2005 Featured Speakers
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Louis M. Staudt, MD, Ph.D. - National Cancer Institute |
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Background Dr. Staudt received his B.A. from Harvard College in 1976 and his M.D. and Ph.D. from the University of Pennsylvania in 1982. His Ph.D. thesis in the laboratory of Walter Gerhard defined somatic hypermutation as a rapid mechanism of antibody diversification during normal immune responses. Following internal medicine training, he joined David Baltimore’s laboratory where he cloned and characterized the lymphoid-restricted transcription factor Oct-2. He established his laboratory in the Metabolism Branch, NCI, in 1988, and currently studies the molecular basis of human lymphoid malignancies. |
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Research
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Our laboratory studies the molecular pathogenesis of
human lymphoid malignancies and has three primary goals: to
establish a new molecular diagnosis of human lymphoid malignancies
using gene expression profiling, to elucidate the oncogenic pathways
that result in malignant transformation of normal B lymphocytes, and
to identify molecular targets for development of novel therapeutics
for these cancers. To provide a molecular basis for the diagnosis of human lymphoid malignancies, we are exploiting DNA microarray technology to profile gene expression in these cancers on a genomic scale. The laboratory created a novel DNA microarray, the "Lymphochip", which is enriched in genes that are expressed in and/or function in lymphocytes (1). We have used Lymphochip and Affymetrix microarrays to profile gene expression in diffuse large B cell lymphoma (DLBCL) (2-4), chronic lymphocytic leukemia (CLL) (5, 6), mantle cell lymphoma (7), follicular lymphoma (8), multiple myeloma (9), and in a wide variety of normal lymphoid subsets (2, 10-13). One central goal of these studies is to relate gene expression to clinical outcome, thereby establishing a quantitative, reproducible and informative molecular diagnosis of the lymphoid malignancies (14). Our studies have revealed previously unknown types of diffuse large B cell lymphoma that are indistinguishable by current diagnostic methods, but which have strikingly distinct gene expression profiles, originate from different stages of B cell differentiation, utilize distinct oncogenic mechanisms, and differ in their ability to be cured by current chemotherapy (2-4). For several lymphoid malignancies, we have identified molecular profiles that predict the length of survival or the ability to be cured by chemotherapy, thereby providing clinically useful prognostic indicators. Our laboratory has mounted a major effort to create a diagnostic microarray that could provide these molecular diagnoses and prognoses to patients with lymphoid malignancies.
Natalie G. Ahn, Ph.D. - Associate Professor, Department of Chemistry and Biochemistry, University Colorado at Boulder
Research A major goal of our research is to understand how phosphorylation controls cell signal transduction, by identifying protein kinases and phosphatases that are controlled by growth factors and examining their mechanisms of regulation. A second goal is to develop new techniques for analyzing post-translational modifications of proteins involved in signaling. An intracellular signal transduction pathway, called the MAP kinase cascade, is rapidly stimulated in response to growth factors. Enzymes in this pathway include pp90 ribosomal S6 kinase, mitogen-activated protein (MAP) kinase, and MAP kinase kinase (MKK, also referred to as MEK), which form three tiers of a protein kinase cascade in which pp90rsk is phosphorylated and activated by MAPK, and MAPK is phosphorylated and activated by MPKK. MPKK is phosphorylated and activated by any of three protein kinases, Raf-1, MEK kinase, and Mos, and thus is a convergence point for diverse signaling pathways triggered upon cell surface receptor activation. Several transcription factors are downstream targets for these kinases, thus, the MAP kinase cascade is a key pathway allowing transcriptional regulation by extracellular stimuli. Our aim is to understand the regulation of MKK and its role in cell growth and differentiation. We have identified several phosphorylation sites on this enzyme, and are examining their contributions to kinase activation. Mutants of MKK that are either constitutively active or catalytically inactive were designed, which, upon transfection into cultured mammalian cells respectively, enhance or block signal transduction through the MAP kinase pathway. Overexpression of constitutively active MKK mutants led to transformation in several types of cells. Interestingly, cell types respond instead by cell cycle arrest and differentiation. We are currently examining downstream cellular targets of MAP kinase and MKK to explain how such different cellular responses can be controlled by the same pathway. Electrospray ionization mass spectrometry (ESI-MS) enables determination of protein or nucleic acid masses to uncertainties of 1 in 10,000 Da. Experimental approaches we use in our laboratory include (i) liquid chromatography coupled to mass spectrometry (LC/MS), which separates molecules by high performance liquid chromatography followed by mass determination, (ii) tandem mass spectrometry (MS/MS, LC/MS/MS), which fragments ions by collision-induced dissociation, allowing peptide and nucleic acid sequences and specific sites of covalent modification to be determined, and (iii) deuterium exchange coupled to mass spectrometry, which measures solvent accessibility of protein backbone amides. LC/MS and LC/MS/MS are being used to identify phosphorylation sites and autophosphorylation sites on MKK, and other proteins. Deuterium exchange experiments are being combined with site directed mutagenesis to identify conformational differences between inactive vs. active forms of MKK and MAPK. We are also using mass spectrometry to examine masses of components within large protein/nucleic acid complexes with the aim of identifying novel protein post-translational modifications under growth factor control.
