Proteomic characterization of endogenous substrates of mammalian ubiquitin ligase Hrd1

Cell & Bioscience
Yilin YeTing Zhang

Abstract

Endoplasmic reticulum (ER)-associated degradation (ERAD) regulates protein homeostasis in the secretory pathway by targeting misfolded or unassembled proteins for degradation by the proteasome. Hrd1 is a conserved multi-spanning membrane bound ubiquitin ligase required for ubiquitination of many aberrant ER proteins, but few endogenous substrates of Hrd1 have been identified to date. Using a SILAC-based quantitative proteomic approach combined with CRISPR-mediated gene silencing, we searched for endogenous physiological substrates of Hrd1. We used RNA microarray, immunoblotting, cycloheximide chase combined with chemical genetics to define the role of Hrd1 in regulating the stability of endogenous ERAD substrates. We identified 58 proteins whose levels are consistently upregulated in Hrd1 null HEK293 cells. Many of these proteins function in pathways involved in stress adaptation or immune surveillance. We validated OS9, a lectin required for ERAD of glycoproteins as a highly upregulated protein in Hrd1 deficient cells. Moreover, the abundance of OS9 is inversely correlated with Hrd1 level in clinical synovium samples isolated from osteoarthritis and rheumatoid arthritis patients. Intriguingly, immunoblotting detects two OS9 va...Continue Reading

References

Sep 17, 2003·Genes & Development·Tetsuya AmanoToshihiro Nakajima
Apr 14, 2004·The Journal of Cell Biology·Shilpa Vashist, Davis T W Ng
May 19, 2004·Genes to Cells : Devoted to Molecular & Cellular Mechanisms·Osamu HoriSatoshi Ogawa
Oct 18, 2006·The Journal of Cell Biology·Britta MuellerHidde L Ploegh
Dec 30, 2008·Molecular Cell·Erin M QuanJonathan S Weissman
Nov 16, 2010·Current Opinion in Cell Biology·Riccardo Bernasconi, Maurizio Molinari
Sep 14, 2011·Molecular and Cellular Biology·Yoshiki TsuchiyaAkira Kobayashi
Oct 12, 2011·The Journal of Biological Chemistry·Kimberly A LeeJohn R Doedens
Nov 26, 2011·Science·Melanie H SmithJonathan S Weissman
Oct 26, 2012·Molecular Biology of the Cell·Ryan E TylerRon R Kopito
Jul 31, 2013·Nature Chemical Biology·Paola MagnaghiAntonella Isacchi
Aug 10, 2013·Proceedings of the National Academy of Sciences of the United States of America·Marian L BurrPaul J Lehner
Dec 25, 2013·The Journal of Biological Chemistry·Chih-Hsiang HuangXin Chen
Jan 24, 2014·Proceedings of the National Academy of Sciences of the United States of America·Shengyi SunLing Qi
Mar 19, 2014·The Journal of Cell Biology·Annamaria RuggianoPedro Carvalho
Apr 5, 2014·Nature Structural & Molecular Biology·John C Christianson, Yihong Ye
Oct 28, 2016·Nature Cell Biology·Fiorenza FumagalliMaurizio Molinari
Feb 20, 2018·The Journal of Clinical Investigation·Geun Hyang KimLing Qi

❮ Previous
Next ❯

Citations

Oct 31, 2019·Journal of Oncology·Gábor TaxPietro Roversi
Aug 28, 2020·International Journal of Molecular Sciences·Alekya S TanikellaJulia Thom Oxford

❮ Previous
Next ❯

Methods Mentioned

BETA
ubiquitination
protein folding
Protein Assay
electrophoresis
X-ray

Software Mentioned

Affymetrix
MaxQuant
Odyssey
ImageGauge

Related Concepts

Related Feeds

CRISPR Ribonucleases Deactivation

CRISPR-Cas system enables the editing of genes to create or correct mutations. This feed focuses on mechanisms that underlie deactivation of CRISPR ribonucleases. Here is the latest research.

Aneurysm

Aneurysms are outward distensions or bulges that occurs in a weakened wall of blood vessels. Discover the latest research on aneurysms here.

CRISPR (general)

Clustered regularly interspaced short palindromic repeats (CRISPR) are DNA sequences in the genome that are recognized and cleaved by CRISPR-associated proteins (Cas). CRISPR-Cas system enables the editing of genes to create or correct mutations. Discover the latest research on CRISPR here.

Aortic Aneurysm

An aortic aneurysm is the weakening and bulging of the blood vessel wall in the aorta. This causes dilatation of the aorta, which is usually asymptomatic but carries the risk of rupture and hemorrhage. Find the latest research on aortic aneurysms here.