scholarly journals Yos9p assists in the degradation of certain nonglycosylated proteins from the endoplasmic reticulum

2011 ◽  
Vol 22 (16) ◽  
pp. 2937-2945 ◽  
Author(s):  
Laura A. Jaenicke ◽  
Holger Brendebach ◽  
Matthias Selbach ◽  
Christian Hirsch
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Ubiquitin Ligase ◽  
Quantitative Proteomics ◽  
Structural Defects ◽  
Sterol Synthesis ◽  
Mutant Variant ◽  
Substrate Spectrum

The HRD ubiquitin ligase recognizes and ubiquitylates proteins of the endoplasmic reticulum that display structural defects. Here, we apply quantitative proteomics to characterize the substrate spectrum of the HRD complex. Among the identified substrates is Erg3p, a glycoprotein involved in sterol synthesis. We characterize Erg3p and demonstrate that the elimination of Erg3p requires Htm1p and Yos9p, two proteins that take part in the glycan-dependent turnover of aberrant proteins. We further show that the HRD ligase also mediates the breakdown of Erg3p and CPY* engineered to lack N-glycans. The degradation of these nonglycosylated substrates is enhanced by a mutant variant of Yos9p that has lost its affinity for oligosaccharides, indicating that Yos9p has a previously unrecognized role in the quality control of nonglycosylated proteins.

scholarly journals Overlapping function of Hrd1 and Ste24 in translocon quality control provides robust channel surveillance

2020 ◽  
Vol 295 (47) ◽  
pp. 16113-16120
Author(s):  
Avery M. Runnebohm ◽  
Kyle A. Richards ◽  
Courtney Broshar Irelan ◽  
Samantha M. Turk ◽  
Halie E. Vitali ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Ubiquitin Ligase ◽  
Biological Membranes ◽  
The Other ◽  
Growth Defect ◽  
Zinc Metalloprotease

Translocation of proteins across biological membranes is essential for life. Proteins that clog the endoplasmic reticulum (ER) translocon prevent the movement of other proteins into the ER. Eukaryotes have multiple translocon quality control (TQC) mechanisms to detect and destroy proteins that persistently engage the translocon. TQC mechanisms have been defined using a limited panel of substrates that aberrantly occupy the channel. The extent of substrate overlap among TQC pathways is unknown. In this study, we found that two TQC enzymes, the ER-associated degradation ubiquitin ligase Hrd1 and zinc metalloprotease Ste24, promote degradation of characterized translocon-associated substrates of the other enzyme in Saccharomyces cerevisiae. Although both enzymes contribute to substrate turnover, our results suggest a prominent role for Hrd1 in TQC. Yeast lacking both Hrd1 and Ste24 exhibit a profound growth defect, consistent with overlapping function. Remarkably, two mutations that mildly perturb post-translational translocation and reduce the extent of aberrant translocon engagement by a model substrate diminish cellular dependence on TQC enzymes. Our data reveal previously unappreciated mechanistic complexity in TQC substrate detection and suggest that a robust translocon surveillance infrastructure maintains functional and efficient translocation machinery.

scholarly journals In Vivo Action of the HRD Ubiquitin Ligase Complex: Mechanisms of Endoplasmic Reticulum Quality Control and Sterol Regulation

2001 ◽  
Vol 21 (13) ◽  
pp. 4276-4291 ◽  
Author(s):  
Richard G. Gardner ◽  
Alexander G. Shearer ◽  
Randolph Y. Hampton
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Ubiquitin Ligase ◽  
Sequence Similarity ◽  
High Specificity ◽  
Specific Sequence ◽  
Ubiquitin Ligase Complex ◽  
Ubiquitin Conjugating Enzyme ◽  
Conjugating Enzyme

ABSTRACT Ubiquitination is used to target both normal proteins for specific regulated degradation and misfolded proteins for purposes of quality control destruction. Ubiquitin ligases, or E3 proteins, promote ubiquitination by effecting the specific transfer of ubiquitin from the correct ubiquitin-conjugating enzyme, or E2 protein, to the target substrate. Substrate specificity is usually determined by specific sequence determinants, or degrons, in the target substrate that are recognized by the ubiquitin ligase. In quality control, however, a potentially vast collection of proteins with characteristic hallmarks of misfolding or misassembly are targeted with high specificity despite the lack of any sequence similarity between substrates. In order to understand the mechanisms of quality control ubiquitination, we have focused our attention on the first characterized quality control ubiquitin ligase, the HRD complex, which is responsible for the endoplasmic reticulum (ER)-associated degradation (ERAD) of numerous ER-resident proteins. Using an in vivo cross-linking assay, we directly examined the association of the separate HRDcomplex components with various ERAD substrates. We have discovered that the HRD ubiquitin ligase complex associates with both ERAD substrates and stable proteins, but only mediates ubiquitin-conjugating enzyme association with ERAD substrates. Our studies with the sterol pathway-regulated ERAD substrate Hmg2p, an isozyme of the yeast cholesterol biosynthetic enzyme HMG-coenzyme A reductase (HMGR), indicated that the HRD complex discerns between a degradation-competent “misfolded” state and a stable, tightly folded state. Thus, it appears that the physiologically regulated, HRD-dependent degradation of HMGR is effected by a programmed structural transition from a stable protein to a quality control substrate.

