Degradation of Proteins from the ER of S. cerevisiae Requires an Intact Unfolded Protein Response Pathway

Abstract

To dissect the requirements of membrane protein degradation from the ER, we expressed the mouse major histocompatibility complex class I heavy chain H-2K(b) in yeast. Like other proteins degraded from the ER, unassembled H-2K(b) heavy chains are not transported to the Golgi but are degraded in a proteasome-dependent manner. The overexpression of H-2K(b) heavy chains induces the unfolded protein response (UPR). In yeast mutants unable to mount the UPR, H-2K(b) heavy chains are greatly stabilized. This defect in degradation is suppressed by the expression of the active form of Hac1p, the transcription factor that upregulates UPR-induced genes. These results indicate that induction of the UPR is required for the degradation of protein substrates from the ER.

Decreased ER-associated degradation of α-TCR induced by Grp78 depletion with the SubAB cytotoxin

“…This agrees with the finding that treatment of human fibroblasts with SubAB during the early phase of CMV infection downregulates the expression of viral proteins even as late as 96 h after addition of the toxin (Buchkovich et al, 2008). Many reports have indicated that ERAD components are upregulated as part of UPR (Casagrande et al, 2000;Friedlander et al, 2000). However, we have not observed an upregulation of proteins involved in ERAD, such as proteasome subunits, p97/VCP or the channel forming protein derlin 1 even after 16 h of incubation with SubAB.…”

“…ERAD constitutively counteracts ER stress, eliminating misfolded proteins from the ER, however UPR induces its further activation through upregulation of its components (Casagrande et al, 2000;Friedlander et al, 2000). We therefore expected that treatment with SubAB would accelerate ERAD of αTCR.…”

“…Since ER stress is usually associated with increased ER-associated degradation (ERAD) of proteins (Casagrande et al, 2000;Friedlander et al, 2000)., we also looked at different components of the ubiquitin and proteasome system (UPS). The levels of polyubiquitinated proteins were similar in all three cell lines and were not affected by either wild type or mutant SubAB.…”

“…Thus, UPR is an adaptative response allowing the cell to cope with ER stress: However, when UPR is persistent, it leads to apoptosis via transcriptional induction of CHOP/GADD153, TRAF-mediated activation of cJUN N-terminal kinase, and/ or the activation of caspase-12/4 (Boyce and Yuan, 2006;Wu and Kaufman, 2006). ERAD constitutively counteracts ER stress, eliminating misfolded proteins from the ER, but it is further activated as part of UPR (Casagrande et al, 2000;Friedlander et al, 2000). Expression of individual TCR subunits in cell lines which endogenously do not express TCR have served in numerous studies to resolve the molecular determinants for rapid degradation of free TCR subunits (Bonifacino et al, 1989;Bonifacino et al, 1990;Fang et al, 2001;Fayadat and Kopito, 2003;Huppa and Ploegh, 1997;Lenk et al, 2002;Tiwari and Weissman, 2001;Travers et al, 2000;Wileman et al, 1990;Wileman et al, 1993;Yang et al, 1998;Yu et al, 1997;Yu and Kopito, 1999).…”

Proteins of the endoplasmic-reticulum-associated degradation pathway: domain detection and function prediction

“…The transmembrane kinase and endoribonuclease Ire1p is a key member of the latter pathway, since deletion of its gene impairs protein degradation [53]. Ire1p spans the ER membrane with its N-terminal portion within the lumen, and with its catalytic kinase and endoribonuclease domains in the cytosol or nucleus.…”