TGFβ1 rapidly activates Src through a non-canonical redox signaling mechanism.
TGFβ1 rapidly activates Src through a non-canonical redox signaling mechanism.
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DOI：
10.1016/j.abb.2015.01.001
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发表时间：
2015-02-15
影响因子：
3.9
通讯作者：
Forman, Henry Jay
中科院分区：
文献类型：
--
作者：
Zhang, Hongqiao;Davies, Kelvin J. A.;Forman, Henry Jay
转化生长因子-β1（TGF-β 1）通过Src激活参与多种细胞过程。在经典途径中，Src激活由pTyr 530去磷酸化引发，随后是允许Tyr 419自磷酸化的构象变化。已经报道了一种非经典途径，其中半胱氨酸的氧化允许绕过pTyr 530去磷酸化。在这里，我们研究了TGF-β如何激活小细胞肺癌细胞系H358细胞中的Src。TGF-β增加Src Tyr 419磷酸化，但令人惊讶的是，Tyr 530磷酸化增加而不是减少。钒酸盐（一种蛋白酪氨酸磷酸酶抑制剂）刺激Src活化本身，但不是抑制TGF-β对Src的活化，钒酸盐的活化与TGF-β是相加的，表明不需要pTyr 530去磷酸化。因此，怀疑参与了非典型的氧化活化。TGF-β瞬时增加细胞外H2 O2，而GSH-酯和过氧化氢酶消除TGF-β对Src的激活。夹竹桃麻素是一种NADPH氧化酶抑制剂，可抑制TGF-β刺激的H2 O2产生。此外，半胱氨酸突变为丙氨酸，248 C/A，277 C/A，或501 C/A废除，而490 C/A显着降低，TGF-β介导的Src激活。综上所述，结果表明TGF-β介导的Src活化主要通过氧化还原依赖性机制起作用，这是由于通过NADPH氧化酶产生的H2 O2增加，并且半胱氨酸248、277、490和501对于该活化是关键的。
Transforming growth factor-β1 (TGF-β) is involved in multiple cellular processes through Src activation. In the canonical pathway, Src activation is initiated by pTyr530 dephosphorylation followed by a conformational change allowing Tyr419 auto-phosphorylation. A non-canonical pathway in which oxidation of cysteine allows bypassing of pTyr530 dephosphorylation has been reported. Here, we examined how TGF-β activates Src in H358 cells, a small cell lung carcinoma cell line. TGF-β increased Src Tyr419 phosphorylation, but surprisingly, Tyr530 phosphorylation was increased rather than decreased. Vanadate, a protein tyrosine phosphatase inhibitor, stimulated Src activation itself, but rather than inhibiting Src activation by TGF-β, activation by vanadate was additive with TGF-β showing that pTyr530 dephosphorylation was not required. Thus, the involvement of the non-canonical oxidative activation was suspected. TGF-β increased extracellular H2O2 transiently while GSH-ester and catalase abrogated Src activation by TGF-β. Apocynin, a NADPH oxidase inhibitor, inhibited TGF-β-stimulated H2O2 production. Furthermore, mutation of cysteines to alanine, 248C/A, 277C/A, or 501C/A abrogated, while 490C/A significantly reduced, TGF-β-mediated Src activation. Taken together, the results indicate that TGF-β-mediated Src activation operates largely through a redox dependent mechanism, resulting from enhanced H2O2 production through an NADPH oxidase and that cysteines 248, 277, 490, and 501 are critical for this activation.
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