Biological activity of human N-ras and K-ras genes containing the Asn17 dominant negative mutation.
Substitution of asparagine for serine at position 17 of human H-ras results in an impaired GTP-binding activity, causing the mutant Ras protein to be locked in a constitutively inactive GDP-bound state. Expression of this mutant in NIH 3T3 cells inhibits cell proliferation by blocking endogenous ras function. Plasmids that encode the analogous dominant negative mutation at position 17 in human N- and K-ras were constructed. These mutant ras genes, driven by a heavy metal-inducible sheep metallothionein promoter, were introduced by transfection into a variety of animal cell lines. All three mutant ras genes displayed an inhibitory phenotype when expressed in NIH 3T3 cells. This inhibition could be overcome by cotransfection with either activated H-ras or v-raf. These data indicate that the three human Ras proteins probably act through the same signal transduction pathway in NIH 3T3 cells and suggest that these mutations may confer similar phenotypes to other GTP/GDP-binding proteins.
Kaplan
J B
JB
Oncology and Immunology Research Department, Lederle Laboratories, Pearl River, NY 10965, USA.
D016475:3T3 Cells; D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine; D002461:Cell Line, Transformed; D003062:Codon / Q000235:genetics*; D003114:Colony-Forming Units Assay; D005799:Genes, Dominant; D011905:Genes, ras; D005816:Genetic Complementation Test; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D027384:Oncogene Proteins v-raf; D010957:Plasmids; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D016283:Proto-Oncogene Proteins p21(ras) / Q000145:classification / Q000235:genetics / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics / Q000502:physiology; D012694:Serine; D015398:Signal Transduction; D014162:Transfection
D003062:Codon; D011993:Recombinant Fusion Proteins; D015682:Retroviridae Proteins, Oncogenic; D012694:Serine; D001216:Asparagine; D027384:Oncogene Proteins v-raf; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)