scholarly journals The degradation pathway of a model misfolded protein is determined by aggregation propensity

2018 ◽  
Vol 29 (12) ◽  
pp. 1422-1434 ◽  
Author(s):  
Zhihao Sun ◽  
Jeffrey L. Brodsky
Keyword(s):  
Quality Control ◽  
Secretory Pathway ◽  
Multivesicular Body ◽  
Degradation Pathway ◽  
Misfolded Proteins ◽  
Protein Homeostasis ◽  
Substrate Selection ◽  
Er Quality Control ◽  
Aggregation Propensity ◽  
Novel Model

Protein homeostasis in the secretory pathway is maintained by a hierarchy of quality control checkpoints, including endoplasmic reticulum–associated degradation (ERAD), which leads to the destruction of misfolded proteins in the ER, as well as post-ER proteolysis. Although most aberrant proteins are degraded by ERAD, some misfolded proteins escape the ER and are degraded instead by lysosomal/vacuolar proteases. To date, it remains unclear how misfolded membrane proteins are selected for these different fates. Here we designed a novel model substrate, SZ*, to investigate how substrate selection is mediated in yeast. We discovered that SZ* is degraded by both the proteasome and vacuolar proteases, the latter of which occurs after ER exit and requires the multivesicular body pathway. By interrogating how various conditions affect the fate of SZ*, we also discovered that heat-shock and substrate overexpression increase ERAD targeting. These conditions also increase substrate aggregation. We next found that aggregation of the membrane-free misfolded domain in SZ* is concentration dependent, and fusion of this misfolded domain to a post-ER quality control substrate instead targets the substrate for ERAD. Our data indicate that a misfolded membrane protein with a higher aggregation propensity is preferentially retained in the ER and targeted for ERAD.

scholarly journals Misfolded proteins are sorted by a sequential checkpoint mechanism of ER quality control

2004 ◽  
Vol 165 (1) ◽  
pp. 41-52 ◽  
Author(s):  
Shilpa Vashist ◽  
Davis T.W. Ng
Keyword(s):  
Quality Control ◽  
Membrane Proteins ◽  
Control Mechanism ◽  
Degradation Pathway ◽  
Misfolded Proteins ◽  
Er Quality Control ◽  
Golgi Transport ◽  
Test Face ◽  
Folded Proteins ◽  
Quality Control Mechanism

Misfolded proteins retained in the endoplasmic reticulum (ER) are degraded by the ER-associated degradation pathway. The mechanisms used to sort them from correctly folded proteins remain unclear. Analysis of substrates with defined folded and misfolded domains has revealed a system of sequential checkpoints that recognize topologically distinct domains of polypeptides. The first checkpoint examines the cytoplasmic domains of membrane proteins. If a lesion is detected, it is retained statically in the ER and rapidly degraded without regard to the state of its other domains. Proteins passing this test face a second checkpoint that monitors domains localized in the ER lumen. Proteins detected by this pathway are sorted from folded proteins and degraded by a quality control mechanism that requires ER-to-Golgi transport. Although the first checkpoint is obligatorily directed at membrane proteins, the second monitors both soluble and membrane proteins. Our data support a model whereby “properly folded” proteins are defined biologically as survivors that endure a series of distinct checkpoints.

scholarly journals A post-ER degradation pathway that relies on protease-dependent internalization from the vacuolar membrane

2019 ◽  
Author(s):  
Leticia Lemus ◽  
Zrinka Matić ◽  
Veit Goder
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Proteolytic Activity ◽  
Secretory Pathway ◽  
Model Organism ◽  
Degradation Pathway ◽  
Vacuolar Membrane ◽  
List Type ◽  
Er Quality Control

