A Nucleus-targeted Alternately Spliced Nix/Bnip3L Protein Isoform Modifies Nuclear Factor κB (NFκB)-mediated Cardiac Transcription
A Nucleus-targeted Alternately Spliced Nix/Bnip3L Protein Isoform Modifies Nuclear Factor κB (NFκB)-mediated Cardiac Transcription
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DOI：
10.1074/jbc.m113.452342
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发表时间：
2013-05-31
影响因子：
4.8
通讯作者：
Dorn, Gerald W., II
中科院分区：
文献类型：
--
作者：
Chen, Yun;Decker, Keith F.;Dorn, Gerald W., II
几种Bcl 2家族蛋白质既表达为线粒体靶向全长，又表达为胞浆截短的交替剪接亚型。恶性表达的较短Bcl 2家族同种型可以异型结合并阻止其全长类似物的线粒体定位，从而通过螯合抑制其活性。这种“海绵”作用需要1：1的表达化学计量;如果没有这种作用，则建议替代作用。在此，RNA测序揭示了心脏中仅BH 3蛋白Nix/Bnip 3L（Nix）及其交替剪接可溶形式（sNix）的协调调节，但相对sNix/Nix表达类似于1：10。因此，我们研究了sNix的其他假定功能。虽然Nix在H9 c2大鼠成肌细胞中表达定位于线粒体，但sNix表现出可变的细胞质和细胞核分布。肿瘤坏死因子α（TNF α）诱导快速且完全的sNix核质易位，同时伴有NF kappa B的p65/RelA亚基的核易位。TNF α刺激后sNix与p65/RelA共定位和共沉淀; TNF α诱导的sNix核转位在p65/RelA缺失的鼠胚胎成纤维细胞中未发生。TNF α刺激的H9 c2细胞的ChIP测序显示sNix抑制p65/RelA与较弱DNA结合位点的亚组的结合，解释了其改变培养细胞和体内小鼠心脏中基因表达的能力。这些发现揭示了TNF α刺激的交替剪接的非线粒体Nix同种型的细胞质-核穿梭，并揭示了sNix作为TNF α/NF κ B刺激的心脏基因表达的调节剂的作用。sNix和Nix的转录共调节与TNF α对sNix的翻译后调节相结合，包含了外源性死亡受体和内源性线粒体凋亡途径之间分子串扰的先前未知机制。
Several Bcl2 family proteins are expressed both as mitochondrial-targeted full-length and as cytosolic truncated alternately spliced isoforms. Recombinantly expressed shorter Bcl2 family isoforms can heterotypically bind to and prevent mitochondrial localization of their full-length analogs, thus suppressing their activity by sequestration. This "sponge" role requires 1:1 expression stoichiometry; absent this an alternate role is suggested. Here, RNA sequencing revealed coordinate regulation of BH3-only protein Nix/Bnip3L (Nix) and its alternately spliced soluble form (sNix) in hearts, but relative sNix/Nix expression of similar to 1:10. Accordingly, we examined other putative functions of sNix. Although Nix expressed in H9c2 rat myoblasts localized to mitochondria, sNix showed variable cytoplasmic and nuclear distribution. Tumor necrosis factor alpha (TNF alpha) induced rapid and complete sNix nucleoplasmic translocation concomitant with nuclear translocation of the p65/RelA subunit of NF kappa B. sNix co-localized and co-precipitated with p65/RelA after TNF alpha stimulation; TNF alpha-induced sNix nuclear translocation did not occur in p65/RelA null murine embryonic fibroblasts. ChIP sequencing of TNF alpha-stimulated H9c2 cells revealed sNix suppression of p65/RelA binding to a subset of weaker DNA binding sites, accounting for its ability to alter gene expression in cultured cells and in vivo mouse hearts. These findings reveal TNF alpha-stimulated cytoplasmic-nuclear shuttling of the alternately spliced non-mitochondrial Nix isoform and uncover a role for sNix as a modulator of TNF alpha/NF kappa B-stimulated cardiac gene expression. Transcriptional co-regulation of sNix and Nix, combined with sNix posttranslational regulation by TNF alpha, comprises a previously unknown mechanism for molecular cross-talk between extrinsic death receptor and intrinsic mitochondrial apoptosis pathways.