scholarly journals Aberrant substrate engagement of the ER translocon triggers degradation by the Hrd1 ubiquitin ligase

2012 ◽  
Vol 197 (6) ◽  
pp. 761-773 ◽  
Author(s):  
Eric M. Rubenstein ◽  
Stefan G. Kreft ◽  
Wesley Greenblatt ◽  
Robert Swanson ◽  
Mark Hochstrasser
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Ubiquitin Ligase ◽  
Apolipoprotein B ◽  
Secretory Pathway ◽  
Protein A ◽  
Proteasomal Degradation ◽  
Membrane Localization ◽  
General Role

Little is known about quality control of proteins that aberrantly or persistently engage the endoplasmic reticulum (ER)-localized translocon en route to membrane localization or the secretory pathway. Hrd1 and Doa10, the primary ubiquitin ligases that function in ER-associated degradation (ERAD) in yeast, target distinct subsets of misfolded or otherwise abnormal proteins based primarily on degradation signal (degron) location. We report the surprising observation that fusing Deg1, a cytoplasmic degron normally recognized by Doa10, to the Sec62 membrane protein rendered the protein a Hrd1 substrate. Hrd1-dependent degradation occurred when Deg1-Sec62 aberrantly engaged the Sec61 translocon channel and underwent topological rearrangement. Mutations that prevent translocon engagement caused a reversion to Doa10-dependent degradation. Similarly, a variant of apolipoprotein B, a protein known to be cotranslocationally targeted for proteasomal degradation, was also a Hrd1 substrate. Hrd1 therefore likely plays a general role in targeting proteins that persistently associate with and potentially obstruct the translocon.

scholarly journals Human XTP3-B Forms an Endoplasmic Reticulum Quality Control Scaffold with the HRD1-SEL1L Ubiquitin Ligase Complex and BiP

2008 ◽  
Vol 283 (30) ◽  
pp. 20914-20924 ◽  
Author(s):  
Nobuko Hosokawa ◽  
Ikuo Wada ◽  
Koji Nagasawa ◽  
Tatsuya Moriyama ◽  
Katsuya Okawa ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Ubiquitin Ligase ◽  
Ubiquitin Ligase Complex ◽  
Endoplasmic Reticulum Quality Control

scholarly journals USP13 antagonizes gp78 to maintain functionality of a chaperone in ER-associated degradation

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Yanfen Liu ◽  
Nia Soetandyo ◽  
Jin-gu Lee ◽  
Liping Liu ◽  
Yue Xu ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Substrate Specificity ◽  
Ubiquitin Ligase ◽  
Proteolytic Processing ◽  
Misfolded Proteins ◽  
Physiological Adaptation ◽  
Er Quality Control ◽  
Proteotoxic Stress ◽  
Chaperone Complex

Physiological adaptation to proteotoxic stress in the endoplasmic reticulum (ER) requires retrotranslocation of misfolded proteins into the cytoplasm for ubiquitination and elimination by ER-associated degradation (ERAD). A surprising paradox emerging from recent studies is that ubiquitin ligases (E3s) and deubiquitinases (DUBs), enzymes with opposing activities, can both promote ERAD. Here we demonstrate that the ERAD E3 gp78 can ubiquitinate not only ERAD substrates, but also the machinery protein Ubl4A, a key component of the Bag6 chaperone complex. Remarkably, instead of targeting Ubl4A for degradation, polyubiquitination is associated with irreversible proteolytic processing and inactivation of Bag6. Importantly, we identify USP13 as a gp78-associated DUB that eliminates ubiquitin conjugates from Ubl4A to maintain the functionality of Bag6. Our study reveals an unexpected paradigm in which a DUB prevents undesired ubiquitination to sharpen substrate specificity for an associated ubiquitin ligase partner and to promote ER quality control.

scholarly journals Mechanisms for Rescue of Correctable Folding Defects in CFTRΔF508

2009 ◽  
Vol 20 (18) ◽  
pp. 4059-4069 ◽  
Author(s):  
Diane E. Grove ◽  
Meredith F.N. Rosser ◽  
Hong Yu Ren ◽  
Anjaparavanda P. Naren ◽  
Douglas M. Cyr
Keyword(s):  
Cystic Fibrosis ◽  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Ubiquitin Ligase ◽  
Therapeutic Approach ◽  
Er Quality Control ◽  
Amino Terminal ◽  
Large Pool ◽  
Cellular Environment ◽  
R Domain