SummaryNewly synthesized proteins of the secretory pathway are quality-controlled inside the endoplasmic reticulum (ER) and, if not properly folded, are retained. An exception are glycosylphosphatidylinositol-anchored proteins (GPI-APs) which can leave the ER even when misfolded and are routed to the vacuole/lysosome for degradation by largely unknown mechanisms linked to post-ER quality control. Using yeast as model organism, we show that Gas1*, an ER-exported misfolded GPI-AP, is diverted from the secretory pathway to endosomes for transport to the vacuole. However, Gas1* is not sorted into endosomal intraluminal vesicles but internalizes directly from the vacuolar membrane. There, the vacuolar protease Pep4, but not any other known vacuolar protease, is required for Gas1* internalization. Our data reveal novel and unexpected mechanisms for invaginations from the vacuolar membrane.HighlightsER-exited misfolded GPI-anchored proteins are routed to the vacuole via endosomes but do not internalize into intraluminal vesiclesInternalization occurs directly from the vacuolar membrane into intravacuolar mobile structuresInternalization from the vacuolar membrane depends on the proteolytic activity of the vacuolar protease Pep4

scholarly journals Limited ER quality control for GPI-anchored proteins

2016 ◽  
Vol 213 (6) ◽  
pp. 693-704 ◽  
Author(s):  
Natalia Sikorska ◽  
Leticia Lemus ◽  
Auxiliadora Aguilera-Romero ◽  
Javier Manzano-Lopez ◽  
Howard Riezman ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Protein Folding ◽  
Misfolded Proteins ◽  
Control Mechanisms ◽  
Gpi Anchor ◽  
Er Quality Control ◽  
Entire Class ◽  
Er Export ◽  
Export Signal

Endoplasmic reticulum (ER) quality control mechanisms target terminally misfolded proteins for ER-associated degradation (ERAD). Misfolded glycophosphatidylinositol-anchored proteins (GPI-APs) are, however, generally poor ERAD substrates and are targeted mainly to the vacuole/lysosome for degradation, leading to predictions that a GPI anchor sterically obstructs ERAD. Here we analyzed the degradation of the misfolded GPI-AP Gas1* in yeast. We could efficiently route Gas1* to Hrd1-dependent ERAD and provide evidence that it contains a GPI anchor, ruling out that a GPI anchor obstructs ERAD. Instead, we show that the normally decreased susceptibility of Gas1* to ERAD is caused by canonical remodeling of its GPI anchor, which occurs in all GPI-APs and provides a protein-independent ER export signal. Thus, GPI anchor remodeling is independent of protein folding and leads to efficient ER export of even misfolded species. Our data imply that ER quality control is limited for the entire class of GPI-APs, many of them being clinically relevant.

scholarly journals Reglucosylation by UDP-glucose:glycoprotein glucosyltransferase 1 delays glycoprotein secretion but not degradation

2015 ◽  
Vol 26 (3) ◽  
pp. 390-405 ◽  
Author(s):  
Abla Tannous ◽  
Nishant Patel ◽  
Taku Tamura ◽  
Daniel N. Hebert
Keyword(s):  
Quality Control ◽  
Secretory Pathway ◽  
Selection Process ◽  
Control Process ◽  
Degradation Pathway ◽  
Carbohydrate Binding ◽  
Wild Type ◽  
Mutant Proteins ◽  
Quality Control Process ◽  
Glycoprotein Secretion

UDP-glucose:glycoprotein glucosyltransferase 1 (UGT1) is a central quality control gatekeeper in the mammalian endoplasmic reticulum (ER). The reglucosylation of glycoproteins supports their rebinding to the carbohydrate-binding ER molecular chaperones calnexin and calreticulin. A cell-based reglucosylation assay was used to investigate the role of UGT1 in ER protein surveillance or the quality control process. UGT1 was found to modify wild-type proteins or proteins that are expected to eventually traffic out of the ER through the secretory pathway. Trapping of reglucosylated wild-type substrates in their monoglucosylated state delayed their secretion. Whereas terminally misfolded substrates or off-pathway proteins were most efficiently reglucosylated by UGT1, the trapping of these mutant substrates in their reglucosylated or monoglucosylated state did not delay their degradation by the ER-associated degradation pathway. This indicated that monoglucosylated mutant proteins were actively extracted from the calnexin/calreticulin binding-reglucosylation cycle for degradation. Therefore trapping proteins in their monoglucosylated state was sufficient to delay their exit to the Golgi but had no effect on their rate of degradation, suggesting that the degradation selection process progressed in a dominant manner that was independent of reglucosylation and the glucose-containing A-branch on the substrate glycans.

