDT into a clinically effective treatment for chronic OM. Our third research area involves novel antimicrobial treatments for orthopedic/prosthetic infections. Infections after orthopedic intervention, including knee/hip replacements and insertion of prosthetic devices, are devastating to the patient and these infections will likely increase over the next 20 years. This is particularly relevant to the military where improvised explosive devices cause severe extremity injuries requiring amputation. Antibiotic-resistant biofilms are the primary source of these infections. In collaboration with colleagues at UB, we are testing a novel electrical stimulation method for prevention/eradication of biofilm infections on implant materials. The goals of this research are to define the optimal antimicrobial parameters that are broadly effective against multiple pathogens, including Staphylococcus aureus, Acinetobacter baumannii, Staphylococcus epidermidis and Klebsiella pneumoniae. The members of my research team typically include a combination of graduate students, lab technicians and a junior faculty member. In the summer, I usually mentor medical students or undergraduates who are interested in the fundamentals of basic science and translational research focused on microbial pathogenesis.
Allergy and Immunology; Medical Microbiology; Infectious Disease; Microbiology; Genomics and proteomics; Immunology; Microbial Pathogenesis; Molecular and Cellular Biology; Molecular Basis of Disease; Molecular genetics; Gene Expression; Signal Transduction; Protein Function and Structure; Bacterial Pathogenesis
Research efforts in my laboratory are focused in the fields of immunology and bacterial pathogenesis, two diverse fields of biomedical research for which I have two separate research groups. Projects in both fields are performed by undergraduates, doctoral and master’s degree students, postdoctoral fellows and senior research associates. One major focus of my laboratory is studying the regulation of mucosal immune responses. We investigate the cellular and molecular events by which Type II heat-labile enterotoxins (HLTs), produced by certain strains of Escherichia coli, modulate immune responses. We have demonstrated that LT-Ilia, LT-IIb and LT-IIc, when co-administered with an antigen, have the capacity to enhance antibody and cellular immune responses to that antigen. Using a variety of immunological and cellular technologies, including flow cytometry, fluorescence resonance energy transfer (FRET) detection, cytokine multiplex analysis, mutagenesis, quantitative Reverse Transcription PCR (qRT-PCR), RNA-sequencing (RNA-Seq) and a variety of transgenic mice, we are investigating the mechanisms by which these immunomodulators productively interact with various immunocompetent cells (T cells, B cells, dendritic cells, macrophages) to induce or suppress cytokine production, costimulatory ligand expression and cellular proliferation. A practical outgrowth of these experiments is the potential to engineer novel recombinant vaccines by genetically fusing antigens from different pathogens to the enterotoxins. Recent experiments have shown that these HLT are lethal for triple-negative breast cancer cells, which has opened a new area of oncological research for the lab. A second focus of my laboratory is to investigate the molecular mechanisms by which adherent-invasive Escherichia coli (AIEC) induce, exacerbate or prolong the symptoms of inflammatory bowel disease (IBD) and Crohn’s disease, two acute and chronic inflammatory diseases of the human gut. In vitro, AIEC strains invade into the cytoplasm of several epithelial cell lines. Using recombinant screening methods and RNA-Seq technologies, we are identifying the genes of AIEC that are required to attach and to invade gut cells.
