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Functional analysis of chaperones in mammals

哺乳动物伴侣的功能分析

基本信息

批准号：
14037257
负责人：
TERADA Kazutoyo
金额：
$ 82.5万
依托单位：
Kumamoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2006
项目状态：
已结题

项目摘要

The mitochondrial serine protease Omi/HtrA2 has a proapoptotic role in mammalian cells. However, neither the topology nor the processing of Omi in mitochondria is clearly understood. To determine the topology of Omi in the mitochondrial IMS, EGFP fusions were expressed with the entire N-terminal segment of full-length Omi, and that without the transmembrane region in the cells. As a result, the transmembrane region determines the topology of Omi in the IMS and anchors the pro form into the inner membrane. Mature Omi largely exists in the IMS as a soluble form. The processing sites of the precursor protein were examined, and we could reveal the importance of three arginine residues for the processing of the N-terminal segment of Omi. (BBRC 2007)Anti-apoptotic Bcl-2 localizes in the membranes of mitochondria and endoplasmic reticulum (ER) and resists a broad range of apoptotic stimuli. However, the precise function of Bcl-2 in ER is still unclear. We herein examined the anti-apoptotic potencies of Bcl-2 in mitochondria and ER in vitro. The mitochondria isolated from HeLa cells, which have little or practically no Bcl-2, were apoptosis-competent. That is, membrane-bound Bax was activated and cytochrome c was released. In the presence of tBid, Bcl-2 in mitochondria and ER are similarly potent in inhibiting Bax-associated apoptosis of other mitochondria, but are regulated by tBid differently. (ECR 2007)Bim, a proapoptotic BH3-only member of the Bcl-2 family is required for initiation of apoptosis. We found that Bim is essential for ER stress-induced apoptosis through two novel pathways, involving protein phosphatase 2A-mediated dephosphorylation, which prevents its ubiquitination and proteasomal degradation and CHOP-C/EBP-mediated direct transcriptional induction. (Cell 2007)

线粒体丝氨酸蛋白酶Omi/HtrA 2在哺乳动物细胞中具有促凋亡作用。然而，线粒体中Omi的拓扑结构和加工都不清楚。为了确定Omi在线粒体IMS中的拓扑结构，用全长Omi的整个N-末端片段和细胞中没有跨膜区的片段表达EGFP融合物。因此，跨膜区决定了顶S中Omi的拓扑结构，并将前体锚定到内膜中。成熟的近江大部分以可溶形式存在于IMS中。对前体蛋白的加工位点进行了研究，我们可以揭示三个精氨酸残基对Omi N-末端片段加工的重要性。(BBRC抗凋亡Bcl-2定位于线粒体和内质网（ER）的膜中，并抵抗广泛的凋亡刺激。然而，Bcl-2在ER中的确切功能仍不清楚。我们在此研究了Bcl-2在体外线粒体和ER中的抗凋亡效力。从HeLa细胞中分离的线粒体，其中有很少或几乎没有Bcl-2，是approximatosis-competent。也就是说，膜结合的Bax被激活，细胞色素c被释放。在存在tBid的情况下，线粒体和ER中的Bcl-2在抑制其他线粒体的Bcl-2相关凋亡中同样有效，但受tBid的调节不同。(ECR 2007）Bim，Bcl-2家族的促凋亡BH 3-唯一成员，是启动凋亡所必需的。我们发现Bim通过两种新的途径对ER应激诱导的细胞凋亡至关重要，包括蛋白磷酸酶2A介导的去磷酸化，这阻止了其泛素化和蛋白酶体降解以及CHOP-C/EBP介导的直接转录诱导。(Cell（2007年）