Apoptosis and cell death; Bioinformatics; Molecular Basis of Disease; Molecular and Cellular Biology; Molecular genetics; Neurobiology; Regulation of metabolism
My laboratory studies the cell-autonomous and non-cell-autonomous mechanisms of axon degeneration, a process akin to programmed cell death. In other words, we are attempting to elucidate what causes axon breakdown from within neurons and which external (glial) events trigger axon loss. Degeneration of axons is a hallmark in many neurodegenerative conditions, including those associated with abnormal glia. We have great hope that understanding why and how axons degenerate may lead to more efficient neuroprotective therapies tailored specifically to support axons and their surrounding glia. Axons are the longest cellular projections of neurons relaying electrical and biochemical signals in nerves and white-matter tracts of the nervous system. As such, they are critical for neuronal wiring and transport of neuronal maintenance signals. Because of their incredible length and energetic demand (human motor neurons can be one meter long), however, axons are very vulnerable and at continuous risk of damage. Axons do not exist in isolation but are inextricably and intimately associated with their enwrapping glia (Schwann cells and oligodendrocytes) to form a unique axon-glia unit. The most relevant neurological symptoms in a number of debilitating neurodegenerative conditions are due to compromised axon integrity. Thus, neuroprotective therapies promoting axon stability have great potential for more effective treatment. Recent studies indicate that axonal degeneration, at least in experimental settings, is an active and highly regulated process akin to programmed cell death (‘axonal auto-destruction’). Moreover, it is increasingly realized that axonal maintenance relies not only on neuron-derived provisions but also on trophic support from their enwrapping glia. The mechanism for this non-cell-autonomous support function remains unknown, but emerging evidence indicates that it is distinct from the glial role in insulating axons with myelin. We are pursuing the intriguing question of whether abolished support by aberrant delivery of metabolites and other trophic factors from glia into axons is mechanistically linked to the induction of axonal auto-destruction. This concept is supported by our recent finding that metabolic dysregulation exclusively in Schwann cells is sufficient to trigger axon breakdown.
Molecular genetics; Protein Function and Structure; DNA Replication, Recombination and Repair; Bacterial Pathogenesis
My associates and I use a combination of biochemical and biophysical approaches to study the molecular basis of stalled DNA replication fork rescue. Our model organism is the well-characterized bacterium Escherichia coli (E. coli), since the majority of the proteins thought to be involved in fork rescue are known. Most of our experimental work is concerned with the function and regulation of the complexes that control fork rescue, with studies focused primarily on the role of the single-strand DNA binding protein (SSB) and several recombination DNA helicases. Comparative studies are also underway using selected components of some medically relevant bacterial organisms. We collaborate with scientists from the National Institutes of Health (NIH) and other research institutions. The team working in my lab consists of undergraduate and graduate students, postdoctoral fellows and a technician. We seek to understand fork rescue utilizing both bulk-phase and single molecule techniques. Typically, studies focus initially on purification and characterization of the various proteins (there are now more than 10 being studied). We study DNA binding, unwinding and the hydrolysis of adenosine triphosphate (ATP) using a combination of modern spectroscopic (both ultraviolet–visible and fluorescence) and equilibrium binding methods. The goal of these initial studies is to understand the range of DNA substrates on which an enzyme can act, as a means to understanding its role in vivo. This is followed by careful single molecule studies using a technique I pioneered that combines optical tweezers, microfluidics and high-resolution fluorescence microscopy. My research team is also pursuing a new area of research targeted at developing small molecule inhibitors. These are aimed at disrupting binding between SSB and the 12-14 proteins comprising the SSB-interactome. As SSB is an essential protein and its binding to interactome partners is required for viability, the goal of these studies is to identify inhibitors that will be further developed into novel antibiotics.