Infectious Disease; Infectious Disease; Microbial Pathogenesis; Vitamins and Trace Nutrient
I care for patients who are hospitalized at Erie County Medical Center where I also serve as the hospital epidemiologist addressing infection control. I teach medical students, residents, and fellows in both hospital and classroom settings. In UB’s schools of medicine and dentistry, I teach a variety of topics including microbiology, pharmacology and toxicology, oral biology, and gastrointestinal systems, host defenses, and global health. I also conduct laboratory research on diarrhea-producing strains of E. coli bacteria. My lab focuses on enteropathogenic Escherichia coli (EPEC), Shiga-toxigenic E. coli (STEC, aka EHEC) and enterotoxigenic E. coli (ETEC). We are working on the role of intestinal host defenses such as nitric oxide and on the immune modulatory effects of adenosine. We have discovered that zinc can directly inhibit the virulence of pathogenic bacteria, and we are working on turning these laboratory findings into treatments. In our work on zinc we collaborate with Michael Duffey, PhD, in the Department of Physiology and Biophysics. Recently we have discovered that zinc can inhibit the development of resistance to antibiotics in Escherichia coli and other bacteria. Zinc does this by its ability to inhibit the SOS response, a bacterial stress response triggered by damage to the bacterial DNA. We are collaborating with Dr. Mark Sutton of Biochemistry to better determine the mechanism of zinc in this regard. I am interested in international medicine and global health and participate in an annual medical mission trip to Honduras, a trip in which student volunteers are encouraged to participate. Closer to home, I am a volunteer physician at Good Neighbors Health Center, a free clinic for the underserved on Jefferson Avenue in Buffalo. Resident physicians are encouraged to volunteer, and students may also be able to arrange clinical experiences. I am Co-Medical Director, with Dr. Ryosuke Osawa, of the Erie County TB Clinic. Learning experiences in my laboratory, in infection prevention and hospital epidemiology, or in international health, may be available for motivated students, residents, and fellows.
The human immunodeficiency virus (HIV) is now considered a chronic disease in the developed world. In underdeveloped areas where access to antiretroviral therapy is limited, however, it remains a devastating disease contributing to grave socioeconomic problems. The goal of my research is to expand our knowledge of pathogen interactions with cellular membranes by developing a detailed understanding of the mechanism of HIV entry and by studying co-infection of HIV with the pathogenic fungus Cryptococcus neoformans in human macrophages. The first step of HIV infection is HIV entry when the envelope protein complex on the surface of the virus comes into contact with the cellular receptors, glycoprotein CD4 and coreceptor, and mediates merging of the viral and cellular membranes leading to delivery of the viral genetic material. Mechanistic studies help to inform the development of inhibitors to HIV entry that will be beneficial on both therapeutic and prophylactic levels. The envelope protein complex is the machinery that gets the virus into the cell; as such, it is also a prime target for the development of vaccines. HIV/AIDS often kills by priming the host for opportunistic infections. Cryptococcal meningitis is one of the leading killers of AIDS patients. The human macrophage is the cell type tasked with ingesting and clearing microbes. In my lab, we are working to define the role of the human macrophage in the copathogenesis of the opportunistic fungus Cryptococcus neoformans and HIV during AIDS progression. The mechanisms of host-microbe interactions also serve as templates for the design of novel drug regimens, including immunotherapy. We have recently utilized our extensive experience in the study of how HIV enters the cell to begin studies in Ebolavirus entry which has a similar mechanism. We are developing inhibitors to the process of Ebolavirus entry and using developments in inhibition to study the mechanism of attachment and membrane fusion into multiple cell types. It is my objective throughout my career to provide vital basic research in virology and cell biology in order to advance medical treatment and prevention. As an academic researcher, I put a strong emphasis on the training and mentoring of young scientists in my lab, and I participate in the T35 training grant from the National Institutes of Health that UB and Roswell Park Cancer Institute jointly secured. I train master’s and PhD students as well as postdoctoral fellows in the departments of Microbiology and Immunology and Biochemistry. I also mentor undergraduates in research projects; these students may come to me independently or through UB’s Center for Undergraduate Research and Creative Activities (CURCA) in which I am active. I direct undergraduate studies for my department, and I am the course director for Biomedical Microbiology, my department’s large undergraduate basic science course.
