Williger B T, Liscovitch M
Department of Biological Regulation, Weizmann Institute of Science, Rehovot, Israel.
FEBS Lett. 1997 Feb 10;403(1):35-9. doi: 10.1016/s0014-5793(97)00027-6.
Oncogenic transformation by v-Src is accompanied by marked morphological changes and cytoskeletal reorganization. Yet, the cytoskeleton-associated proteins with which v-Src interacts are largely unknown. We have studied the binding of v-Src-SH3 domain to cellular proteins utilizing a blot overlay procedure with a GST-v-Src-SH3 fusion protein as probe. A major 62-64 kDa v-Src-SH3-binding protein, present in detergent-insoluble cellular fractions, was identified as p21ras-GTPase-activating protein-associated p62 (GAPA62). In non-transformed cells, including NIH 3T3 cells, GAPA62 was present in both the RIP A-soluble and RIP A-insoluble fractions, but only the latter form was tyrosine-phosphorylated. In contrast, in polyoma middle T antigen-transformed 3T3 cells, GAPA62 was present only in the RIP A-insoluble fraction, where it was highly phosphorylated. It is suggested that tyrosine phosphorylation of GAPA62 may be an important determinant of its cellular localization and its possible function as a mediator of v-Src actions.
v-Src介导的致癌转化伴随着显著的形态变化和细胞骨架重组。然而，与v-Src相互作用的细胞骨架相关蛋白在很大程度上仍不清楚。我们利用以GST-v-Src-SH3融合蛋白为探针的印迹覆盖法研究了v-Src-SH3结构域与细胞蛋白的结合。一种主要的62-64 kDa的v-Src-SH3结合蛋白存在于去污剂不溶性细胞组分中，被鉴定为p21ras-鸟苷三磷酸酶激活蛋白相关p62（GAPA62）。在包括NIH 3T3细胞在内的未转化细胞中，GAPA62存在于RIPA可溶性和RIPA不溶性组分中，但只有后者形式发生酪氨酸磷酸化。相比之下，在多瘤病毒中T抗原转化的3T3细胞中，GAPA62仅存在于RIPA不溶性组分中，且在该组分中高度磷酸化。提示GAPA62的酪氨酸磷酸化可能是其细胞定位的重要决定因素，以及其作为v-Src作用介质的可能功能。