Reproductive Endocrinology; Apoptosis and cell death; Cell growth, differentiation and development; Endocrinology; Gene Expression; Molecular genetics; Signal Transduction; Toxicology and Xenobiotics; Vitamins and Trace Nutrient
I am an Associate Professor in the Department of Pharmacology & Toxicology of the School of Medicine and Biomedical Sciences. I have a secondary appointment as Adjunct Associate Professor in the Department of Oral Biology in the School of Dental Medicine. My research interests focus on the study of how hormones and nutrients affect cell growth, differentiation, and survival. I study these processes in bone osteoblasts and breast normal epithelial cells as well as cancers of both tissues. I have discovered how natural estrogens as well as dietary phytochemicals sustain osteoblast longevity and contribute to bone growth. In collaboration with Dr Atif Awad of the University at Buffalo School of Public Health, I have identified dietary factors that inhibit the growth of cancers of the prostate and breast. We have published primary research papers, significant review articles, and two books entitled "Nutrition and Cancer Prevention" and Adipose Tissue and Inflammation".
Bioinformatics; Genomics and proteomics; Molecular and Cellular Biology; Molecular genetics; Gene Expression; Transcription and Translation
Our research group is interested in how regulatory proteins are targeted to the correct DNA binding sites at the correct time. Transcription factors are directed to their genomic targets by DNA sequence, local chromatin structure, and protein-protein interactions. These modulators of transcription factor binding are not independent but function both cooperatively and competitively to regulate where transcription factors bind. Understanding how these modulators affect transcription factor binding in vivo remains a major unsolved biological problem. We use the model organism Saccharomyces cerevisiae to address the disconnect between the presence of the correct DNA binding sequence and true regulatory protein binding, integrating both experimental and computational approaches to: i) investigate transcription factor binding in response to environmental stress, ii) identify and characterize the mechanisms directing transcription factor target selection, and iii) and develop bioinformatics tools to analyze and interpret ChIP-seq experiments and chromatin structural patterns.
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.
Autoimmunity; Bioinformatics; Genomics and proteomics; Immunology; Infectious Disease; Molecular and Cellular Biology; Molecular genetics; Neurobiology
My primary research is in the field of biomedical ontology development. An ontology is a controlled, structured vocabulary intended to represent knowledge within a particular domain. Terms in an ontology have logical relationships to each other and to terms in other ontologies, to allow for reasoning and inference across the ontology. Biomedical ontologies allow annotation and integration of scientific data within particular fields of science and medicine, and their careful curation and logical structure facilitate data analysis. My work in biomedical ontology is strongly informed by my earlier experience in laboratory research in immunology, genetics, molecular biology and virology. My research group works on ontologies for both basic and clinical applications, in collaboration with researchers both at UB and other institutions. I led efforts to revise and extend the Cell Ontology, which is intended to represent in vivo cell types from across biology. We worked extensively to bring it up to community-accepted standards in ontology development, placing particular emphasis on improving the representation of hematopoietic cells and neurons. We are developing the Cell Ontology as a metadata standard for annotation and analysis of experimental data in immunology in support of the National Institute of Allergy and Infectious Diseases (NIAID) ImmPort Immunology Database and Analysis Portal and Human Immunology Project Consortium. We have also developed ways to use the Cell Ontology in support of the analysis of gene expression data linked to cell types and have contributed to the Functional Annotation of the Mammalian Genome (FANTOM) 5 Consortium‘s work on identifying gene transcription start sites across multiple cell types and tissues. My research team is also developing the Neurological Disease Ontology to represent clinical and basic aspects of neurological diseases in order to support translational research in this area. In collaboration with clinical colleagues at UB, we are initially focusing on Alzheimer’s disease and dementia, multiple sclerosis and stroke. We have as well developed a companion ontology, the Neuropsychological Testing Ontology, to aid in the annotation and analysis of neuropsychological testing results used as part of the diagnosis of Alzheimer‘s disease and other neurological diseases. I am a long-term member of the Gene Ontology (GO) Consortium and have a particular interest in the representation of immunology and neuroscience in the GO. I am also involved in UB’s contribution to the Protein Ontology and contribute as well to the work of the Infectious Disease Ontology Consortium, Immunology Ontology Consortium and Vaccine Ontology Consortium. I teach and mentor students at the master’s and doctoral levels, and advise undergraduate, graduate, and medical students in summer research projects as well.