Infectious Disease; Microbiology; Molecular and Cellular Biology; Molecular genetics; DNA Replication, Recombination and Repair; Virology; Genome Integrity
The major focus of my laboratory is in understanding the molecular machines that make up the DNA replication forks of the small human DNA viruses, polyoma- and papillomaviruses. Papillomaviruses and polyomaviruses are human pathogens; human papillomavirus (HPV) results in a vast number of human cancers, and the human polyomaviruses JC and BK cause serious disease and death in immunocompromised patients. Both viral systems provide important models for the study of human DNA replication mechanisms and have allowed for vital insights into eukaryotic DNA replication. The study of polyomavirus DNA replication led to the first identification of many cellular DNA replication complexes and processes; papillomavirus has provided the best structures and models to date of replicative hexameric DNA helicases and how they function. I typically train undergraduate, master’s and doctoral students and postdoctoral scholars, assistant research professors and laboratory technicians. My laboratory focuses on two primary areas. One is elucidating the dynamic protein-protein interactions that allow the series of enzymes required to replicate DNA to act in concert and in the correct sequence required to duplicate the genome. My laboratory has been at the forefront of identifying the interactions between the one critical HPV DNA replication protein, the origin-binding DNA helicase, E1, and cellular DNA replication proteins. Understanding these interactions and the roles they play in the HPV DNA replication process has helped our understanding of, and continues to lead to information that tells us more about how both viral and eukaryotic DNA replication forks function. In addition, as we identify protein-protein interactions between HPV E1 and cellular factors that are essential for HPV DNA synthesis, we will uncover potential targets for development of broad-range HPV antivirals that could act to block HPV replication. We recently obtained a large multilaboratory NIH research grant to investigate just this possibility for the interaction between HPV E1 and the human DNA replication protein, Topoisomerase I. The second primary area of investigation is elucidating how the cellular DNA damage response (DDR) pathways inhibit DNA replication when cells are subjected to DNA damage. For many years, the DDR field focused on the effects of DDR on the cell cycle kinases as the only method by which DNA replication was arrested. In the mid- to late-2000s, researchers recognized that in mammalian cells there is also a substantial (tenfold) inhibition of elongation of DNA replication following DDR. The mechanisms for this inhibition are unknown. Using both in vitro and cell-based simian virus 40 (SV40) DNA replication systems, we have shown that SV40 DNA replication is also shut down in response to DDR kinase pathways and that this is not based on cell cycle kinase action. Therefore, SV40 provides a useful model system for determining how elongation of DNA replication is inhibited by DDR. Furthermore, we have shown that in contrast HPV DNA replication does not respond to DDR, providing us an important control DNA replication system for these studies. (The lack of DDR arrest of HPV DNA replication likely explains why HPV integrates so readily into host cell chromosomes−an important step for HPV-induced carcinogenesis). Our studies on the DDR effect on polyoma and papilloma virus DNA replication will lead to insights into the effect of DDR on cellular DNA replication as well as an understanding of how HPV integrates into host cell chromosomes causing HPV-induced cancers.
Microbial Pathogenesis; Molecular and Cellular Biology; Gene Expression; Regulation of metabolism
The adaptive success of bacteria depends, in part, on the ability to sense and respond to their environment. Metals such as iron and manganese are important nutrients that can often be limiting, and therefore cellular metabolism must be modified to either scavenge the nutrients or use alternative processes that do not require the metal. Bradyrhizobium japonicum belongs to a group of related organisms that form a close or intracellular relationship with eukaryotes in a pathogenic or symbiotic context. This bacterium serves as a model to study related pathogens that are refractive to genetic and biochemical study. Our lab seeks to understand the mechanisms by which cells maintain iron homeostasis at the level of gene expression. We discovered the global transcriptional regulator Irr that controls iron-dependent processes. Irr is stable only under iron limitation, where it positively and negatively controls target genes. We are interested in understanding the mechanism of this conditional stability, how Irr regulates genes, and the functions of numerous genes under its control. Moreover, Irr integrates iron homeostasis with manganese metabolism, providing a link between the two nutrients. Identifying the Irr regulon and iron stimulon has given us important clues into how bacteria traffic iron into and out of the cell. Recent work suggests that B. japonicum not only adapts at the level of gene expression, but can mutate rapidly to accommodate new nutritional sources in the environment.
