DDI2 Is a Ubiquitin-Directed Endoprotease Responsible for Cleavage of Transcription Factor NRF1
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DDI2 Is a Ubiquitin-Directed Endoprotease Responsible for Cleavage of Transcription Factor NRF1. / Dirac-Svejstrup, A. Barbara; Walker, Jane; Faull, Peter; Encheva, Vesela; Akimov, Vyacheslav; Puglia, Michele; Perkins, David; Kümper, Sandra; Hunjan, Suchete S.; Blagoev, Blagoy; Snijders, Ambrosius P.; Powell, David J.; Svejstrup, Jesper Q.
I: Molecular Cell, Bind 79, Nr. 2, 16.07.2020, s. 332-341.e7.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - DDI2 Is a Ubiquitin-Directed Endoprotease Responsible for Cleavage of Transcription Factor NRF1
AU - Dirac-Svejstrup, A. Barbara
AU - Walker, Jane
AU - Faull, Peter
AU - Encheva, Vesela
AU - Akimov, Vyacheslav
AU - Puglia, Michele
AU - Perkins, David
AU - Kümper, Sandra
AU - Hunjan, Suchete S.
AU - Blagoev, Blagoy
AU - Snijders, Ambrosius P.
AU - Powell, David J.
AU - Svejstrup, Jesper Q.
N1 - Funding Information: This work was supported by the Francis Crick Institute (FCI), which receives its core funding from Cancer Research UK ( FC001166 ), the UK Medical Research Council ( FC001166 ), and the Wellcome Trust ( FC001166 ), and by a grant from the European Research Council, Agreements 693327 (TRANSDAM) to JQS, while work in the Blagoev lab was funded by the Danish National Research Foundation (ATLAS, DNRF grant No. 141 ) and the Novo Nordisk Foundation ( NNF18OC0052768 ). We thank Cell Services, the Crick Flow Cytometry Platform, and the High Throughput Screening Facility of the FCI for their support and time spent on this project and Mao Xiang Chen at GSK who designed the lentivirus constructs expressing human DDI2 and generated the infected HEK293 cell pools. Publisher Copyright: © 2020 The Author(s)
PY - 2020/7/16
Y1 - 2020/7/16
N2 - The Ddi1/DDI2 proteins are ubiquitin shuttling factors, implicated in a variety of cellular functions. In addition to ubiquitin-binding and ubiquitin-like domains, they contain a conserved region with similarity to retroviral proteases, but whether and how DDI2 functions as a protease has remained unknown. Here, we show that DDI2 knockout cells are sensitive to proteasome inhibition and accumulate high-molecular weight, ubiquitylated proteins that are poorly degraded by the proteasome. These proteins are targets for the protease activity of purified DDI2. No evidence for DDI2 acting as a de-ubiquitylating enzyme was uncovered, which could suggest that it cleaves the ubiquitylated protein itself. In support of this idea, cleavage of transcription factor NRF1 is known to require DDI2 activity in vivo. We show that DDI2 is indeed capable of cleaving NRF1 in vitro but only when NRF1 protein is highly poly-ubiquitylated. Together, these data suggest that DDI2 is a ubiquitin-directed endoprotease.
AB - The Ddi1/DDI2 proteins are ubiquitin shuttling factors, implicated in a variety of cellular functions. In addition to ubiquitin-binding and ubiquitin-like domains, they contain a conserved region with similarity to retroviral proteases, but whether and how DDI2 functions as a protease has remained unknown. Here, we show that DDI2 knockout cells are sensitive to proteasome inhibition and accumulate high-molecular weight, ubiquitylated proteins that are poorly degraded by the proteasome. These proteins are targets for the protease activity of purified DDI2. No evidence for DDI2 acting as a de-ubiquitylating enzyme was uncovered, which could suggest that it cleaves the ubiquitylated protein itself. In support of this idea, cleavage of transcription factor NRF1 is known to require DDI2 activity in vivo. We show that DDI2 is indeed capable of cleaving NRF1 in vitro but only when NRF1 protein is highly poly-ubiquitylated. Together, these data suggest that DDI2 is a ubiquitin-directed endoprotease.
KW - Bortezomib
KW - Ddi1
KW - DDI2
KW - NFE2L1
KW - NRF1
KW - proteasome
KW - proteasome inhibition
KW - ubiquitin
KW - ubiquitin protease
U2 - 10.1016/j.molcel.2020.05.035
DO - 10.1016/j.molcel.2020.05.035
M3 - Journal article
C2 - 32521225
AN - SCOPUS:85086917252
VL - 79
SP - 332-341.e7
JO - Molecular Cell
JF - Molecular Cell
SN - 1097-2765
IS - 2
ER -
ID: 331575594