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Published December 2014 | Supplemental Material + Accepted Version
Journal Article Open

Rsp5/Nedd4 is the main ubiquitin ligase that targets cytosolic misfolded proteins following heat stress


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.

Additional Information

© 2014 Macmillan Publishers Limited. Received 07 January 2014. Accepted 17 September 2014. Published online 26 October 2014. We thank N. Stoynov for his support for the mass spectrometry analyses, all colleagues cited in Methods who provided reagents and a previous anonymous reviewer who suggested investigating Rsp5. This work was supported by a CIHR grant. R.J.D. is an HHMI investigator and T.M. is a CIHR and MSFHR new investigator.

Attached Files

Accepted Version - nihms6319.pdf

Supplemental Material - ncb3054-s1.pdf

Supplemental Material - ncb3054-s2.xlsx

Supplemental Material - ncb3054-s3.xlsx

Supplemental Material - ncb3054-s4.xlsx

Supplemental Material - ncb3054-s5.xlsx

Supplemental Material - ncb3054-s6.xlsx

Supplemental Material - ncb3054-sf1.jpg

Supplemental Material - ncb3054-sf2.jpg

Supplemental Material - ncb3054-sf3.jpg

Supplemental Material - ncb3054-sf4.jpg

Supplemental Material - ncb3054-sf5.jpg

Supplemental Material - ncb3054-sf6.jpg

Supplemental Material - ncb3054-sf7.jpg

Supplemental Material - ncb3054-sf8.jpg


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Additional details

August 20, 2023
October 17, 2023