TY - JOUR
T1 - Cullin 2-RBX1 E3 ligase and USP2 regulate antithrombin ubiquitination and stability
AU - Xu, Dacai
AU - Wu, Jiawen
AU - Chen, Jinghong
AU - Jiang, Liling
AU - Chen, Juan
AU - Bao, Wenhao
AU - Chen, Xin
AU - Yang, Qianqian
AU - Zhang, Xiaolan
AU - Yao, Leyi
AU - Su, Huabo
AU - Liu, Jinbao
N1 - Funding Information:
The study was supported by the National Funds for Developing Local Colleges and Universities (B16056001), the National Natural Science Foundation of China (81900365), the Natural Science Foundation research team of Guangdong Province (2018B030312001), the Science and Technology Program of Guangzhou (201604020001), Innovative AcademicTeam of Guangzhou Education System (1201610014), and the Research Team of Department of Education of Guangdong Province (2017KCXTD027).
Publisher Copyright:
© 2021 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology
PY - 2021/8
Y1 - 2021/8
N2 - Hemophilia A and B are congenital bleeding disorders caused by a deficiency in pro-coagulant factor VIII or IX that is treated by downregulation of antithrombin. However, the molecular mechanisms that regulate antithrombin expression remain poorly understood. Here, we identified Cullin 2 and USP2 (ubiquitin-specific peptidase-2) as novel regulators of antithrombin expression that act by modulating antithrombin ubiquitination. Inhibition of the proteasome caused accumulation of antithrombin and its ubiquitinated forms in HepG2 and SMMC7721 cells. Notably, inhibition of neddylation with MLN4924 suppressed both ubiquitination and degradation of antithrombin, which is recapitulated by silencing of the neddylation enzymes, NAE1, UBA3, and UBE2M, with small interfering RNA (siRNA). We identified Cullin 2 as the interaction partner of antithrombin, and siRNA-mediated Cullin 2 knockdown reduced antithrombin ubiquitination and increased antithrombin protein. We further found that USP2 interacted with antithrombin and regulated antithrombin expression, showing that overexpression of USP2 inhibits the ubiquitination and proteasomal clearance of antithrombin, whereas pharmacological inhibition or siRNA-mediated knockdown of USP2 downregulates antithrombin. Collectively, these results suggest that Cullin 2 E3 ubiquitin ligase and USP2 coordinately regulate antithrombin ubiquitination and degradation. Thus, targeting Cullin 2 and USP2 could be a potential strategy for treatment of hemophilia.
AB - Hemophilia A and B are congenital bleeding disorders caused by a deficiency in pro-coagulant factor VIII or IX that is treated by downregulation of antithrombin. However, the molecular mechanisms that regulate antithrombin expression remain poorly understood. Here, we identified Cullin 2 and USP2 (ubiquitin-specific peptidase-2) as novel regulators of antithrombin expression that act by modulating antithrombin ubiquitination. Inhibition of the proteasome caused accumulation of antithrombin and its ubiquitinated forms in HepG2 and SMMC7721 cells. Notably, inhibition of neddylation with MLN4924 suppressed both ubiquitination and degradation of antithrombin, which is recapitulated by silencing of the neddylation enzymes, NAE1, UBA3, and UBE2M, with small interfering RNA (siRNA). We identified Cullin 2 as the interaction partner of antithrombin, and siRNA-mediated Cullin 2 knockdown reduced antithrombin ubiquitination and increased antithrombin protein. We further found that USP2 interacted with antithrombin and regulated antithrombin expression, showing that overexpression of USP2 inhibits the ubiquitination and proteasomal clearance of antithrombin, whereas pharmacological inhibition or siRNA-mediated knockdown of USP2 downregulates antithrombin. Collectively, these results suggest that Cullin 2 E3 ubiquitin ligase and USP2 coordinately regulate antithrombin ubiquitination and degradation. Thus, targeting Cullin 2 and USP2 could be a potential strategy for treatment of hemophilia.
KW - coagulation
KW - deubiquitinase
KW - neddylation
KW - ubiquitin
UR - http://www.scopus.com/inward/record.url?scp=85111534872&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85111534872&partnerID=8YFLogxK
U2 - 10.1096/fj.202001146RR
DO - 10.1096/fj.202001146RR
M3 - Article
C2 - 34324733
AN - SCOPUS:85111534872
SN - 0892-6638
VL - 35
JO - FASEB Journal
JF - FASEB Journal
IS - 8
M1 - e21800
ER -