Eukaryotic Pathogenesis; Gene Expression; Infectious Disease; Microbial Pathogenesis; Microbiology; RNA; Signal Transduction
There are estimated to be over one million species of fungi on the earth, yet very few of these species are capable of causing deadly systemic infections in humans. One of the major limiting factors for most fungi is their inability to grow at mammalian core body temperature. We utilize the fungal pathogen Cryptococcus neoformans var. grubii as a representative fungal pathogen to understand how these few fungi have adapted to growth at mammalian body temperature. C. neoformans is a worthy pathogen, as it is estimated to cause over 500,000 deaths from meningoencephalitis per year, primarily in Africa and Southeast Asia as an HIV/AIDS comorbidity. We use the temperature-limited Cryptococcus amylolentus as a comparator; it is an environmental strain that produces similar virulence factors to C. neoformans and is fully virulent in surrogate invertebrate hosts at permissive temperatures. We have discovered that host temperature adaptation in C. neoformans is accompanied by a reprogramming of gene expression at the level of messenger RNA (mRNA) stability. In response to temperature stress, C. neoformans rapidly degrades mRNAs that encode energy consuming machinery such as ribosomes. At the same time, it prioritizes the translation of stress-responsive mRNAs on existing ribosomes. Because mRNA synthesis and decay are coupled processes, we seek to identify the protein components of mRNA complexes that mediate the specificity of this decay process and posttranslational modifications, such as arginine methylation and phosphorylation, that modify their function. In addition, we are investigating the signaling pathways that accelerate or slow mRNA decay in response to specific environmental stimuli such as host temperature and nutrient deprivation. Finally, mRNA decay not only alters gene expression at the posttranscriptional level, but the degradation of abundant mRNAs during stress releases nucleotide intermediates that can be utilized by the stressed cell to promote genome stability. We are investigating the process of mRNA degradation as well as nucleotide metabolic pathways as drug targets in C. neoformans and other fungal pathogens. Our goal is to define the unique attributes of C. neoformans that confer pathogenicity and to identify potential targets for novel therapeutics. Each of my students has a project that contributes to the overall goals of my research team. Students in my laboratory work independently, though with frequent interaction with me regarding the direction of investigation and interpretation of data. Regular meetings allow us to provide input on each other’s projects. I expect my students to present their work at least once per year at a national or international meeting, and I expect them to do the bulk of the work in writing papers describing their findings for publication.
Eukaryotic Pathogenesis; Gene Expression; Genomics and proteomics; Infectious Disease; Microbial Pathogenesis; Microbiology; Molecular and Cellular Biology; Molecular genetics; Protein Function and Structure; RNA
Trypanosoma brucei is a eukaryotic pathogen that causes human African trypanosomiasis, a disease that is invariably fatal if not treated. Essential and novel processes in this parasite may serve as starting platforms for new chemotherapeutics, which are urgently needed. Our laboratory combines biochemical, genetic, genomic and proteomic approaches toward understanding gene regulation and protein modification in this pathogenic eukaryote. One focus in my laboratory is RNA editing, a novel mechanism for regulating mitochondrial gene expression in which sequence information is added to mRNAs after transcription by specific insertion and deletion of uridine residues. RNA editing is essential for creating translatable open reading frames (ORFs). We are performing functional and biochemical characterization of the large, dynamic RNA-protein complex termed MRB1, which coordinates multiple aspects of the RNA editing process. A second focus is on regulating RNA stability and translational control in T. brucei, which constitute the major methods of gene regulation in this organism. We identified an RNA binding protein, DRBD18, that impacts the stabilities of hundreds of mRNAs. Our data support a model in which posttranslational modification of DRBD18 by arginine methylation acts as a switch to change DRBD18 from an mRNA destabilizer to an mRNA stabilizer by regulating specific protein-protein and protein-RNA interactions. We are testing this model in vitro and in vivo using reporter assays, in vivo protein-RNA cross-linking and protein-protein interaction assays. A third focus is on understanding the mechanisms by which protein arginine methylation modulates trypanosome biology. We performed a global proteomic analysis of the arginine methylome of T. brucei, identifying >1100 methylproteins spanning most cellular compartments and a wide array of functional classes. We are now analyzing novel mechanisms of protein arginine methyltransferase regulation and defining the physiological and molecular functions of arginine methylmarks on selected proteins. I foster a collaborative and flexible laboratory environment, and I encourage my students to explore the research topics that interest them.
