Mitochondrial c-Src regulates cell survival through phosphorylation of respiratory chain components.
Mitochondrial c-Src regulates cell survival through phosphorylation of respiratory chain components.
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DOI：
10.1042/bj20120509
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发表时间：
2012-10-15
期刊：
影响因子：
--
通讯作者：
Homma Y
中科院分区：
文献类型：
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作者：
Ogura M;Yamaki J;Homma MK;Homma Y
线粒体蛋白酪氨酸磷酸化是调节线粒体功能的重要机制。在本研究中，我们已经确定了线粒体中的c-Src的新底物，并研究了它们在氧化磷酸化调节中的功能。Src家族激酶抑制剂PP 2 {氨基-5-（4-氯苯基）-7-（叔丁基）吡唑并[3，4d]嘧啶}表现出呼吸的显著降低。从表达激酶死亡的c-Src的细胞中获得了类似的结果，该细胞含有肿瘤靶向序列。磷酸化位点分析选择c-Src靶标，包括呼吸复合物I的Tyr 193处的NDUFV 2（NADH脱氢酶[泛醌]黄素蛋白2）和复合物II的Tyr 215处的SDHA（琥珀酸脱氢酶A）。通过使用表达其磷酸化缺陷突变体的细胞的体内测定支持c-Src对这些位点的磷酸化。表达野生型蛋白及其突变体的细胞的比较揭示，NDUFV 2磷酸化是NADH脱氢酶活性所需的，影响呼吸活性和细胞ATP含量。SDHA磷酸化对酶活性没有影响，但扰乱了电子传递，从而诱导活性氧。在T98 G细胞和表达这些突变体的原代神经元中观察到活力丧失。这些结果表明，线粒体c-Src调节氧化磷酸化系统的磷酸化呼吸成分和c-Src活性是必不可少的细胞活力。
Mitochondrial protein tyrosine phosphorylation is an important mechanism for the modulation of mitochondrial functions. In the present study, we have identified novel substrates of c-Src in mitochondria and investigated their function in the regulation of oxidative phosphorylation. The Src family kinase inhibitor PP2 {amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo [3,4d] pyrimidine} exhibits significant reduction of respiration. Similar results were obtained from cells expressing kinase-dead c-Src, which harbours a mitochondrial-targeting sequence. Phosphorylation-site analysis selects c-Src targets, including NDUFV2 (NADH dehydrogenase [ubiquinone] flavoprotein 2) at Tyr193 of respiratory complex I and SDHA (succinate dehydrogenase A) at Tyr215 of complex II. The phosphorylation of these sites by c-Src is supported by an in vivo assay using cells expressing their phosphorylation-defective mutants. Comparison of cells expressing wild-type proteins and their mutants reveals that NDUFV2 phosphorylation is required for NADH dehydrogenase activity, affecting respiration activity and cellular ATP content. SDHA phosphorylation shows no effect on enzyme activity, but perturbed electron transfer, which induces reactive oxygen species. Loss of viability is observed in T98G cells and the primary neurons expressing these mutants. These results suggest that mitochondrial c-Src regulates the oxidative phosphorylation system by phosphorylating respiratory components and that c-Src activity is essential for cell viability.