The broad, long-term objectives of this project are to characterize the structure, modes of regulation and physiological function of members of the protein tyrosine phosphatase (PTP) family of enzymes. The reversible phosphorylation of tyrosyl residues in proteins, catalyzed by the coordinated actions of protein tyrosine kinases (PTKs) and phosphatases (PTPs), is of critical importance to the regulation of signaling events that underlie such fundamental processes as growth and proliferation, migration differentiation, metabolism and cytoskeletal function. Disturbance of this delicate balance between the PTKS and PTPs has been shown to be a cause of human diseases including cancer, diabetes and inflammation. Clearly a characterization of the PTPs will be a prerequisite to achieving a complete understanding of the physiological consequences of tyrosine phosphorylation and how this process is abrogated in the pathogenesis of human disease. In this competitive renewal, the research program is focused primarily on two non-transmembrane PTPs that have been shown to exert selective effects on signal transduction in vivo - PTP1B, which has been implicated in the regulation of insulin receptor signaling, and TCPTP, which regulates EGF-induced signal transduction. Detailed structure-function analyses will be applied to examine at the molecular level the mechanism of substrate recognition, catalysis and regulation of these enzymes. In addition, substrate trapping mutant forms of these enzymes, a powerful technology developed as part of this research program, will be utilized to identify their physiological substrates, thus yielding critical insights into function. It has been noted that expression of PTP 18 is induced at the transcriptional level in several disease contexts. Therefore, a characterization of its promoter is proposed. Finally, in this post-genomic era of biological research, the proposal is made to develop strategies for profiling PTPs present in defined cell types and subcellular compartments, with the long-term goal of establishing novel links between members of the PTP family and human disease states. In view of these objectives, the Specific Aims are: 1) To conduct a structure-function analysis of PTP1B that will define mechanisms of substrate recognition and regulation of enzymatic activity, 2) To characterize the PTP1B promoter and elucidate mechanisms by which expression of the PTP is altered in human disease, 3) To characterize the regulation and function of TCPTP and 4) To develop proteomics-based strategies for profiling the expression of members of the PTP family.
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