Department of Biochemistry and Molecular Biology, Centre for High-Throughput Biology, University of British Columbia, 2125 East Mall Vancouver, British Columbia V6T1Z4, Canada.
Program in Cell Biology, Hospital for Sick Children, and Biochemistry Department, University of Toronto, Toronto, Ontario M5G 0A4, Canada.
Nat Cell Biol. 2014 Dec;16(12):1227-37. doi: 10.1038/ncb3054. Epub 2014 Oct 26.
The heat-shock response is a complex cellular program that induces major changes in protein translation, folding and degradation to alleviate toxicity caused by protein misfolding. Although heat shock has been widely used to study proteostasis, it remained unclear how misfolded proteins are targeted for proteolysis in these conditions. We found that Rsp5 and its mammalian homologue Nedd4 are important E3 ligases responsible for the increased ubiquitylation induced by heat stress. We determined that Rsp5 ubiquitylates mainly cytosolic misfolded proteins upon heat shock for proteasome degradation. We found that ubiquitylation of heat-induced substrates requires the Hsp40 co-chaperone Ydj1 that is further associated with Rsp5 upon heat shock. In addition, ubiquitylation is also promoted by PY Rsp5-binding motifs found primarily in the structured regions of stress-induced substrates, which can act as heat-induced degrons. Our results support a bipartite recognition mechanism combining direct and chaperone-dependent ubiquitylation of misfolded cytosolic proteins by Rsp5.
热休克反应是一种复杂的细胞程序，它诱导蛋白质翻译、折叠和降解的重大变化，以减轻蛋白质错误折叠引起的毒性。尽管热休克已被广泛用于研究蛋白质稳态，但在这些条件下，错误折叠的蛋白质如何被靶向进行蛋白酶体降解仍不清楚。我们发现，Rsp5 及其哺乳动物同源物 Nedd4 是负责热应激诱导的泛素化增加的重要 E3 连接酶。我们确定 Rsp5 在热休克时主要泛素化细胞质中错误折叠的蛋白质，以进行蛋白酶体降解。我们发现，热诱导底物的泛素化需要 Hsp40 共伴侣 Ydj1，它在热休克时进一步与 Rsp5 相关联。此外，泛素化还受到 PY Rsp5 结合基序的促进，这些基序主要存在于应激诱导底物的结构区域，可作为热诱导的降解基序。我们的研究结果支持一种双识别机制，该机制通过 Rsp5 对细胞质中错误折叠的蛋白质进行直接和伴侣依赖的泛素化。
