Abstract
Although most eukaryotic proteins are targeted for proteasomal degradation by ubiquitination, a subset have been demonstrated to undergo ubiquitin-independent proteasomal degradation (UbInPD). However, little is known about the molecular mechanisms driving UbInPD and the degrons involved. Utilizing the GPS-peptidome approach, a systematic method for degron discovery, we found thousands of sequences that promote UbInPD; thus, UbInPD is more prevalent than currently appreciated. Furthermore, mutagenesis experiments revealed specific C-terminal degrons required for UbInPD. Stability profiling of a genome-wide collection of human open reading frames identified 69 full-length proteins subject to UbInPD. These included REC8 and CDCA4, proteins which control proliferation and survival, as well as mislocalized secretory proteins, suggesting that UbInPD performs both regulatory and protein quality control functions. In the context of full-length proteins, C termini also play a role in promoting UbInPD. Finally, we found that Ubiquilin family proteins mediate the proteasomal targeting of a subset of UbInPD substrates.
| Original language | English |
|---|---|
| Pages (from-to) | 1921-1935.e7 |
| Journal | Molecular Cell |
| Volume | 83 |
| Issue number | 11 |
| DOIs | |
| State | Published - 1 Jun 2023 |
Bibliographical note
Publisher Copyright:© 2023 The Authors
Funding
We thank J. Adler (Y. Shaul lab, Weizmann Institute of Science) for providing PSMD1 and luciferase shRNAs, I. Sahu for help in purifying proteasomes, and M. Sharon (Weizmann Institute of Science) for providing pET28 vector. We are grateful to H. Hauschner for FACS. I.K. is supported by European Research Council (ERC-2020-STG 947709), Israel Science Foundation (ISF grants no. 2380/21 and 3096/21), Alon Fellowship, and The Applebaum Foundation. M.H.G. is funded by ISF 755/19, NSFC-ISF 2512/18 and a Russell Berrie Nanotechnology NEVET grant to A.R.W. and M.H.G. S.J.E. is funded by NIH grant AG11085. S.J.E. is an investigator with the Howard Hughes Medical Institute. R.T.T. is the recipient of a Pemberton-Trinity Fellowship. Conceptualization, S.J.E. and I.K.; supervision, I.K.; methodology, S.J.E. and I.K.; software, Y.M. A.R. M.I.G. S.B. R.T.T. and I.K.; formal analysis, Y.M. R.T.T. A.R.W. M.H.G. and I.K.; investigation, Y.M. A.R. R.T.T. A.R.W. A.B. J.G.-H. M.H.G. and I.K.; writing – original draft, Y.M. and I.K.; writing & editing, Y.M. and I.K. S.J.E. is a member of the Molecular Cell advisory board. We thank J. Adler (Y. Shaul lab, Weizmann Institute of Science) for providing PSMD1 and luciferase shRNAs, I. Sahu for help in purifying proteasomes, and M. Sharon (Weizmann Institute of Science) for providing pET28 vector. We are grateful to H. Hauschner for FACS. I.K. is supported by European Research Council ( ERC-2020-STG 947709 ), Israel Science Foundation (ISF grants no. 2380/21 and 3096/21 ), Alon Fellowship , and The Applebaum Foundation . M.H.G. is funded by ISF 755/19 , NSFC-ISF 2512/18 and a Russell Berrie Nanotechnology NEVET grant to A.R.W. and M.H.G. S.J.E. is funded by NIH grant AG11085 . S.J.E. is an investigator with the Howard Hughes Medical Institute. R.T.T. is the recipient of a Pemberton-Trinity Fellowship .
| Funders | Funder number |
|---|---|
| NSFC-ISF | 2512/18 |
| National Institutes of Health | AG11085 |
| Howard Hughes Medical Institute | |
| Applebaum Foundation | 755/19 |
| European Commission | ERC-2020-STG 947709 |
| Weizmann Institute of Science | |
| Israel Science Foundation | 3096/21, 2380/21 |
Keywords
- C-degron
- Global Protein Stability (GPS) technology
- Ubiquilin
- Ubiquitin
- degron
- proteasome
- ubiquitin-independent degradation