Protein kinase Cε interacts with and inhibits the permeability transition pore in cardiac mitochondria
Protein kinase Cε interacts with and inhibits the permeability transition pore in cardiac mitochondria
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DOI：
10.1161/01.res.0000069215.36389.8d
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发表时间：
2003-05-02
影响因子：
20.1
通讯作者：
Ping, PP
中科院分区：
文献类型：
--
作者：
Baines, CP;Song, CX;Ping, PP
尽管蛋白激酶 Cepsilon (PKCepsilon) 和线粒体之间的功能耦合与心脏保护的发生有关，但实现这种联系的信号转导机制以及 PKCepsilon 调节的线粒体蛋白的身份仍然未知。基于最近的证据表明线粒体通透性转换孔可能参与缺血/再灌注损伤，我们假设 PKCepsilon 和线粒体孔成分之间的蛋白质-蛋白质相互作用可能作为调节孔功能的信号传导机制，从而产生心脏保护作用。小鼠心脏线粒体的共免疫沉淀和基于 GST 的亲和力下拉揭示了 PKCepsilon 与孔的成分，即电压依赖性阴离子通道 (VDAC)、腺嘌呤核苷酸转位酶 (ANT) 和己糖激酶 II (HKII) 的相互作用。 VDAC1、ANT1 和 HKII 在 PKCepsilon 复合体中的存在量分别约为其总表达量的 2%、0.2% 和 1%。此外，体外研究表明 PKCepsilon 可以直接结合并磷酸化 VDAC1。将分离的心脏线粒体与重组 PKCepsilon 一起孵育，可显着抑制 Ca2+ 诱导的线粒体肿胀（孔开放的指标）。此外，活性 PKCepsilon 在小鼠体内的心脏特异性表达具有心脏保护作用，大大增加了 PKCepsilon 与孔成分的相互作用，并抑制 Ca2+ 诱导的孔开放。相反，激酶失活的 PKCepsilon 的心脏表达不影响孔的开放。最后，开孔剂白术苷的施用显着减弱了 PKCepsilon 转基因的梗塞保护作用。总的来说，这些数据表明 PKCepsilon 与心脏线粒体孔的成分形成物理相互作用。这反过来又抑制了孔的病理功能，并有助于 PKCepsilon 诱导的心脏保护作用。
Although functional coupling between protein kinase Cepsilon (PKCepsilon) and mitochondria has been implicated in the genesis of cardioprotection, the signal transduction mechanisms that enable this link and the identities of the mitochondrial proteins modulated by PKCepsilon remain unknown. Based on recent evidence that the mitochondrial permeability transition pore may be involved in ischemia/reperfusion injury, we hypothesized that protein-protein interactions between PKCepsilon and mitochondrial pore components may serve as a signaling mechanism to modulate pore function and thus engender cardioprotection. Coimmunoprecipitation and GST-based affinity pull-down from mouse cardiac mitochondria revealed interaction of PKCepsilon with components of the pore, namely voltage-dependent anion channel (VDAC), adenine nucleotide translocase (ANT), and hexokinase II (HKII). VDAC1, ANT1, and HKII were present in the PKCepsilon complex at approximate to2%, approximate to0.2%, and approximate to1% of their total expression, respectively. Moreover, in vitro studies demonstrated that PKCepsilon can directly bind and phosphorylate VDAC1. Incubation of isolated cardiac mitochondria with recombinant PKCepsilon resulted in a significant inhibition of Ca2+-induced mitochondrial swelling, an index of pore opening. Furthermore, cardiac-specific expression of active PKCepsilon in mice, which is cardioprotective, greatly increased interaction of PKCepsilon with the pore components and inhibited Ca2+-induced pore opening. In contrast, cardiac expression of kinase-inactive PKCepsilon did not affect pore opening. Finally, administration of the pore opener atractyloside significantly attenuated the infarct-sparing effect of PKCepsilon transgenesis. Collectively, these data demonstrate that PKCepsilon forms physical interactions with components of the cardiac mitochondrial pore. This in turn inhibits the pathological function of the pore and contributes to PKCepsilon-induced cardioprotection.