DIMERIZATION OF CDC25P, THE GUANINE-NUCLEOTIDE EXCHANGE FACTOR FOR RAS FROM SACCHAROMYCES-CEREVISIAE, AND ITS INTERACTION WITH SDC25P

The oligomerization state of Cdc25p, the guanine nucleotide exchange f actor for ras from yeast, was analyzed using different complementary a pproaches. The two-hybrid system showed that the C-terminal part of Cd c25p (Cdc25-Ct) can interact with itself but also with Sdc25p-Ct, the corresponding part of Sdc25p, the other guanine exchange factor from y east. The homotropic interaction of Cdc25p-Ct has been confirmed in ye ast using immunoprecipitation experiments with epitope-tagged and beta -galactosidase-fused polypeptides. No other component was required for this interaction, since dimerization was shown to occur with material synthesized in vitro. The size of Cdc25-Ct produced in Escherichia co li has been directly measured on gel filtration columns and correspond s to a dimer. The dimerization domain is localized in the same part of the molecule as the catalytic domain and the portion responsible for membrane localization. The biological relevance of dimerization is sti ll an open question, however by allowing heterodimerization with Sdc25 p it could permit a more complex combinatorial regulation of ras in ye ast.