Abstract
We have developed a generalized approach, using two hybrid interactions, to isolate Ha-Ras effector loop mutations that separate the ability of Ha-Ras to interact with different downstream effectors. These mutations attenuate or eliminate Ha-ras(G12V) transformation of mammalian cells, but retain complementary activity, as demonstrated by synergistic induction of foci of growth-transformed cells, and by the ability to activate different downstream components. The transformation defect of Ha-ras(G12V, E37G) is rescued by a mutant, raf1, that restores interaction. These results indicate that multiple cellular components, including Raf1, are activated by Ha-Ras and contribute to Ha-Ras-induced mammalian cell transformation.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Cell Transformation, Neoplastic / genetics*
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Cells, Cultured
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Chloramphenicol O-Acetyltransferase / analysis
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Gene Library
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Genes, ras / genetics*
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Genetic Complementation Test
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Humans
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Mice
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Mutation
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Protein Kinases / analysis
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Protein Serine-Threonine Kinases / genetics*
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Proto-Oncogene Proteins / genetics*
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Proto-Oncogene Proteins c-raf
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Rats
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Recombinant Fusion Proteins / biosynthesis
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Saccharomyces cerevisiae / genetics
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Schizosaccharomyces / genetics
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Transfection
Substances
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Proto-Oncogene Proteins
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Recombinant Fusion Proteins
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Chloramphenicol O-Acetyltransferase
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Protein Kinases
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-raf