Premature degradation of CFTRΔF508 causes cystic fibrosis (CF). CFTRΔF508 folding defects are conditional and folding correctors are being developed as CF therapeutics. How the cellular environment impacts CFTRΔF508 folding efficiency and the identity of CFTRΔF508's correctable folding defects is unclear. We report that inactivation of the RMA1 or CHIP ubiquitin ligase permits a pool of CFTRΔF508 to escape the endoplasmic reticulum. Combined RMA1 or CHIP inactivation and Corr-4a treatment enhanced CFTRΔF508 folding to 3–7-fold greater levels than those elicited by Corr-4a. Some, but not all, folding defects in CFTRΔF508 are correctable. CHIP and RMA1 recognize different regions of CFTR and a large pool of nascent CFTRΔF508 is ubiquitinated by RMA1 before Corr-4a action. RMA1 recognizes defects in CFTRΔF508 related to misassembly of a complex that contains MSD1, NBD1, and the R-domain. Corr-4a acts on CFTRΔF508 after MSD2 synthesis and was ineffective at rescue of ΔF508 dependent folding defects in amino-terminal regions. In contrast, misfolding caused by the rare CF-causing mutation V232D in MSD1 was highly correctable by Corr-4a. Overall, correction of folding defects recognized by RMA1 and/or global modulation of ER quality control has the potential to increase CFTRΔF508 folding and provide a therapeutic approach for CF.

scholarly journals Intramembrane protease RHBDL4 cleaves oligosaccharyltransferase subunits to target them for ER-associated degradation

2019 ◽  
Author(s):  
Julia D. Knopf ◽  
Nina Landscheidt ◽  
Cassandra L. Pegg ◽  
Benjamin L. Schulz ◽  
Nathalie Kühnle ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Quantitative Proteomics ◽  
Proteasomal Degradation ◽  
Rhomboid Protease ◽  
Protein Fragments ◽  
Clearance Mechanism ◽  
Endogenous Substrate ◽  
Erad Pathway ◽  
Substrate Spectrum

AbstractThe Endoplasmic Reticulum (ER)-resident intramembrane rhomboid protease RHBDL4 generates metastable protein fragments and together with the ER-associated degradation (ERAD) machinery provides a clearance mechanism for aberrant and surplus proteins. However, the endogenous substrate spectrum and with that the role of RHBDL4 in physiological ERAD is mainly unknown. Here, we use a substrate trapping approach in combination with quantitative proteomics to identify physiological RHBDL4 substrates. This revealed oligosacharyltransferase (OST) complex subunits such as the catalytic active subunit STT3A as substrates for the RHBDL4-dependent ERAD pathway. RHBDL4-catalyzed cleavage inactivates OST subunits by triggering dislocation into the cytoplasm and subsequent proteasomal degradation. Thereby, RHBDL4 controls the abundance and activity of OST, suggesting a novel link between the ERAD machinery and glycosylation tuning.

scholarly journals Cooperation of mitochondrial and ER factors in quality control of tail-anchored proteins

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Verena Dederer ◽  
Anton Khmelinskii ◽  
Anna Gesine Huhn ◽  
Voytek Okreglak ◽  
Michael Knop ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Ubiquitin Ligase ◽  
Mitochondrial Membrane ◽  
E3 Ubiquitin Ligase ◽  
Outer Mitochondrial Membrane ◽  
Complex Assembly ◽  
Aaa Atpase ◽  
Er Targeting ◽  
Ta Proteins

Tail-anchored (TA) proteins insert post-translationally into the endoplasmic reticulum (ER), the outer mitochondrial membrane (OMM) and peroxisomes. Whereas the GET pathway controls ER-targeting, no dedicated factors are known for OMM insertion, posing the question of how accuracy is achieved. The mitochondrial AAA-ATPase Msp1 removes mislocalized TA proteins from the OMM, but it is unclear, how Msp1 clients are targeted for degradation. Here we screened for factors involved in degradation of TA proteins mislocalized to mitochondria. We show that the ER-associated degradation (ERAD) E3 ubiquitin ligase Doa10 controls cytoplasmic level of Msp1 clients. Furthermore, we identified the uncharacterized OMM protein Fmp32 and the ectopically expressed subunit of the ER-mitochondria encounter structure (ERMES) complex Gem1 as native clients for Msp1 and Doa10. We propose that productive localization of TA proteins to the OMM is ensured by complex assembly, while orphan subunits are extracted by Msp1 and eventually degraded by Doa10.

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