Rabinowich H, Manciulea M, Herberman R B, Whiteside T L
Department of Pathology, University of Pittsburgh School of Medicine, PA 15213, USA.
J Immunol. 1996 Nov 1;157(9):3860-8.
Adhesive interactions mediated by cell surface receptors have been shown to induce signal transduction pathways that regulate changes in cellular function. We have reported recently that fibronectin (FN) receptors, alpha4beta1 and alpha5beta1 integrins, on NK cells transduce transmembrane signals leading to tyrosine phosphorylation of 60-, 70-, and 120-kDa proteins. In the current study, we have identified a 120-kDa phosphoprotein as the focal adhesion kinase (p125FAK), a structurally unique nonreceptor protein tyrosine kinase that localizes to focal adhesions. Activity of p125FAK was induced by adhesion of NK cells to plastic-immobilized FN, by cross-linking of cell surface-bound FN or FN fragments, FN120 or FN40, with anti-FN mAb, or by cross-linking of alpha4beta1 or alpha5beta1 integrins with alpha-chain-specific Abs. We also observed that enhanced in vitro kinase activity was associated with immunoprecipitates of alpha4beta1 or alpha5beta1 integrins from lysates of FN-adherent NK cells as compared with BSA-treated NK cells. In addition to p125FAK activity, FN-induced kinase activity was also found to be mediated by Fyn, Lyn, and Zap-70, as assessed by in vitro phosphorylation of the immunoprecipitated kinases in the presence of [gamma-32P]ATP. Clustering of FN receptors on NK cells by agonists such as immobilized FN or alpha4- or alpha5-specific Abs also induced association of Fyn and Zap-70 with p125FAK. Our observations indicate that activation and phosphorylation of p125FAK as well as Zap-70 and certain kinases of the src family play an important role in formation of active signaling complexes in response to triggering via beta1 integrins on NK cells. These results also suggest the existence of cross-talk or points of convergence between the beta1 integrin-mediated and other receptor-signaling pathways.
细胞表面受体介导的黏附相互作用已被证明可诱导调节细胞功能变化的信号转导途径。我们最近报道，NK细胞上的纤连蛋白（FN）受体α4β1和α5β1整合素可转导跨膜信号，导致60 kDa、70 kDa和120 kDa蛋白的酪氨酸磷酸化。在当前研究中，我们已确定一种120 kDa的磷蛋白为粘着斑激酶（p125FAK），它是一种结构独特的非受体蛋白酪氨酸激酶，定位于粘着斑。p125FAK的活性可通过NK细胞与塑料固定的FN黏附、细胞表面结合的FN或FN片段（FN120或FN40）与抗FN单克隆抗体交联，或α4β1或α5β1整合素与α链特异性抗体交联来诱导。我们还观察到，与牛血清白蛋白处理的NK细胞相比，FN黏附的NK细胞裂解物中α4β1或α5β1整合素的免疫沉淀物的体外激酶活性增强。除了p125FAK活性外，通过在[γ-32P]ATP存在下对免疫沉淀激酶进行体外磷酸化评估发现，FN诱导的激酶活性也由Fyn、Lyn和Zap-70介导。激动剂（如固定化FN或α4或α5特异性抗体）使NK细胞上的FN受体聚集，也会诱导Fyn和Zap-70与p125FAK结合。我们的观察结果表明，p125FAK以及Zap-70和src家族的某些激酶的激活和磷酸化在响应NK细胞上β1整合素触发形成活性信号复合物中起重要作用。这些结果还表明β1整合素介导的信号通路与其他受体信号通路之间存在串扰或汇聚点。
