Src kinase activity is required for integrin αVβ3-mediated activation of nuclear factor-κB
Src kinase activity is required for integrin αVβ3-mediated activation of nuclear factor-κB
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DOI：
10.1074/jbc.m412555200
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发表时间：
2005-04-01
影响因子：
4.8
通讯作者：
Giachelli, CM
中科院分区：
文献类型：
--
作者：
Courter, DL;Lomas, L;Giachelli, CM
整合素粘附于细胞外基质蛋白保护粘附依赖性细胞免于悬浮诱导的凋亡。先前的研究表明，转录因子核因子-κ B的激活对于整合素α（v）β（3）配体骨桥蛋白保护内皮细胞免受血清戒断引起的凋亡是必要的。在这项研究中，β（3）整合素在平滑肌细胞中过度表达。与载体对照组相比，在骨桥蛋白上培养时，过表达野生型β 3的细胞具有增强的细胞粘附、细胞铺展和核因子-κ B活化。从β 3的C末端去除四个氨基酸（759 X）消除了整合素促进这些过程的能力。单个氨基酸取代表明，磷酸化酪氨酸759是不需要的转录因子的激活，但这个残基似乎发挥了结构性作用，因为突变为丙氨酸显着抑制核因子-κ B激活。酪氨酸激酶的Src家族代表整合素信号传导过程中的重要转导子，并且β（3）的C末端已经被认为是Src的结合位点。免疫沉淀证实Src与野生型β 3整合素结合，但Src与缺乏C末端的整合素（759 X）不形成复合物。Src抑制剂PP 2（4-氨基-5-（4-氯苯基）-7-（叔丁基）吡唑并[3，4-d]嘧啶）的药理学抑制或激酶死亡c-Src的过表达阻断了核因子-κ B的激活。与对照成纤维细胞相比，Src缺陷的小鼠胚胎成纤维细胞在骨桥蛋白上接种时未能激活核因子-κ B。总之，这些实验表明β 3的C末端和Src活性是整联蛋白α v β 3介导的核因子-κ B活化所必需的。
Integrin adhesion to extracellular matrix proteins protects adhesion-dependent cells from suspension-induced apoptosis. Previous studies indicate that activation of the transcription factor nuclear factor-kappa B was necessary for the integrin alpha(v)beta(3) ligand osteopontin to protect endothelial cells from apoptosis caused by serum withdrawal. In this study, beta(3) integrins were over-expressed in smooth muscle cells. When plated on osteopontin, cells overexpressing wild-type beta(3) had enhanced cell adhesion, cell spreading, and nuclear factor-kappa B activation compared with vector control. Removal of four amino acids (759X) from the C terminus of beta(3) eliminated the ability of the integrin to promote these processes. Single amino acid substitutions indicated that phosphorylation at tyrosine 759 was not required for activation of the transcription factor, however this residue appeared to play a structural role, because mutation to alanine significantly inhibited nuclear factor-kappa B activation. The Src family of tyrosine kinases represents important transducers during integrin signaling, and the C terminus of beta(3) has been implicated as the binding site for Src. Immunoprecipitations demonstrated that Src associated with wild-type beta(3) integrins, but Src and integrins lacking the C terminus (759X) did not form a complex. Pharmacological inhibition with the Src inhibitor PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine) or overexpression of kinase-dead c-Src blocked nuclear factor-kappa B activation. Mouse embryonic fibroblasts deficient for Src failed to activate nuclear factor-kappa B when plated on osteopontin, in contrast to control fibroblasts. Together, these experiments indicate that the C terminus of beta(3) and Src activity are required for integrin alpha(v)beta(3)-mediated nuclear factor-kappa B activation.