INTERACTION OF ACTIVATED RAS WITH RAF-1 ALONE MAY BE SUFFICIENT FOR TRANSFORMATION OF RAT2 CELLS

v-H-ras effector mutants have been assessed for transforming activity and for the ability of the encoded proteins to interact with Raf-1-, B -Raf-, byr2-, ralGDS-, and CDC25-encoded proteins in the yeast two-hyb rid system. Transformation was assessed in rat2 cells as well as in a mutant cell line, rv68BUR, that affords a more sensitive transformatio n assay. Selected mutant Ras proteins were also examined for their abi lity to interact with an amino-terminal fragment of Raf-1 in vitro. Fi nally, possible cooperation between different v-H-ras effector mutants and between effector mutants and overexpressed Raf-1 was assessed. Ra s transforming activity was shown to correlate best with the ability o f the encoded protein to interact with Raf-1. No evidence for cooperat ion between v-H-ras effector mutants was found. Signaling through the Raf1-MEK-mitogen-activated protein kinase cascade may be the only effe ctor pathway contributing to RAS transformation in these cells.