MAMMALIAN RASGAP RELATED GENES

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This project proposes to explore the biological functions of two newly discovered Ras GTPase activating protein (RasGAP)-related human proteins, called IQGAP1&2, which may play roles in human disease and which may present novel targets for therapy. Both novel proteins share extensive similarity in their C-terminal halves with the sar1/gap1 putative RasGAP from Schizosaccharomyces pombe and may be homologs of this fission yeast Ras regulator. Further implicating IQGAPs as Ras regulators, one of these proteins resembled neurofibromin in its ability to suppress the growth of a human H=Ras dependent yeast strain. In addition, it was surprising to find that both proteins harbor IQ motifs and form stable complexes with calmodulin. Since IQGAP2 is uniquely expressed in liver and hepatocyte cell line, whereas IQGAP1 mRNA levels are especially high in placenta, lung, kidney, and peripheral blood leukocytes, these proteins may thus perform novel roles as perhaps epthelial cell specific integrators of the signaling pathways mediated by Ras and Ca2+/calmodulin. However, while in vitro experiments have suggested Ras binding, in preliminary studies to stimulation of the GTPase of Ras or its immediate relatives has yet been detected, indicating either that the activities of IQGAP1&2 are tightly regulated by Ca2+/calmodulin, or that these proteins are Ras interactors without prominent GAP activity. Since the central role of Ras and its regulators in mitogenic signal transduction is reflected by a high frequency of oncogenic Ras pathway mutations, a combination biochemical, expression, cell biological and genetic studies is proposed to further explore the biological properties and potential tumor suppressor roles of these intriguing new proteins. Specifically, this project includes biochemical studies to define the potentially calimodulin-regulated GTPase modulating activities of IQGAP1&2, expression studies to determine the exact tissues and cell types in which these proteins perform their function, cell biological analyses to test whether IQGAP1&2 form transient protein complexes in stimulated cells, as is suggested by the unexpected observation of a calcium ionophore-dependent nuclear translocation, and genetic studies to explore potential tumor suppressor roles. Beyond the fact that NF1 and p120GAP are both mutated in specific cancers, the latter studies are also proposed because IQGAP1 and IQGAP2 are heterogenously expressed in lung and liver cancer cell lines, because loss-of- heterozygosity has been detected with IQGAP1 probes in lung tumors, and because the IQGAP1 gene maps to a region involved in a recurring lung adenocarcinoma chromosome translocation. In addition to providing clues to the biological functions of these intriguing new protein, these studies may also contribute to a better understanding of other RasGAPs and provide a structural basis for often suspected links between Ras and calmodulin- mediated signal transduction.