Neurology; Cytoskeleton and cell motility; Molecular and Cellular Biology; Molecular Basis of Disease; Molecular genetics; Neurobiology; Signal Transduction; Inherited Metabolic Disorders; Transgenic organisms
My laboratory seeks to understand the molecular basis of myelination and myelin diseases. Myelin is a multi-lamellar sheath that invests large axons and permits rapid conduction of nerve signals. Failure in myelin synthesis and myelin breakdown cause several important neurological diseases, including multiple sclerosis, leukodystrophies and peripheral dysmyelinating neuropathies. In some of these diseases, genetic mutations cause defects in cytoskeletal, adhesion and signaling molecules. I work with a team of undergraduate and graduate students, postdoctoral fellows, technicians, senior scientists and many international collaborators to discover how these molecules normally coordinate cell-cell and cell-extracellular matrix interactions to generate the cytoarchitecture of myelinated axons. We use a variety of approaches, including generation of mice carrying genetic abnormalities, cultures of myelinating glia and neurons, imaging, biochemistry and morphology to understand the role of these molecules in normal and pathological development. By comparing normal myelination to the abnormalities occurring in human diseases, we aim to identify molecular mechanisms that pharmacological intervention might correct. For example, we described how the protein dystroglycan associates with different proteins, some of which impact human neuropathies, depending on a proteolitic cleavage that can be regulated to improve the disease. Similarly, we found that molecules such as integrins and RhoGTPAses are required for glia to extend large processes that will become myelin around axons. In certain neuromuscular disorders, defective signaling pathways that converge on these molecules cause failure to produce or mantain an healthy myelin Finally, in collaborations with scientists and clinicians in the Hunter J. Kelly Research Institute, we are generating transgenic forms of GalC, an enzyme deficient in Krabbe leukodystrophy, to investigate which cells requires the enzyme. Investigating how GalC is handled may help find a cure for this devastating disease.
Neurology; Neurodegenerative disorders; Pathophysiology; Apoptosis and cell death; Cytoskeleton and cell motility; Molecular and Cellular Biology; Molecular genetics; Neurobiology; Protein Folding; Gene Expression; Transcription and Translation; Signal Transduction; Toxicology and Xenobiotics
My research is aimed at finding the cause and a cure for Parkinson’s disease. Parkinson’s disease (PD) is defined by a characteristic set of locomotor symptoms (rest tremor, rigidity, bradykinesia and postural instability) that are believed to be caused by the selective loss of dopaminergic (DA) neurons in substantia nigra. The persistent difficulties in using animals to model this human disease suggest that human nigral dopaminergic neurons have certain vulnerabilities that are unique to our species. One of our unique features is the large size of the human brain (1350 grams on average) relative to the body. A single nigral dopaminergic neuron in a rat brain (2 grams) has a massive axon arbor with a total length of 45 centimeters. Assuming that all mammalian species share a similar brain wiring plan, we can estimate (using the cube root of brain weight) that a single human nigral dopaminergic neuron may have an axon with gigantic arborization that totals 4 meters. Another unique feature of our species is our strictly bipedal movement, which is affected by Parkinson’s disease, in contrast to the quadrupedal movement of almost all other mammalian species. The much more unstable bipedal movement may require more dopamine, which supports the neural computation necessary for movement. The landmark discovery of human induced pluripotent stem cells (iPSC) made it possible to generate patient-specific human midbrain dopaminergic neurons to study Parkinson’s disease. A key problem for dopaminergic neurons is the duality of dopamine as a signal required for neural computation and a toxin as its oxidation produces free radicals. Our study using iPSC-derived midbrain dopaminergic neurons from PD patients with parkin mutations and normal subjects shows that parkin sustains this necessary duality by maintaining the precision of the signal while suppressing the toxicity. Mutations of parkin cause increased spontaneous release of dopamine and reduced dopamine uptake, thereby disrupting the precision of dopaminergic transmission. On the other hand, transcription of monoamine oxidase is greatly increased when parkin is mutated. This markedly increases dopamine oxidation and oxidative stress. These phenomena have not been seen in parkin knockout mice, suggesting the usefulness of parkin-deficient iPSC-derived midbrain DA neurons as a cellular model for Parkinson’s disease. Currently, we are using iPS cells and induced DA neurons to expand our studies on parkin to idiopathic Parkinson’s disease. We are also utilizing the molecular targets identified in our studies to find small-molecule compounds that can mimic the beneficial functions of parkin. The availability of human midbrain DA neurons should significantly speed up the discovery of a cure for Parkinson’s disease.