scholarly journals ER stress causes widespread protein aggregation and prion formation

2017 ◽  
Vol 216 (8) ◽  
pp. 2295-2304 ◽  
Author(s):  
Norfadilah Hamdan ◽  
Paraskevi Kritsiligkou ◽  
Chris M. Grant
Keyword(s):  
Protein Aggregation ◽  
Er Stress ◽  
Secretory Pathway ◽  
Cellular Protein ◽  
Protein Homeostasis ◽  
Er Quality Control ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Er Homeostasis

Disturbances in endoplasmic reticulum (ER) homeostasis create a condition termed ER stress. This activates the unfolded protein response (UPR), which alters the expression of many genes involved in ER quality control. We show here that ER stress causes the aggregation of proteins, most of which are not ER or secretory pathway proteins. Proteomic analysis of the aggregated proteins revealed enrichment for intrinsically aggregation-prone proteins rather than proteins which are affected in a stress-specific manner. Aggregation does not arise because of overwhelming proteasome-mediated degradation but because of a general disruption of cellular protein homeostasis. We further show that overexpression of certain chaperones abrogates protein aggregation and protects a UPR mutant against ER stress conditions. The onset of ER stress is known to correlate with various disease processes, and our data indicate that widespread amorphous and amyloid protein aggregation is an unanticipated outcome of such stress.

scholarly journals Approaches to imaging unfolded secretory protein stress in living cells

2014 ◽  
Vol 1 (1) ◽  
Author(s):  
Patrick Lajoie ◽  
Elena N. Fazio ◽  
Erik L. Snapp
Keyword(s):  
Quality Control ◽  
Signaling Pathways ◽  
Protein Function ◽  
Secretory Pathway ◽  
Transcriptional Response ◽  
Secretory Protein ◽  
Model Systems ◽  
Er Quality Control ◽  
Unfolded Protein ◽  
Point Of Entry

AbstractThe endoplasmic reticulum (ER) is the point of entry of proteins into the secretory pathway. Nascent peptides interact with the ER quality control machinery that ensures correct folding of the nascent proteins. Failure to properly fold proteins can lead to loss of protein function and cytotoxic aggregation of misfolded proteins that can lead to cell death. To cope with increases in the ER unfolded secretory protein burden, cells have evolved the Unfolded Protein Response (UPR). The UPR is the primary signaling pathway that monitors the state of the ER folding environment. When the unfolded protein burden overwhelms the capacity of the ER quality control machinery, a state termed ER stress, sensor proteins detect accumulation of misfolded peptides and trigger the UPR transcriptional response. The UPR, which is conserved from yeast to mammals, consists of an ensemble of complex signaling pathways that aims at adapting the ER to the new misfolded protein load. To determine how different factors impact the ER folding environment, various tools and assays have been developed. In this review, we discuss recent advances in live cell imaging reporters and model systems that enable researchers to monitor changes in the unfolded secretory protein burden and activation of the UPR and its associated signaling pathways.

scholarly journals A Striking Quality Control Subcompartment in Saccharomyces cerevisiae: The Endoplasmic Reticulum-associated Compartment