Periodontics; Operative Dentistry; Oral Biology; Pediatric Dentistry
Research in my laboratory encompasses the general area of oral infection and immunity with a major focus on adhesin-mediated interactions of oral bacteria with host salivary or cellular receptors. We investigate the glycoproteins in saliva that are recognized by lectin-like microbial adhesins. It is our long-term goal to better understand the modulating role of salivary glycoproteins in supporting tissue tropism of a benign commensal oral microflora to the human oral cavity and in host defense against pathogenic microorganisms. A more detailed description of our current research activities can be found on the following website:
Infectious Disease; Infectious Disease; Microbial Pathogenesis
I am an expert in infectious diseases, and I care for hospitalized patients at the Buffalo VA Medical Center (Buffalo VAMC). I have an active, nationally funded translational research program. My research focuses on Gram-negative bacilli (GNB), including Escherichia coli, Acinetobacter baumannii and a new hypervirulent variant of Klebsiella pneumoniae. These GNB cause infection in nearly every nonintestinal site in the body. The hypervirulent variant of K. pneumoniae is both fascinating and worrisome. Unlike its predecessors, it is capable of causing infection in young, healthy hosts and spreading nearly anywhere in the body from the initial infected site, including the eyes and brain. GNB-caused infections result in the loss of billions of health care dollars, millions of work days and hundreds of thousands of lives each year. GNB are becoming increasingly resistant to antibiotics, including strains that have become resistant to all available antibiotics. Unfortunately, there are virtually no new antimicrobial agents active against highly resistant GNB in the pharmaceutical “pipeline.” To address this formidable clinical challenge, my collaborators and I have increased our understanding of the bacterial factors that are critical for these GNB to cause infection. We use this information to develop vaccines that will prevent infection and antibodies that can be used to treat infection. My UB collaborators include Dr. Campagnari (microbiology), Dr. Gulick (structural biology) and Drs. Elkin and Zola (biomedical informatics). My research also involves identifying potential bacterial drug targets; this information will be used to develop new classes of antibiotics. I intermittently have students in my lab, and I participate in a grant designed to encourage medical students to become physician-scientists. I welcome interested students to contact me about conducting research with me. The Buffalo VAMC is the site of my clinical teaching. I teach first- and second-year medical students in lecture settings and small group sessions, including courses in lung respiration, musculoskeletal, renal and microbiology-immunology. Residents attend my grand rounds; I also teach fellows in all aspects of their training and mentor those who perform their research projects in my lab.
Retroviruses comprise a large and diverse family of RNA viruses that can infect a variety of hosts and can lead to immune system dysfunction and cancer. The most well known member of this family is HIV (Human Immunodeficiency Virus), which is responsible for millions of deaths every year. Immune cells, such as CD4+ T cells, are the targets of HIV infection resulting in their subsequent destruction and the overall impairment of the immune system. As a result of the deterioration of the immune system, the host is unable to fight effectively infections and some other diseases. Opportunistic infections or cancers take advantage of the weakened host, which can prove to be lethal. Other members of the retrovirus family, Murine Leukemia Virus (MLV) and Mouse Mammary Tumor Virus (MMTV) are common pathogens of mice that are used in research to study the interplay between the host and retroviruses and have served as models for HIV and other human retroviruses. In the context of the constant struggle between host cells and pathogens, cells have developed early innate immune and cell-intrinsic strategies to counteract retroviruses. Therefore, retroviruses have developed a variety of sophisticated mechanisms to counteract cellular responses and allow for productive infections to occur. Due to the complexity of the antiviral immune response, a full understanding of host-pathogen interactions requires the integration of in vitro and in vivo data, where the role of cellular restriction factors and the innate immune response are examined in a living organism. Infections in mouse models have provided important and at times surprising insights into the relationship between hosts and pathogens. Thus one of the areas of focus for our lab will be the integration of in vivo and in vitro models to study the interaction of novel cellular host factors and retroviruses. HIV and other lentiviruses devote a relatively large portion of their genome to accessory proteins that counteract those cellular host factors. Understanding the function of these accessory proteins has provided insight into the intrinsic defenses utilized by the cell to block viral infections, as well as generated potential targets for antiviral interventions. However, there is no currently tractable in vivo model for studying cell host restriction factor/accessory protein interactions. Hence, the second major focus of our lab will be the development of in vivo models to examine the interplay between HIV accessory proteins and host cell intrinsic immunity.
