Burridge K, Turner C E, Romer L H
Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill 27599.
J Cell Biol. 1992 Nov;119(4):893-903. doi: 10.1083/jcb.119.4.893.
Cells in culture reveal high levels of protein tyrosine phosphorylation in their focal adhesions, the regions where cells adhere to the underlying substratum. We have examined the tyrosine phosphorylation of proteins in response to plating cells on extracellular matrix substrata. Rat embryo fibroblasts, mouse Balb/c 3T3, and NIH 3T3 cells plated on fibronectin-coated surfaces revealed elevated phosphotyrosine levels in a cluster of proteins between 115 and 130 kD. This increase in tyrosine phosphorylation was also seen when rat embryo fibroblasts were plated on laminin or vitronectin, but not on polylysine or on uncoated plastic. Integrin mediation of this effect was suggested by finding the same pattern of elevated tyrosine phosphorylation in cells plated on the cell-binding fragment of fibronectin and in cells plated on a synthetic polymer containing multiple RGD sequences. We have identified one of the proteins of the 115-130-kD cluster as pp125FAK, a tyrosine kinase recently localized in focal adhesions (Schaller, M. D., C. A. Borgman, B. S. Cobb, R. R. Vines, A. B. Reynolds, and J. T. Parsons. 1992. Proc. Natl. Acad. Sci. USA. 89:5192). A second protein that becomes tyrosine phosphorylated in response to extracellular matrix adhesion is identified as paxillin, a 70-kD protein previously localized to focal adhesions. Treatment of cells with the tyrosine kinase inhibitor herbimycin A diminished the adhesion-induced tyrosine phosphorylation of these proteins and inhibited the formation of focal adhesions and stress fibers. These results suggest a role for integrin-mediated tyrosine phosphorylation in the organization of the cytoskeleton as cells adhere to the extracellular matrix.
培养的细胞在其粘着斑（即细胞与下层基质附着的区域）中显示出高水平的蛋白质酪氨酸磷酸化。我们研究了细胞接种在细胞外基质上时蛋白质的酪氨酸磷酸化情况。接种在纤连蛋白包被表面的大鼠胚胎成纤维细胞、小鼠Balb/c 3T3细胞和NIH 3T3细胞，在115至130 kD的一组蛋白质中显示出磷酸酪氨酸水平升高。当大鼠胚胎成纤维细胞接种在层粘连蛋白或玻连蛋白上时，也观察到酪氨酸磷酸化的增加，但接种在聚赖氨酸或未包被的塑料上时则没有。在接种于纤连蛋白细胞结合片段的细胞以及接种于含有多个RGD序列的合成聚合物的细胞中发现相同的酪氨酸磷酸化升高模式，提示整合素介导了这种效应。我们已确定115 - 130 kD蛋白簇中的一种蛋白质为pp125FAK，这是一种最近定位于粘着斑的酪氨酸激酶（Schaller, M. D., C. A. Borgman, B. S. Cobb, R. R. Vines, A. B. Reynolds, and J. T. Parsons. 1992. Proc. Natl. Acad. Sci. USA. 89:5192）。另一种因细胞外基质黏附而发生酪氨酸磷酸化的蛋白质被鉴定为桩蛋白，这是一种先前定位于粘着斑的70 kD蛋白质。用酪氨酸激酶抑制剂赫伯霉素A处理细胞可减少这些蛋白质的黏附诱导的酪氨酸磷酸化，并抑制粘着斑和应力纤维的形成。这些结果表明，在细胞黏附于细胞外基质时，整合素介导的酪氨酸磷酸化在细胞骨架组织中发挥作用。
