Biological and biochemical activity of v-Crk chimeras containing the SH2/SH3 regions of phosphatidylinositol-specific phospholipase C-gamma and Src.
The chicken CT10 virus oncogene product, P47gag-crk, contains SH2/SH3 domains that have been identified as conserved domains among proteins involved in signal transduction. We studied the functional similarity of the SH2/SH3 domains by replacing those of v-Crk with those of phosphatidylinositol-specific phospholipase C-gamma, v-Src, or c-Src. The transforming activity of v-Crk was partially retained in a mutant with a v-Src SH3 domain but not in the other mutants with heterologous SH2/SH3 domains. Mutant viruses with Crk-SH2/SH2' domains induced tyrosine phosphorylation of cellular proteins, but mutants with phosphatidylinositol-specific phospholipase C-gamma or Src SH2/SH2' domains did not. However, the mutant proteins with heterologous SH2/SH2' regions were able to weakly associate with some phosphotyrosine-containing proteins in vitro. These results indicate that in the context of the P47gag-crk structure, the requirement of Crk-SH2/SH3 is more stringent for its activity to induce cell transformation than to cause phosphorylation of cellular proteins. The substitution with heterologous sequences least perturbs the capacity to bind phosphotyrosine-containing proteins. In each case, the SH3 domain is more flexible to substitution than is the SH2 domain.
Rockefeller University, New York, New York 10021-6399.
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Matsuda
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Reichman
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Hanafusa
D000818:Animals; D001483:Base Sequence; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002642:Chick Embryo; D002678:Chimera; D003001:Cloning, Molecular; D004279:DNA, Viral; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D009154:Mutation; D015688:Oncogene Protein pp60(v-src) / Q000235:genetics / Q000378:metabolism*; D050736:Oncogene Protein v-crk; D010716:Phosphatidylinositols / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics / Q000378:metabolism*; D012191:Retroviridae Proteins / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D010738:Type C Phospholipases / Q000235:genetics / Q000378:metabolism*; D014780:Viruses / Q000235:genetics
D004279:DNA, Viral; D050736:Oncogene Protein v-crk; D010716:Phosphatidylinositols; D011518:Proto-Oncogene Proteins; D012191:Retroviridae Proteins; C060731:p47 (gag-crk) protein, Avian sarcoma virus; D015688:Oncogene Protein pp60(v-src); D016392:Proto-Oncogene Proteins pp60(c-src); D010738:Type C Phospholipases