Bioinformatics; Genomics and proteomics; Immunology; Membrane Transport (Ion Transport); Molecular and Cellular Biology; Molecular Basis of Disease; Molecular genetics; Neurobiology; Gene Expression
The current focus of my lab is on iron metabolism in animals and humans. From the practical viewpoint, iron is an important nutrient, but its ability to act in the ferrous and ferric state also makes it toxic. Thus, iron deficiency is the most frequent disorder in the world and hereditary hemochromatosis (HH) is the most common Mendelian disorder in the United States. Our research is related to erythroid differentiation on the fundamental level and to genetic and acquired diseases on the applied level, with four long-term themes: 1.) analysis of the molecular basis of differential gene expression among tissues and during development, with hemoglobin synthesis and red blood cell (RBC) development as models; 2.) application of molecular and genetic advances to inherited diseases; 3.) iron metabolism; 4.) study of gene variation in populations and divergence of gene loci during evolution. New vistas have opened recently for the anemia of chronic diseases, leading us to re-exam how microbes and their human hosts fight for iron. We approach these issues by working on rodent models like the Belgrade rat, plus a series of genetically engineered mice. The rat has a hypochromic, microcytic anemia inherited as an autosomal recessive. The defect is in an iron transporter called DMT1 (or slc11a2, previously called Nramp2 or DCT1) that is responsible for iron uptake by enterocytes and is also responsible for iron exiting endosomes in the transferrin cycle. The rats appear to have a severe iron deficiency, and although dietary iron and iron injection increase the number of RBCs, they do not restore the RBCs nor the rat itself to a normal phenotype. Recent discoveries show that DMT1 is ubiquitous and responsible for transport of other metals such as Mn and Ni. It occurs in the kidney, brain and lung at even higher levels than in the GI tract or in erythroid cells. It also has multiple isoforms, and we have cloned them and developed cell lines that express high levels of particular isoforms. We have specific antibodies to the isoforms and assays for each of the mRNAs too. Future projects in my lab will continue to address whether DMT1 is dysregulated in HH. We will also tackle how DMT1 functions in neurons, pneumocytes and other tissues, look at isoforms of DMT1 under circumstances where we suspect that they must have different functions from one another, and examine DMT1’s relevance to iron metabolism and human disease. Because we cloned the gene and identified the mutation, a number of molecular and cellular approaches can now be used. As evidence indicates that metal ion homeostasis fails in Parkinson’s disease, Alzheimer’s disease and Huntington’s disease, research on DMT1 has opened new vistas for these disorders.
Bioinformatics; Cell growth, differentiation and development; Genomics and proteomics; Molecular and Cellular Biology; Molecular Basis of Disease; Molecular genetics; Neurobiology; Gene Expression; Stem Cells; Transgenic organisms
My research goal is to gain a better understanding of how proteins that interact with DNA regulate RNA transcription, DNA replication and metazoan development. I mentor undergraduate and graduate students in my lab; we focus on the structure and function of the Nuclear Factor I (NFI) family of site-specific DNA binding proteins, and we are investigating their roles in development. Our work has been made possible by our development of loss-of-function mutations of the NFI genes in the mouse and C. elegans. We are addressing four major questions in my laboratory and in collaboration with a number of talented collaborators: What is the structure of the NFI DNA-binding domain? How does NFI recognize and interact with DNA? Does NFI change the structure of DNA when it binds? What proteins interact with NFI to stimulate RNA transcription and/or DNA replication? These research questions are explored in my lab through two major projects focused on the role of NFIB in lung development and the role of NFIX in brain development. When NFIB is deleted from the germline of mice the animals die at birth because their lungs fail to mature normally. This provides a good model for the problems that occur with premature infants, whose lungs also fail to mature normally. We are using this model to determine how NFIB promotes lung maturation with the goal of being able to stimulate this process in premature infants. In our NFIX knockout animals, the brains of the animals are actually larger than normal and contain large numbers of cells in an area known to be the site of postnatal neurogenesis. We have evidence that NFIX may regulate the proliferation and differentiation of neural stem cells, which produce new neurons throughout adult life. Our aim is to understand the specific target genes that NFIX regulates in the adult brain to control this process of neurogenesis.
