Fukui Y, Kornbluth S, Jong S M, Wang L H, Hanafusa H
Rockefeller University, New York, New York 10021.
Oncogene Res. 1989;4(4):283-92.
We have assayed immunoprecipitates of several oncogene products from retrovirally infected chicken embryo fibroblasts (CEF) for phosphatidylinositol (PI) kinase activity. Immunoprecipitates of P68gag-ros, P130gag-tps, P47gag-crk, polyoma middle T (mT)-p60c-src complex, and mT-p62c-yrs complex exhibited PI kinase activity when assayed without detergents. This activity was sensitive to the nonionic detergent, Triton X-100, and the product was indistinguishable from phosphatidylinositol-3-phosphate, the product of kinase type I. Immunoprecipitates of p21Ha-ros protein did not contain any PI kinase type I activity. It has been suggested that an 81 kD protein phosphorylated in in vitro kinase assays of immunoprecipitates from mT-transformed rodent cells is responsible for the PI kinase type I activity seen in these immunoprecipitates. We have detected a chicken homologue of this 81 kD protein in immunoprecipitates of lysates from mT-transformed CEF. However, the chicken 81 kD protein sedimented more quickly than the PI kinase activity in sucrose gradients. In addition, the 81 kD protein was not detectable in protein kinase assays of immunoprecipitates of P68gag-ros or P130gag-fps. These results suggest that the 81 kD protein may not be the PI kinase.
我们检测了来自逆转录病毒感染的鸡胚成纤维细胞（CEF）的几种癌基因产物的免疫沉淀物的磷脂酰肌醇（PI）激酶活性。在不使用去污剂进行检测时，P68gag-ros、P130gag-tps、P47gag-crk、多瘤病毒中T抗原（mT）-p60c-src复合物以及mT-p62c-yrs复合物的免疫沉淀物表现出PI激酶活性。这种活性对非离子去污剂Triton X-100敏感，且产物与I型激酶的产物磷脂酰肌醇-3-磷酸无法区分。p21Ha-ros蛋白的免疫沉淀物不包含任何I型PI激酶活性。有人提出，在mT转化的啮齿动物细胞免疫沉淀物的体外激酶检测中被磷酸化的一种81kD蛋白，是这些免疫沉淀物中所见I型PI激酶活性的原因。我们在mT转化的CEF裂解物的免疫沉淀物中检测到了这种81kD蛋白的鸡同源物。然而，在蔗糖梯度中，鸡81kD蛋白的沉降速度比PI激酶活性更快。此外，在P68gag-ros或P130gag-fps免疫沉淀物的蛋白激酶检测中未检测到81kD蛋白。这些结果表明，81kD蛋白可能不是PI激酶。
