p66shc inhibits pro-survival epidermal growth factor receptor/ERK signaling during severe oxidative stress in mouse renal proximal tubule cells
p66shc inhibits pro-survival epidermal growth factor receptor/ERK signaling during severe oxidative stress in mouse renal proximal tubule cells
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DOI：
10.1074/jbc.m708799200
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发表时间：
2008-03-07
影响因子：
4.8
通讯作者：
Safirstein, Robert L.
中科院分区：
文献类型：
--
作者：
Arany, Istvan;Faisal, Amir;Safirstein, Robert L.
完全执行的表皮生长因子受体 (EGFR)/Ras/MEK/ERK 通路在中度氧化应激下的肾上皮细胞中发挥促生存作用。然而，我们和其他人已经证明，在严重的氧化应激过程中，在培养的肾近曲小管细胞以及缺血/再灌注损伤后的肾近曲小管中，活化的 EGFR 与 ERK 激活断开，导致坏死性死亡。研究表明，ShcA 接头蛋白家族的酪氨酸磷酸化 p46/52 同工型将激活的 EGFR 与 Ras 和 ERK 的激活联系起来，而 p66(shc) 同工型可以抑制 p46/52(shc) 功能。在这里，我们确定严重的氧化应激（在短暂的激活后）会终止 Ras/MEK/ERK 通路的激活，这与 p66(shc) 的 ERK/JNK 依赖性 Ser(36) 磷酸化相一致。 p66(shc) 的异构体特异性敲除或 Ser(36) 突变为 Ala，但不突变为 Asp，在体外减弱了严重氧化应激介导的 ERK 抑制和细胞死亡。此外，严重的氧化应激（与配体刺激和中度氧化应激不同，两者都支持生存）增加了 p66(shc) 与激活的 EGFR 和 Grb2 的结合。这种结合使 SOS1 接头蛋白与 EGFR 招募的信号复合物分离，导致 Ras/MEK/ERK 激活终止。值得注意的是，体内缺血/再灌注损伤后的肾脏中也发生了 p66(shc) 的 Ser(36) 磷酸化及其与 EGFR 结合的增加。与此同时，SOS1 与 EGFR 的结合下降，与体外研究结果相似。因此，我们提出的体外机制提供了一种通过抑制 p66(shc) Ser(36) 磷酸化或体内敲低 p66(sh)c 表达来改善氧化应激诱导的细胞损伤的方法。
The fully executed epidermal growth factor receptor (EGFR)/Ras/MEK/ERK pathway serves a pro-survival role in renal epithelia under moderate oxidative stress. We and others have demonstrated that during severe oxidative stress, however, the activated EGFR is disconnected from ERK activation in cultured renal proximal tubule cells and also in renal proximal tubules after ischemia/reperfusion injury, resulting in necrotic death. Studies have shown that the tyrosine-phosphorylated p46/52 isoforms of the ShcA family of adaptor proteins connect the activated EGFR to activation of Ras and ERK, whereas the p66(shc) isoform can inhibit this p46/52(shc) function. Here, we determined that severe oxidative stress ( after a brief period of activation) terminates activation of the Ras/MEK/ERK pathway, which coincides with ERK/JNK-dependent Ser(36) phosphorylation of p66(shc). Isoform-specific knockdown of p66(shc) or mutation of Ser(36) to Ala, but not to Asp, attenuated severe oxidative stress-mediated ERK inhibition and cell death in vitro. Also, severe oxidative stress ( unlike ligand stimulation and moderate oxidative stress, both of which support survival) increased binding of p66(shc) to the activated EGFR and Grb2. This binding dissociated the SOS1 adaptor protein from the EGFR-recruited signaling complex, leading to termination of Ras/MEK/ERK activation. Notably, Ser(36) phosphorylation of p66(shc) and its increased binding to the EGFR also occurred in the kidney after ischemia/reperfusion injury in vivo. At the same time, SOS1 binding to the EGFR declined, similar to the in vitro findings. Thus, the mechanism we propose in vitro offers a means to ameliorate oxidative stress-induced cell injury by either inhibiting Ser(36) phosphorylation of p66(shc) or knocking down p66(sh)c expression in vivo.