SITE-DIRECTED MUTAGENESIS OF RAS ONCOGENES

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We have studied the properties of mutant v-ras-H genes to determine the effects of these mutations on their transformation potential. The expression of fully transforming v-ras-H genes correlates with a reduction in the level of MHC expression on the surface of NIH3T3 cells. Certain mutant transformed cells show relatively little MHC reduction, and these mutants show reduced tumorigenicity in immunocompetent mice and fail to form metastases. We have shown that certain non-transforming mutants, which encode a p2l product exhibiting reduced GTP binding, are able to reduce the focus-forming ability of wt v-ras-H and other oncogenes in cotransfection experiments. We have isolated a series of flat revertants (RT) following transfection of the 116Y mutant ras plasmid into v-ras-H transformed 3T3 cells. RT cells express v-ras-H and, in comparison to the parental transformed cells, grow poorly in serum-free or reduced serum media, fail to form tumors in immunocompetent mice and express increased levels of MHC class I antigen on their surfaces. They form tumors in nude mice at reduced rates and grow to higher densities in culture than non-transformed 3T3 cells. RT cells are also resistant to retransformation by bat ras and mos, suggesting that they represent a novel class of revertants.