Robert M. Nerem, Ph.D.- Director of the Parker H. Petit Institute for Bioengineering & Bioscience
Background
Research Dr. Nerem's current research projects include the development of a blood vessel substitute for use in bypass surgery, the investigation of hemodynamics as a regulator of vascular biology, and the mathematical modeling of the dynamic response of mammalian cells. The focus of Dr. Nerem's laboratory is cellular and tissue engineering as applied to the vascular system. The projects range from understanding the role of flow and the associated shear stress on vascular endothelial biology, the influence of cyclic stretch on smooth muscle cell biology, and the tissue engineering of blood vessel substitutes. The work of the laboratory extends from the study of actin dynamics to gene expression. Of particular interest is nitric oxide and various vascular cell adhesion molecules. This tissue engineering effort uses an approach that incorporates a smooth muscle cell seeded collagen gel which has been plated with an endothelial cell interface. Both a slab version of this model of a blood vessel wall and a tubular construct version exist. The facilities for this work are rather extensive. Important is the ability to do in vitro flow and cyclic stretch experiments. The laboratory includes a basic cell culture facility, a fluorescence microscope room, instrumentation for 2D electrophoresis and for horizontal and vertical electrophoresis using agarose and SDS PAGE gels, northern and western blotting, an ultracentrifuge for cell fractionation, a spectrophotometer, and the necessary instrumentation to do radioisotope measurements. A special facility exists for making time resolved intracellular calcium measurements. Sponsors of Dr. Nerem's work are the National Science Foundation, the National Institutes of Health, the Whitaker Foundation, the National Aeronautics and Space Administration, and Advanced Tissue Sciences. Mike Ehlers, M.D, Ph.D. - Associate Professor, Department of Neurobiology, Duke University Medical Center
Background Dr. Ehlers completed his undergraduate work at the California Institute of Technology in 1991 and received his M.D. and Ph.D. degrees from the Johns Hopkins University in 1998. His thesis research focused on localization and calcium control of NMDA-type glutamate receptors in the lab of Richard Huganir. Presently, Dr. Ehlers is an associate professor in the Department of Neurobiology at the Duke University Medical Center. Research Neuronal signaling relies on the precise spatial arrangement of receptors and ion channels at specialized membrane sites over the cell body, axon, and dendrites. This arrangement requires selective localization or stabilization of receptors in the neuronal plasma membrane, as well as the highly coordinated intracellular trafficking of receptors and channels through the secretory and endocytic pathways. Despite significant progress in our understanding of membrane trafficking in general, and the role of endocytosis in synaptic vesicle biogenesis in the presynaptic nerve terminal, very little is known about secretory and endocytic trafficking in neuronal dendrites or the contribution of protein degradation to synapse function. The research in the Ehlers Lab is directed at understanding protein trafficking and turnover in dendrites and its relationship to synapse formation and function. The complex morphology of the neuron, with its elaborately branched dendrites onto which impinge hundreds to thousands of individual synapses, requires that highly specialized mechanisms exist for localizing, maintaining, and removing proteins at the synapse. Such mechanisms are crucial for the initial establishment of postsynaptic specializations during synaptogenesis, and for activity-dependent changes in synaptic strength that underlie experience-dependent plasticity. Moreover, alterations in the trafficking or stabilization of neurotransmitter receptors at the synapse may contribute to pathologic changes in neurologic and psychiatric disease.
John D. Scott, Ph.D. - Senior Scientist - The Vollum Institute at Oregon Health Sciences University
Background John D. Scott is a senior scientist of the Vollum Institute at Oregon Health Sciences University and an associate investigator of the Howard Hughes Medical Institute. He received his Ph.D. in Biochemistry from the University of Aberdeen and worked as a postdoctoral fellow at the University of Washington. He held positions at the University of Washington and the University of California at Irvine before he was appointed to the faculty of Oregon Health Sciences University. Research John Scott and his associates use recombinant DNA techniques, protein chemistry, and enzymology to study the actions of protein kinase A (PKA) within cells. The biochemical effects of many peptide hormones proceed through pathways that lead to activation of PKA. However, individual hormones may promote PKA-mediated phosphorylation of distinct sets of proteins. This may be because different hormones activate different subtypes of the PKA enzyme. Alternatively, individual hormones may activate specific pools of PKA. A potential mechanism to explain this phenomenon is that individual PKA pools might be compartmentalized inside the cell at their site of action, close to the proteins that they will ultimately phosphorylate. A specific pool could be activated only when the appropriate hormone elevates cAMP in a particular microenvironment. Scott's laboratory has shown that Type II PKA is tethered at particular subcellular locations by specific A-kinase anchoring proteins (AKAPs). Scott and his group have demonstrated that the AKAPs are a diverse family of functionally related proteins. So far, more than 30 AKAP genes have been isolated by interaction cloning techniques. Each AKAP contains a conserved amphipathic helix motif of 20 or so residues that is responsible for high affinity interaction with the regulatory subunit of PKA. Each anchoring protein has a unique targeting site that is responsible for association with membranes or subcellular structures. These targeting domains confer specificity on each AKAP as they direct the localization of the kinase to the cytoskeleton, nuclear matrix, endoplasmic reticulum, peroxisomes, and cell membranes. One anchoring protein called AKAP79 is targeted to the nerve terminals, and experiments have shown that disruption of PKA/AKAP79 impairs the transfer of excitatory neuronal impulses from one nerve to the next. Additional studies have shown that AKAP79 also anchors two other signal transduction enzymes that participate in the transmission of neuronal impulses, the phosphatase calcineurin and protein kinase C. These observations suggest that anchoring proteins may maintain groups of enzymes involved in a particular physiological response close to their substrates. Scott and his laboratory are now working on the hypothesis that selective activation of kinases and phosphatases anchored close to their site of action may determine the specificity of intracellular signal transduction events.
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