Degradation Signals for Ubiquitin-Proteasome Dependent Cytosolic Protein Quality Control (CytoQC) in Yeast.
Degradation Signals for Ubiquitin-Proteasome Dependent Cytosolic Protein Quality Control (CytoQC) in Yeast.
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DOI：
10.1534/g3.116.027953
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发表时间：
2016-07-07
期刊：
影响因子：
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通讯作者：
Michaelis S
中科院分区：
文献类型：
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作者：
Maurer MJ;Spear ED;Yu AT;Lee EJ;Shahzad S;Michaelis S
细胞蛋白质量控制（PQC）系统选择性地靶向错误折叠或其他异常蛋白，由泛素-蛋白酶体系统（UPS）降解。细胞如何区分异常蛋白和正常蛋白仍不完全清楚，但部分涉及到泛素E3连接酶和暴露在错误折叠蛋白中的降解信号（degrons）之间的识别。PQC在细胞中是区隔的，近年来人们对er相关降解（ERAD）和核质量控制有了大量的了解。相比之下，细胞质质量控制（CytoQC）的全面观点尚未出现，并将受益于一组定义良好的模型底物的发展。在这项研究中，我们在酿酒酵母中建立了一个等基因的“degron文库”，该文库由附在报告蛋白Ura3的c端上的短序列组成。这些含降解蛋白中约有一半是完整膜E3连接酶Doa10的底物，该酶在ERAD和一些核蛋白降解中也起关键作用。值得注意的是，我们的一些降解融合蛋白表现出对E3连接酶Ltn1/Rkr1的降解依赖，显然其机制不同于其在翻译暂停蛋白的核糖体质量控制中的已知作用。Ubr1和San1是参与识别一些错误折叠的CytoQC底物的E3连接酶，在降解含降解蛋白的过程中基本上是必不可少的。有趣的是，Hsp70/Hsp40伴侣蛋白/伴侣蛋白Ssa1、2和Ydj1是所有测试构建体降解所必需的。综上所述，本文提供的综合degron库为检测候选PQC成分和鉴定新成分提供了重要的等基因底物资源。
Cellular protein quality control (PQC) systems selectively target misfolded or otherwise aberrant proteins for degradation by the ubiquitin-proteasome system (UPS). How cells discern abnormal from normal proteins remains incompletely understood, but involves in part the recognition between ubiquitin E3 ligases and degradation signals (degrons) that are exposed in misfolded proteins. PQC is compartmentalized in the cell, and a great deal has been learned in recent years about ER-associated degradation (ERAD) and nuclear quality control. In contrast, a comprehensive view of cytosolic quality control (CytoQC) has yet to emerge, and will benefit from the development of a well-defined set of model substrates. In this study, we generated an isogenic “degron library” in Saccharomyces cerevisiae consisting of short sequences appended to the C-terminus of a reporter protein, Ura3. About half of these degron-containing proteins are substrates of the integral membrane E3 ligase Doa10, which also plays a pivotal role in ERAD and some nuclear protein degradation. Notably, some of our degron fusion proteins exhibit dependence on the E3 ligase Ltn1/Rkr1 for degradation, apparently by a mechanism distinct from its known role in ribosomal quality control of translationally paused proteins. Ubr1 and San1, E3 ligases involved in the recognition of some misfolded CytoQC substrates, are largely dispensable for the degradation of our degron-containing proteins. Interestingly, the Hsp70/Hsp40 chaperone/cochaperones Ssa1,2 and Ydj1, are required for the degradation of all constructs tested. Taken together, the comprehensive degron library presented here provides an important resource of isogenic substrates for testing candidate PQC components and identifying new ones.