Bioinformatics; Gene Expression; Genomics and proteomics
The recent development of high-throughput genomics technologies is revolutionizing many aspects of modern biology. However, the lack of computational algorithms and resources for analyzing massive data generated by these techniques has become a rate-limiting factor for scientific discoveries in biology research. In my laboratory, we study machine learning, data mining and bioinformatics and their applications to cancer informatics and metagenomics. Our work is based on solid mathematical and statistical theories. The main focus of our research is on developing advanced algorithms to help biologists keep pace with the unprecedented growth of genomics datasets available today and enable them to make full use of their massive, high-dimensional data for various biological enquiries. My research team is working on two major projects. The first is focused on metagenomics, currently funded by the National Institutes of Health (NIH), the National Science Foundation (NSF) and the Women’s Health Initiative. Our goal is to develop an integrated suite of computational and statistical algorithms to process millions or even hundreds of millions of microbial genome sequences to: 1) derive quantitative microbial signatures to characterize various infectious diseases, 2) interactively visualize the complex structure of a microbial community, 3) study microbe-microbe interactions and community dynamics and 4) identify novel species. We collaborate with researchers throughout the University at Buffalo, notably those in the School of Medicine and Biomedical Sciences, the School of Public Health and Health Professions and the College of Arts and Sciences. The second project focuses on cancer progression modeling. We use advanced computational algorithms to integrate clinical and genetics data from thousands of tumor and normal tissue samples to build a model of cancer progression. Delineating the disease dynamic process and identifying the molecular events that drive stepwise progression to malignancy would provide a wealth of new insights. Results of this work also would guide the development of improved cancer diagnostics, prognostics and targeted therapeutics. The bioinformatics algorithms and software developed in our lab have been used by more than 200 research institutes worldwide to process large, complex data sets that are core to a wide variety of biological and biomedical research.
DNA Replication, Recombination and Repair; Gene Expression; Genome Integrity; Microbiology; Molecular and Cellular Biology; Protein Function and Structure; Signal Transduction
We are interested in developing an integrated mechanistic view of how organisms coordinate the actions of their DNA replication machinery with those of other cellular factors involved in DNA repair and damage tolerance. Failure to properly coordinate these functions leads to mutations, genome instability, and in extreme cases, cell death. We utilize a combination of biochemical, biophysical, and genetic approaches to investigate the molecular mechanisms of DNA replication, DNA repair, and error-prone DNA damage tolerance functions in Escherichia coli. The primary mechanism for damage tolerance involves direct bypass of damaged bases in the DNA. This process is inherently error-prone, and is the basis for most mutations. Current efforts are focused on understanding the mechanisms by which the actions of high fidelity and error-prone lesion bypass DNA polymerases are coordinated with each other, as well as other proteins involved in DNA metabolism. Our goal in this work is to develop methods that enable us to control the fidelity of DNA repair for therapeutic gain. We are also interested in understanding the mechanisms that contribute to DNA mutagenesis in the opportunistic human pathogen, P. aeruginosa. P. aeruginosa is a particular problem for individuals afflicted with cystic fibrosis. Persistent colonization of cystic fibrosis airways with P. aeruginosa serves as a major source of morbidity and mortality for these patients. The ability of P. aeruginosa to persist in the airways relies in part on its ability to adapt to the continuously changing environment within the diseased airways. We are particularly interested in determining the contribution of mutagenesis and DNA repair to adaptive mutations that contribute to clonal expansion and pathoadaptation of P. aeruginosa during colonization of cystic fibrosis airways.