Bioinformatics; Cell growth, differentiation and development; Gene Expression; Genomics and proteomics; Molecular genetics; Signal Transduction
Research in my laboratory investigates the genetic regulatory circuitry that controls how cell fates are determined during development. We focus on two key aspects, intercellular signaling and transcriptional regulation, using primarily the fruit fly Drosophila melanogaster due to its extremely well-annotated genome and amenability to experimental manipulation. All conclusions, however, are expected to relate directly to mammalian (including human) gene regulation. Recently, we have also started investigating the regulatory genomics of other insect species of both medical and agricultural importance, beginning with the development of methods for regulatory element discovery in species with fully sequenced genomes but little functional, experimental data. A defining feature of my laboratory is that it takes both wet-lab and computational/bioinformatics approaches to studying the same set of problems about development and transcriptional regulation; hypotheses and ideas generated using one set of methods are tested and explored using the other. Current research in the laboratory falls into two main areas: 1) discovery and characterization of transcriptional cis-regulatory modules (CRMs), and 2) mechanisms of specificity for receptor tyrosine kinase (RTK) signaling. The combined results of these studies will provide insight into gene regulation, genome structure, intercellular signaling, and the regulatory networks that govern embryonic development. My group is also heavily involved in biocuration through our development and maintenance of REDfly, an internationally-recognized curated database of known Drosophila transcriptional cis-regulatory modules (CRMs) and transcription factor binding sites (TFBSs). Despite more than 25 years of experimental determination of these elements, the data have never been collected into a single searchable database. REDfly seeks to include all experimentally verified fly regulatory elements along with their DNA sequence, their associated genes, and the expression patterns they direct. REDfly is by far the most comprehensive database of regulatory elements for the higher eukaryotes and serves as an important resource for the fly and bioinformatics communities.
Gene Expression; Immunology; Molecular and Cellular Biology; Molecular genetics; Signal Transduction
I am the administrator for flow cytometry in the Confocal Microscopy and Flow Cytometry Core Facility that serves investigators throughout the university. In that role, I oversee the use of the LSRFortessa and the FACSCalibur analytical flow cytometers, providing instruction on their use and the analysis of acquired data and serving as a consultant on the design and interpretation of experiments. I also operate the FACSAria cell sorter, providing sterile live cell sorting. In addition, I operate and provide assistance to users on the application of cytometric bead array analysis on the FACSArray, as well as Elispot analysis using the Zeiss KS-ELISPOT microscope. My own research centers on investigating the responsiveness of human T cells in the tumor microenvironment of lung and ovarian cancer and lymphomas. In that research, I am a coinvestigator in a collaborative group of oncologists and immunologists coordinated by Richard Bankert, PhD. We have observed that T cells in the tumor microenvironment are hyporesponsive to T cell receptor-mediated activation and that factor(s) present in ovarian tumors and associated ascites fluid can cause this hyporesponsiveness. We are investigating the mechanism(s) of this phenomenon. Also, as an approach to the in vivo study of the immune response to human tumor associated antigens, our group has established a novel xenograft model by injecting human tumor cell aggregates of solid ovarian tumor biopsies intraperitoneally into immune-deficient NSG mice. The result is a human tumor microenvironment in the greater omentum of the mice, i.e., the omental tumor xenograft (OTX) model. The progression of the human tumor xenograft closely approximates the characteristics of the tumor in cancer patients, and it is possible to quantify the presence of tumor cells and stromal cells in the OTX model. These findings have led to our program goals to: 1.) determine whether the OTX model can be used as a predictive tool of the outcome of therapeutic approaches for the treatment of human ovarian cancer and B cell lymphoma, and 2.) determine whether the inhibition of activation in the tumor microenvironment can be reversed so that the antitumor T cell response can be reactivated.
