Mimnaugh E G, Worland P J, Whitesell L, Neckers L M
Clinical Pharmacology Branch, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Biol Chem. 1995 Dec 1;270(48):28654-9. doi: 10.1074/jbc.270.48.28654.
The abundant, cytoplasmic 90-kDa heat-shock protein associates transiently with the Rous sarcoma virus oncogenic protein tyrosine kinase, pp60v-src, directs its cellular trafficking and negatively regulates its kinase activity. Here we report that the serine/threonine phosphatase inhibitor, okadaic acid, destabilized the heat-shock protein 90-pp60v-src chaperone complex in v-src-transfected cells. Concomitant with complex destabilization by okadaic acid, phosphoserine was doubled and phosphothreonine was increased 20-fold in the heat-shock protein 90. Although phosphorylation of the total pool of immunoprecipitable pp60v-src was unchanged, okadaic acid slightly increased phosphoserine and phosphothreonine levels specifically in pp60v-src bound to heat-shock protein 90. The low level of tyrosine phosphorylation in the pp60v-src complexed with heat-shock protein 90 was further decreased by okadaic acid. Interestingly, okadaic acid-stabilized hyperphosphorylation of the heat-shock protein 90-pp60v-src complex lowered the level of pp60v-src in cell membranes, the functional location for pp60v-src. We suggest that serine/threonine phosphorylation of heat-shock protein 90 and/or pp60v-src functions as a regulatory molecular trigger to release pp60v-src from the chaperone complex at the inner surface of cell membranes.
丰富的细胞质90-kDa热休克蛋白与劳氏肉瘤病毒致癌蛋白酪氨酸激酶pp60v-src短暂结合，指导其细胞内运输并负向调节其激酶活性。我们在此报告，丝氨酸/苏氨酸磷酸酶抑制剂冈田酸使v-src转染细胞中的热休克蛋白90-pp60v-src伴侣复合物不稳定。与冈田酸导致的复合物不稳定同时发生的是，热休克蛋白90中的磷酸丝氨酸增加了一倍，磷酸苏氨酸增加了20倍。尽管可免疫沉淀的pp60v-src总池的磷酸化没有变化，但冈田酸特异性地略微增加了与热休克蛋白90结合的pp60v-src中的磷酸丝氨酸和磷酸苏氨酸水平。与热休克蛋白90复合的pp60v-src中的低水平酪氨酸磷酸化被冈田酸进一步降低。有趣的是，冈田酸稳定的热休克蛋白90-pp60v-src复合物的过度磷酸化降低了细胞膜中pp60v-src的水平，而细胞膜是pp60v-src的功能定位。我们认为，热休克蛋白90和/或pp60v-src的丝氨酸/苏氨酸磷酸化作为一种调节分子触发因素，在细胞膜内表面将pp60v-src从伴侣复合物中释放出来。
