A GH3-like domain in reaper is required for mitochondrial localization and induction of IAP degradation
A GH3-like domain in reaper is required for mitochondrial localization and induction of IAP degradation
复制标题
DOI：
10.1074/jbc.m308055200
复制
发表时间：
2003-11-07
影响因子：
4.8
通讯作者：
Kornbluth, S
中科院分区：
文献类型：
--
作者：
Olson, MR;Holley, CL;Kornbluth, S
Reaper是一种有效的促凋亡蛋白，最初是在筛选凋亡诱导缺陷的果蝇突变体中鉴定的。该蛋白具有多种功能，包括抑制IAP（细胞凋亡抑制剂）;诱导IAP降解;抑制蛋白质翻译;以及当在脊椎动物细胞中表达时，诱导线粒体细胞色素c释放。Reaper的结构/功能分析已经确定了一个极端的N-末端基序，似乎足以抑制IAP功能。我们在这里报告说，这个域，虽然需要IAP不稳定，是不够的。此外，我们已经确定了一个小区域的收割机，类似于GH 3结构域的Grim，这是需要本地化的收割机线粒体，诱导IAP降解，并有效的细胞杀伤。尽管缺乏GH 3结构域的突变型Reaper蛋白在这些性质上是缺陷的，但是这些缺陷可以通过附加来自Bcl-x（L）的C-末端线粒体靶向序列或来自促凋亡蛋白HID的同源区域来完全纠正。总之，这些数据强烈表明，IAP不稳定的收割机在完整的细胞需要收割机定位到线粒体和诱导IAP不稳定的收割机是重要的，在果蝇细胞的细胞凋亡的有效诱导。
Reaper is a potent pro-apoptotic protein originally identified in a screen for Drosophila mutants defective in apoptotic induction. Multiple functions have been ascribed to this protein, including inhibition of IAPs (inhibitors of apoptosis); induction of IAP degradation; inhibition of protein translation; and when expressed in vertebrate cells, induction of mitochondrial cytochrome c release. Structure/function analysis of Reaper has identified an extreme N-terminal motif that appears to be sufficient for inhibition of IAP function. We report here that this domain, although required for IAP destabilization, is not sufficient. Moreover, we have identified a small region of Reaper, similar to the GH3 domain of Grim, that is required for localization of Reaper to mitochondria, induction of IAP degradation, and potent cell killing. Although a mutant Reaper protein lacking the GH3 domain was deficient in these properties, these defects could be fully rectified by appending either the C-terminal mitochondrial targeting sequence from Bcl-x(L) or a homologous region from the pro-apoptotic protein HID. Together, these data strongly suggest that IAP destabilization by Reaper in intact cells requires Reaper localization to mitochondria and that induction of IAP instability by Reaper is important for the potent induction of apoptosis in Drosophila cells.