Infectious Disease; Microbiology; Microbial Pathogenesis; Molecular and Cellular Biology; Gene Expression; Transcription and Translation; Protein Function and Structure; RNA; Eukaryotic Pathogenesis
In my laboratory, we use molecular biological and biochemical approaches to study Trypanosoma brucei, the causative agent of African sleeping sickness, and Trypanosoma cruzi, which causes Chagas disease in South and Central America. Treatment for these diseases is severely limited due to increasing drug resistance and lack of available drugs. The goal of our work is to discover and exploit critical events that occur in the parasite life cycle that may be used to prevent growth or transmission of the parasite. The major project in my laboratory examines the ribosome, the complex molecular machine that drives protein synthesis. While many features of the ribosome and its assembly pathway are conserved in the parasites we study, we have identified features that are very different from the human host. Our laboratory discovered a pair of trypanosome-specific RNA binding proteins, P34 and P37, that are part of a unique preribosomal complex that is essential for ribosomal biogenesis and survival of trypanosomes. This may suggest that the interaction of these proteins with other components of the ribosomal assembly pathway can be developed as targets for chemotherapy. We are developing a high-throughput screen for small molecules that disrupt the complex in trypanosomes and do not harm the human host. My team and I also collaborate with Dr. Joachim Frank at Columbia University on a project to examine the structure of the ribosome and intermediates in the pathway of assembly using cryo-electron microscopy (cryo-EM). These experiments will provide important information about the unique features of the structure and function of the trypanosome ribosome and further our discovery of potential drug targets. In addition, we continue in a long-standing collaboration with Dr. Beatriz Garat at the Universidad de la Républica in Uruguay, examining both DNA and RNA binding proteins which regulate gene expression in Trypanosoma cruzi. The balance of graduate, undergraduate and medical students and postdoctoral researchers I mentor changes from year to year, though the international quality I strive to maintain has distinguished my laboratory for years: I enjoy having students from around the world as part of my research team. I am the course director for, and lecture in Critical Analysis and Eukaryotic Pathogens. I am also the course director for Eukaryotic Gene Expression and the co-course director for Molecular Parasitology.
Immunology; Infectious Disease
The focus of my laboratory is to understand regulatory mechanisms during infection and autoimmunity at mucosal sites, particularly within the gastrointestinal tract. The adult human intestine alone contains up to 100 trillion micro-organisms─and no other tissue is submitted to a greater level of antigenic pressure than the gut, which is constantly exposed to food and environmental antigens and the threat of invasion by pathogens. At birth, for example, the human gastrointestinal tract undergoes a massive exposure to these antigens, and throughout the average human life there are multiple instances of the remodeling of the gut flora following infection. All these occurrences impose a unique challenge to the gastrointestinal environment. In response, to maintain immune homeostasis, the intestinal immune system has evolved redundant regulatory strategies. Several subsets of immune cells with immune modulatory function reside within the gastrointestinal tract. Specifically, we study Foxp3 expressing regulatory T cells (Tregs), which play a central role in controlling intestinal homeostasis. Recent studies have demonstrated that the ability of Tregs to control defined polarized settings requires plasticity, the acquisition of characteristics specific to the glycoprotein CD4+ T effector subsets. Such adaptation comes with an inherent cost, however; as my research team and other researchers have demonstrated, in extreme instances of inflammation such adaptation can actually be associated with the expression of pro-inflammatory effector cytokines (i.e., interferon gamma and interleukin 17A). We recently identified GATA3, the canonical Th2 transcription factor, as a critical regulator of Treg adaptation during inflammation in tissues. Our goal is to understand how GATA3 regulates this and to identify other factors involved in Treg adaptation during inflammation. Our laboratory employs natural enteric parasitic infections of mice and the T cell dependent model of colitis to decipher both the environmental cues and cell- intrinsic requirements for Treg cell plasticity, stability and function at mucosal sites. The ultimate goal of our research is to clarify the pathogenesis of inflammatory bowel disease (IBD) and develop novel treatment modalities for patients.