MUTAGENIC ANALYSIS OF FUNCTIONAL DOMAINS OF THE MOS PROTOONCOGENE ANDIDENTIFICATION OF THE SITES IMPORTANT FOR MAPK ACTIVATION AND DNA-BINDING

We constructed in-frame deletion/replacement mutations in the Xenopus mos proto-oncogene that lie within conserved Mos-specific codons, but outside of the regions that are conserved among the src kinase family of genes, All gene products were assayed in vine for kinase activity a nd in vivo for their ability to induce oocyte maturation, embryonic cl eavage arrest and cellular transformation, Most mutations in Mos elimi nated both kinase and biological activity, However, a mutation in Mos that removed two basic amino acid residues (R94 and K97) downstream fr om the lysine at the ATP binding site (K90) markedly enhanced autophos phorylation activity, Moreover, this mutant displayed markedly reduced biological activity, lacked transforming activity, and failed to acti vate mitogen activated protein kinase (MAPK), A second mutant Mos prod uct, lacking amino acids R45-A54, displayed a five-fold increase in ce llular transforming activity, This Mos mutant specifically localized t o the cytoplasm; in contrast to wild-type (wt) Mos that localized to b oth the nucleus and the cytoplasm, These data indicate that Mos transf orming activity is mediated via signalling exerted in the cytoplasm, p resumably through MAPK, and that nuclear localization of the oncogene product interferes with transforming activity, We also show that amino acids R45-A54 are important for Mos DNA binding activity.