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.
Bioinformatics; Cell growth, differentiation and development; Gene Expression; Genomics and proteomics; Molecular genetics; Stem Cells; Transcription and Translation; Transgenic organisms; Vision science
My lab is interested in how global gene expression advances from one state to the next in time and space during development to promote the specification and differentiation of individual retinal cell types from multi-potent neural progenitor cells. We focus on the gene regulatory network (GRN) involved in the formation of one retinal cell type, retinal ganglion cells (RGCs). RGCs are the only projection neurons in the retina and connect the retina to the brain through the optic nerve. Death of RGCs is cause of vision loss in glaucoma and other retinal diseases. Several key transcription factors (TFs) functioning at different stages of RGC development have been identified; Math5 is essential for RGC fate specification, whereas Pou4f2 and Isl1 are required for their differentiation. Our previous study has established a tentative model for the RGC GRN, in which these TFs occupy key node positions. Current projects in the lab are aimed at further understanding how these transcription factors specifically regulate their target genes and how they interact with each other. Considerable efforts are also placed on identifying novel key regulators in the GRN. Our studies employ a combined approach of genetics, genomics and bioinformatics. Our eventual goal is to use the knowledge learned from our studies to develop new therapies for various retinal diseases.
Gene Expression; Molecular Basis of Disease; Molecular and Cellular Biology; Molecular genetics; Protein Function and Structure; Transcription and Translation
Our laboratory utilizes combined genetic, biochemical and molecular biological approaches to investigate the molecular mechanisms involved in the initiation and regulation of eukaryotic transcription. Previous work in our laboratory utilizing both the budding yeast Saccharomyces cerevisiae and human cells resulted in the identification and biochemical characterization of mutants of nuclear RNA polymerase II (RNAPII) and the general transcription factors TFIIB and TFIIF that coordinately affect transcription start site utilization and transcript elongation. These studies supported a model where yeast and human TFIIF induce global conformational changes in RNAPII that result in structural and functional changes in the polymerase active center. Our current studies are focused on elucidating the mechanisms of kinetoplast transcription by the mitochondrial RNA polymerase of Trypanosoma brucei. T. brucei is a protozoan parasite that is the causative agent of African sleeping sickness (trypanosomiasis) in humans and nagana in animals. Procyclic trypanosomes growing in the midgut of the tsetse fly have a fully functional mitochondrion whereas trypanosomes in the mammalian bloodstream exhibit repressed mitochondrial function. The mitochondrial DNA in trypanosomes is unusual in its structure, comprising a highly catenated network of maxicircles and minicircles termed kinetoplast DNA (kDNA). Surprisingly, very little is known about the cis-acting elements and the trans-acting factors governing the transcription of maxicircles and minicircles. Our objective is to elucidate the mechanisms and regulation of T. brucei kDNA transcription with the ultimate goal of developing therapeutic agents.
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.
DNA Replication, Recombination and Repair; Genome Integrity; Molecular and Cellular Biology; Molecular genetics; Protein Function and Structure
In my laboratory, we are interested in the general problem of maintaining genome stability. To this end, we focus on two distinct aspects of genome stability: 1) the roles of mismatch (MMR) proteins in multiple pathways for DNA repair and 2) the manner in which regulation of dNTP pools, through the regulation of ribonucleotide reductase (RNR) activity, impacts genome integrity. 1) MMR proteins recognize many different types of DNA lesions and then target the lesion for the appropriate repair pathway. We are interested in the mechanism(s) by which recognition of a lesion is translated into the appropriate DNA repair pathway, using the yeast Saccharomyces cerevisiae as a model system. Is it through differential protein-nucleic acid or protein-protein interactions? To address these questions as well as the regulation of DNA repair pathway selection, we use a combination of genetic, biochemical and biophysical approaches. 2) RNR activity modulates the level of dNTPs that are available in a cell at a given time. Higher levels of dNTPs lead to higher mutation rates. We are interested in the various ways in which misregulated dNTP pools might affect cellular metabolism and affect the stability of the genome.