Lebrun P, Mothe-Satney I, Delahaye L, Van Obberghen E, Baron V
INSERM U145, Avenue de Valombrose, 06107 Nice Cédex 2, France.
J Biol Chem. 1998 Nov 27;273(48):32244-53. doi: 10.1074/jbc.273.48.32244.
Insulin receptor substrate-1 (IRS-1) is a major substrate of insulin and insulin-like growth factor-I receptors, which upon phosphorylation on tyrosine docks several signaling molecules. Recently, IRS-1 was found to interact with alphav beta3 integrins upon insulin stimulation. Integrins are transmembrane proteins that play an important role in adhesion between cells and between cells and extracellular matrix. One of the major proteins implicated in integrin signaling is pp125(FAK), a cytosolic tyrosine kinase, which upon integrin engagement becomes tyrosine-phosphorylated and subsequently binds to c-Src. Here, we established a mammalian two-hybrid system to show that pp125(FAK) binds to IRS-1. This association depends largely on the C terminus of pp125(FAK) but not on pp125(FAK) tyrosine kinase activity. Furthermore, we observed co-immunoprecipitation of pp125(FAK) with IRS-1 in 293 cells, suggesting a possible biological function of this association. When IRS-1 was expressed in 293 cells together with pp125(FAK) or Src, we found extensive IRS-1 tyrosine phosphorylation. In pp125(FAK)-expressing cells, this was concomitant with increased association of IRS-1 with Src homology 2-containing proteins such as growth factor receptor-bound protein 2, phosphatidylinositol (PI) 3-kinase p85alpha subunit, and Src homology 2-containing protein-tyrosine phosphatase-2. In addition, pp125(FAK)-induced association of IRS-1 with PI 3-kinase resulted in increased PI 3-kinase activity. In contrast, no change in mitogen-activated protein kinase activity was observed, indicating that pp125(FAK)-induced association between IRS-1 and growth factor receptor-bound protein 2 does not affect the mitogen-activated protein kinase pathway. Moreover, we found that engagement of integrins induced IRS-1 tyrosine phosphorylation. Considering our results together, we suggest that integrins and insulin/insulin-like growth factor-I receptor signaling pathways converge at an early point in the signaling cascade, which is the IRS-1 protein.
胰岛素受体底物-1（IRS-1）是胰岛素和胰岛素样生长因子-I受体的主要底物，其酪氨酸磷酸化后可结合多种信号分子。最近发现，胰岛素刺激时IRS-1可与αvβ3整合素相互作用。整合素是跨膜蛋白，在细胞间以及细胞与细胞外基质间的黏附中起重要作用。参与整合素信号传导的主要蛋白之一是pp125（FAK），一种胞质酪氨酸激酶，整合素激活后它会发生酪氨酸磷酸化，随后与c-Src结合。在此，我们建立了一个哺乳动物双杂交系统，以证明pp125（FAK）与IRS-1结合。这种结合很大程度上依赖于pp125（FAK）的C末端，但不依赖于pp125（FAK）的酪氨酸激酶活性。此外，我们在293细胞中观察到pp125（FAK）与IRS-1的共免疫沉淀，提示这种结合可能具有生物学功能。当IRS-1与pp125（FAK）或Src在293细胞中共同表达时，我们发现IRS-1发生广泛的酪氨酸磷酸化。在表达pp125（FAK）的细胞中，这伴随着IRS-1与含Src同源2结构域的蛋白（如生长因子受体结合蛋白2、磷脂酰肌醇（PI）3激酶p85α亚基和含Src同源2结构域的蛋白酪氨酸磷酸酶-2）的结合增加。此外，pp125（FAK）诱导的IRS-1与PI 3激酶的结合导致PI 3激酶活性增加。相反，未观察到丝裂原活化蛋白激酶活性的变化，表明pp125（FAK）诱导的IRS-1与生长因子受体结合蛋白2之间的结合不影响丝裂原活化蛋白激酶途径。此外，我们发现整合素的激活可诱导IRS-1酪氨酸磷酸化。综合我们的结果，我们认为整合素和胰岛素/胰岛素样生长因子-I受体信号通路在信号级联反应的早期交汇，交汇点是IRS-1蛋白。
