Trencia Alessandra, Fiory Francesca, Maitan Maria Alessandra, Vito Pasquale, Barbagallo Alessia Paola Maria, Perfetti Anna, Miele Claudia, Ungaro Paola, Oriente Francesco, Cilenti Lucia, Zervos Antonis S, Formisano Pietro, Beguinot Francesco
Dipartimento di Biologia e Patologia Cellulare e Molecolare and Istituto di Endocrinologia ed Oncologia Sperimentale del CNR, Università degli Studi di Napoli Federico II, Naples 80131, Italy.
J Biol Chem. 2004 Nov 5;279(45):46566-72. doi: 10.1074/jbc.M406317200. Epub 2004 Aug 24.
ped/pea-15 is a ubiquitously expressed 15-kDa protein featuring a broad anti-apoptotic function. In a yeast two-hybrid screen, the pro-apoptotic Omi/HtrA2 mitochondrial serine protease was identified as a specific interactor of the ped/pea-15 death effector domain. Omi/HtrA2 also bound recombinant ped/pea-15 in vitro and co-precipitated with ped/pea-15 in 293 and HeLa cell extracts. In these cells, the binding of Omi/HtrA2 to ped/pea-15 was induced by UVC exposure and followed the mitochondrial release of Omi/HtrA2 into the cytoplasm. Upon UVC exposure, cellular ped/pea-15 protein expression levels decreased. This effect was prevented by the ucf-101 specific inhibitor of the Omi/HtrA2 proteolytic activity, in a dose-dependent fashion. In vitro incubation of ped/pea-15 with Omi/HtrA2 resulted in ped/pea-15 degradation. In intact cells, the inhibitory action of ped/pea-15 on UVC-induced apoptosis progressively declined at increasing Omi/HtrA2 expression. This further effect of Omi/HtrA2 was also inhibited by ucf-101. In addition, ped/pea-15 expression blocked Omi/HtrA2 co-precipitation with the caspase inhibitor protein XIAP and caspase 3 activation. Thus, in part, apoptosis following Omi/HtrA2 mitochondrial release is mediated by reduction in ped/pea-15 cellular levels. The ability of Omi/HtrA2 to relieve XIAP inhibition on caspases is modulated by the relative levels of Omi/HtrA2 and ped/pea-15.
ped/pea - 15是一种广泛表达的15 kDa蛋白，具有广泛的抗凋亡功能。在酵母双杂交筛选中，促凋亡的Omi/HtrA2线粒体丝氨酸蛋白酶被鉴定为ped/pea - 15死亡效应结构域的特异性相互作用分子。Omi/HtrA2在体外也能与重组ped/pea - 15结合，并在293和HeLa细胞提取物中与ped/pea - 15共沉淀。在这些细胞中，Omi/HtrA2与ped/pea - 15的结合是由紫外线（UVC）照射诱导的，并且伴随着Omi/HtrA2从线粒体释放到细胞质中。在UVC照射后，细胞内ped/pea - 15蛋白表达水平下降。Omi/HtrA2蛋白水解活性的特异性抑制剂ucf - 101能够以剂量依赖的方式阻止这种效应。ped/pea - 15与Omi/HtrA2在体外孵育会导致ped/pea - 15降解。在完整细胞中，随着Omi/HtrA2表达的增加，ped/pea - 15对UVC诱导的凋亡的抑制作用逐渐减弱。Omi/HtrA2的这种进一步作用也被ucf - 101抑制。此外，ped/pea - 15的表达阻止了Omi/HtrA2与半胱天冬酶抑制剂蛋白XIAP的共沉淀以及半胱天冬酶3的激活。因此，Omi/HtrA2从线粒体释放后的凋亡部分是由细胞内ped/pea - 15水平的降低介导的。Omi/HtrA2解除XIAP对半胱天冬酶抑制的能力受到Omi/HtrA2和ped/pea - 15相对水平的调节。
