The regulation of cell division in mammalian cells is a complex process; coordinating growth promoting signals transduced across the cytoplasmic membrane initiated by growth factors, with signals initiated by elements monitoring the nutritional status of the cells. The discovery of cellular oncogenes provide a means to study, at least in part, the biochemical mechanisms controlling cell division. One group of these oncogenes, the ras genes, are found to be mutationally activated in a large number and wide variety of human tumors. The ras genes are highly conserved in evolution, suggesting they play a fundamental role in the biology of the cell. Both the function of the ras proteins and how the ras proteins are regulated remains a mystery for mammalian cells. The discovery of the yeast RAS genes made possible a systematic approach to study RAS protein function because of the genetic manipulation possible in that organism. This proposal is aimed at studying the CDC25 proteins in yeast which can regulate, not only yeast RAS protein function but the human H-ras protein as well. This regulation is likely due to a direct protein-protein interaction between CDC25 and RAS proteins. The study of this protein-protein interaction and its conservation t rough evolution provides a unique means to identify the mammalian molecule which regulates ras protein function. We will develop DNA probes and immunological reagents which encode or recognize the domain of the CDC25 protein essential for RAS activation. These reagents will be used to clone cDNAs encoding domains homologous to CDC25. Further, we will use a genetic screen to identify human cDNAs which can complement loss of CDC25 function in yeast. The identification of the molecular mechanism of action of the human ras proteins may allow the design of molecularly rational drugs to reverse the tumorigenic effect of mutant ras proteins.
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