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首页> 外文期刊>Molecular and Cellular Biology >Suppression of src transformation by overexpression of full-length GTPase-activating protein (GAP) or of the GAP C terminus.
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Suppression of src transformation by overexpression of full-length GTPase-activating protein (GAP) or of the GAP C terminus.

机译：通过全长GTPase激活蛋白（GAP）或GAP C末端的过表达来抑制src转化。

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Overexpression of the full-length GTPase-activating protein (GAP) has recently been shown to suppress c-ras transformation of NIH 3T3 cells but not v-ras transformation (36). Here, we show that focus formation induced by c-src was inhibited by approximately 80% when cotransfected with a plasmid encoding full-length GAP. In a similar assay, focus formation by the activated c-src (Tyr-527 to Phe) gene was inhibited by 33%. Cotransfection of the GAP C terminus coding sequences (which encode the GTPase-accelerating domain) with c-src or c-src527F inhibited transformation more efficiently than did the full-length GAP, while the GAP N terminus coding sequences had no effect on src transformation. When cells transformed by c-ras, c-src, c-src527F, or v-src were transfected with GAP or the GAP C terminus sequence in the presence of a selectable marker, 40 to 85% of the resistant colonies were found to be morphologically revertant. The GAP C terminus induced reversion of each src-transformed cell line more efficiently than the full-length GAP, but this was not the case for reversion of c-ras transformation. Biochemical analysis of v-src revertant subclones showed that the reversion correlated with overexpression of full-length GAP or the GAP C terminus. There was no decrease in the level of pp60src expression or the level of protein-tyrosine phosphorylation in vivo. We conclude that GAP can suppress transformation by src via inhibition of endogenous ras activity, without inhibiting in vivo tyrosine phosphorylation of cellular proteins induced by pp60src, and that src may negatively regulate GAP's inhibitory action on endogenous ras.
机译：全长GTPase激活蛋白（GAP）的过表达最近已显示可抑制NIH 3T3细胞的c-ras转化，但不能抑制v-ras转化（36）。在这里，我们显示了当与编码全长GAP的质粒共转染时，由c-src诱导的焦点形成受到大约80％的抑制。在类似的分析中，由激活的c-src（Tyr-527至Phe）基因形成的焦点被抑制了33％。 GAP C末端编码序列（编码GTPase加速域）与c-src或c-src527F的共转染比全长GAP更有效地抑制了转化，而GAP N末端编码序列对src转化没有影响。当在选择标记存在下用GAP或GAP C末端序列转染c-ras，c-src，c-src527F或v-src转化的细胞时，发现40％至85％的抗性菌落是形态可逆的。与全长GAP相比，GAP C末端更有效地诱导了每个src转化的细胞系的回复，但c-ras转化的回复并非如此。 v-src回复子克隆的生化分析表明，该回复与全长GAP或GAP C末端的过表达相关。体内pp60src表达水平或蛋白酪氨酸磷酸化水平均未降低。我们得出的结论是，GAP可以通过抑制内源性ras活性抑制src的转化，而不会抑制pp60src诱导的细胞蛋白的体内酪氨酸磷酸化，并且src可能负面调节GAP对内源性ras的抑制作用。

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