The function of the ras family of oncogene proteins will be investigated utilizing both living cell systems and in vitro biochemical approaches. The human H-ras proto- and oncogene proteins have been produced in an E. coli expression system, and the proteins have been purified to homogeneity and characterized as """"""""native"""""""" entities. We propose to use these intact proteins as antigens for the production of a bank of mouse-derived monoclonal antibodies, which will be subsequently employed in both biochemical experiments and microinjection experiments with normal and transformed cells. As we have recently shown, microinjection of the human H-ras oncogene protein, but not the proto-oncogene protein, into quiescent normal cells results in rapid morphological changes followed by DNA synthesis and proliferation. We wish to follow up these initial experiments by examining this unique assay of the transformation process induced by human oncogene proteins in great detail. And, through the use of the purified human H-ras proteins, we intend to look at the interactions of other cellular proteins with the ras proteins in vitro. (X)
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