Our previous studies have indicated an essential role of p52shc in mediating IGF-I activation of MAPK in smooth muscle cells (SMC). However, the role of the p66 isoform of shc in IGF-I signal transduction is unclear. In the current study, two approaches were employed to investigate the role of p66shc in mediating IGF-I signaling. Knockdown p66shc by small interfering RNA enhanced IGF-I-stimulated p52shc tyrosine phosphorylation and growth factor receptor-bound protein-2 (Grb2) association, resulting in increased IGF-I-dependent MAPK activation. This was associated with enhanced IGF-I-stimulated cell proliferation. In contrast, knockdown of p66shc did not affect IGF-I-stimulated IGF-I receptor tyrosine phosphorylation. Overexpression of p66shc impaired IGF-I-stimulated p52shc tyrosine phosphorylation and p52shc-Grb2 association. In addition, IGF-I-dependent MAPK activation was also impaired, and SMC proliferation in response to IGF-I was inhibited. IGF-I-dependent cell migration was enhanced by p66shc knockdown and attenuated by p66shc overexpression. Mechanistic studies indicated that p66shc inhibited IGF-I signal transduction via competitively inhibiting the binding of Src homology 2 domain-containing protein tyrosine phosphatase-2 (SHP-2) to SHP substrate-1 (SHPS-1), leading to the disruption of SHPS-1/SHP-2/Src/p52shc complex formation, an event that has been shown previously to be essential for p52shc phosphorylation and Grb2 recruitment. These findings indicate that p66shc functions to negatively regulate the formation of a signaling complex that is required for p52shc activation in response to IGF-I, thus leading to attenuation of IGF-I-stimulated cell proliferation and migration.

译文

我们以前的研究表明p52shc在介导平滑肌细胞 (SMC) 中MAPK的igf-i激活中起重要作用。然而,shc的p66亚型在igf-i信号转导中的作用尚不清楚。在本研究中,采用了两种方法来研究p66shc在介导igf-i信号中的作用。通过小干扰RNA敲除p66shc增强了igf-i刺激的p52shc酪氨酸磷酸化和生长因子受体结合蛋白2 (Grb2) 的关联,从而导致igf-i依赖性MAPK激活增加。这与igf-i刺激的细胞增殖增强有关。相反,敲除p66shc不会影响igf-i刺激的igf-i受体酪氨酸磷酸化。p66shc的过表达损害了igf-i刺激的p52shc酪氨酸磷酸化和p52shc-Grb2缔合。此外,igf-i依赖性MAPK激活也受到损害,并且抑制了响应igf-i的SMC增殖。p66shc敲低增强了igf-i依赖性细胞迁移,p66shc过表达减弱了igf-i依赖性细胞迁移。机理研究表明,p66shc通过竞争性抑制含Src同源结构域的蛋白酪氨酸磷酸酶-2 (SHP-2) 与SHP底物-1 (SHPS-1) 的结合来抑制igf-i信号转导,导致SHPS-1/SHP-2/Src/p52shc复合物形成的破坏,先前已证明对p52shc磷酸化和Grb2募集至关重要的事件。这些发现表明,p66shc对响应igf-i的p52shc激活所需的信号复合物的形成产生负调节作用,从而导致igf-i刺激的细胞增殖和迁移的减弱。

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