Analysis of the molecular mechanism ofendoplasmic reticulum-associated protein degradation (ERAD)

内质网相关蛋白降解（ERAD）分子机制分析

基本信息

批准号：
17570161
负责人：
HOSOKAWA Nobuko
金额：
$ 2.24万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

In the present study, I analyzed the molecular mechanism of endoplasmic reticulum-associated protein degradation (ERAD), and the following points were clarified.1.A genetic variant of α1-antitrypsin NHK is known to be degraded intracellularly by ERAD. NHK is a glycoprotein that has three N-linked oligosaccharides. I have constructed a mutant NHK-QQQ, which lacks N-glycosylation sites, and analyzed the kinetics of degradation. The following points have been clarified. (1)NHK-QQQ was degraded more rapidly than NHK. (2)NHK-QQQ was degraded by the proteasome. (3)NHK recycles between the ER and Golgi apparatus, whereas NHK-QQQ is retained in the ER.2.We have identified a TRAP (translocon associated protein) subunit as a gene whose expression is up-regulated by ER stress. TRAP complex is composed of four subunits, and resides in the ER membrane through association with translocon, although the function of this complex is not well understood. We found that (1) the mRNA of all the four subunits of TRAP complex was up-regulated simultaneously by ER stress. (2)This induction depends on the function of Ire1, a transmembrane sensor protein for ER stress, and XBP1, a transcription factor. (3)Knock-down of one TRAP subunit by RNA interference (RNAi) decreased the expression of TRAP complex. (4)The degradation kinetics of misfolded ERAD substrates were attenuated by knock-down of the TRAP complex. Therefore, it was suggested that TRAP complex is involved in the recognition and retrotranslocation of the ERAD substrates.

在本研究中，我分析了内质网相关蛋白降解（ERAD）的分子机制，并阐明了以下几个点。1。α1-抗替替汀NHK的遗传变异已被ERAD在内部降解。 NHK是一种糖蛋白，具有三种N连接的寡糖。我已经构建了一个缺乏N-糖基化位点的突变NHK-QQQ，并分析了降解的动力学。以下几点已经澄清。（1）NHK-QQQ比NHK更快地降解。（2）NHK-QQQ被蛋白质组降解。（3）ER和高尔基体之间的NHK回收，而NHK-QQQ保留在ER中。2。我们已经确定了一个陷阱（转运蛋白相关蛋白）亚基为一个基因，其表达在ER应力上被上调。 TRAP复合物由四个亚基组成，并通过与Cranslocon关联驻留在ER膜中，尽管该复合物的功能尚不清楚。我们发现（1）仅由ER应力上调了所有四个陷阱复合物亚基的mRNA。（2）该诱导取决于IRE1的功能，IRE1是ER应力的跨膜传感器蛋白，而XBP1是转录因子。（3）通过RNA干扰（RNAi）将一个陷阱亚基敲低可改善陷阱复合物的表达。（4）被陷阱综合体的敲除降低了脱落的伊拉德底物的降解动力学。因此，有人提出，陷阱复合物与ERAD底物的识别和逆转转换有关。