2004 ◽  
Vol 15 (2) ◽  
pp. 908-921 ◽  
Author(s):  
Gregory Huyer ◽  
Gaby L. Longsworth ◽  
Deborah L. Mason ◽  
Monica P. Mallampalli ◽  
J. Michael McCaffery ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Secretory Pathway ◽  
Secretory Protein ◽  
Cellular Functions ◽  
Er Quality Control ◽  
Fluorescence And Electron Microscopy ◽  
The Unfolded Protein Response ◽  
Mutant Forms

The folding of nascent secretory and membrane proteins is monitored by the endoplasmic reticulum (ER) quality control system. Misfolded proteins are retained in the ER and can be removed by ER-associated degradation. As a model for the ER quality control of multispanning membrane proteins in yeast, we have been studying mutant forms of Ste6p. Here, we identify mislocalized mutant forms of Ste6p that induce the formation of, and localize to, prominent structures that are absent in normal cells. We have named these structures ER-associated compartments (ERACs), based on their juxtaposition to and connection with the ER, as observed by fluorescence and electron microscopy. ERACs comprise a network of tubulo-vesicular structures that seem to represent proliferated ER membranes. Resident ER lumenal and membrane proteins are present in ERACs in addition to their normal ER localization, suggesting there is no barrier for their entry into ERACs. However, the forms of Ste6p in ERACs are excluded from the ER and do not enter the secretory pathway; instead, they are ultimately targeted for ER-associated degradation. The presence of ERACs does not adversely affect secretory protein traffic through the ER and does not lead to induction of the unfolded protein response. We propose that ERACs may be holding sites to which misfolded membrane proteins are specifically diverted so as not to interfere with normal cellular functions. We discuss the likelihood that related ER membrane proliferations that form in response to certain other mutant or unassembled membrane proteins may be substantially similar to ERACs.

scholarly journals Role of Sec61p in the ER-associated degradation of short-lived transmembrane proteins

2008 ◽  
Vol 181 (7) ◽  
pp. 1095-1105 ◽  
Author(s):  
Daniel C. Scott ◽  
Randy Schekman
Keyword(s):  
Degradation Process ◽  
Ubiquitin Proteasome System ◽  
Degradation Pathway ◽  
Specific Protein ◽  
Cysteine Residue ◽  
Misfolded Proteins ◽  
Er Quality Control ◽  
Substrate Protein ◽  
Ubiquitin Proteasome ◽  
Protein Components

Misfolded proteins in the endoplasmic reticulum (ER) are identified and degraded by the ER-associated degradation pathway (ERAD), a component of ER quality control. In ERAD, misfolded proteins are removed from the ER by retrotranslocation into the cytosol where they are degraded by the ubiquitin–proteasome system. The identity of the specific protein components responsible for retrotranslocation remains controversial, with the potential candidates being Sec61p, Der1p, and Doa10. We show that the cytoplasmic N-terminal domain of a short-lived transmembrane ERAD substrate is exposed to the lumen of the ER during the degradation process. The addition of N-linked glycan to the N terminus of the substrate is prevented by mutation of a specific cysteine residue of Sec61p, as well as a specific cysteine residue of the substrate protein. We show that the substrate protein forms a disulfide-linked complex to Sec61p, suggesting that at least part of the retrotranslocation process involves Sec61p.

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 Protein quality control in the secretory pathway

2019 ◽  
Vol 218 (10) ◽  
pp. 3171-3187 ◽  
Author(s):  
Zhihao Sun ◽  
Jeffrey L. Brodsky
Keyword(s):  
Quality Control ◽  
Protein Folding ◽  
Secretory Pathway ◽  
Protein Quality ◽  
Protein Quality Control ◽  
Misfolded Proteins ◽  
Unfolded Protein ◽  
Protein Biogenesis ◽  
The Unfolded Protein Response ◽  
Protein Response

Protein folding is inherently error prone, especially in the endoplasmic reticulum (ER). Even with an elaborate network of molecular chaperones and protein folding facilitators, misfolding can occur quite frequently. To maintain protein homeostasis, eukaryotes have evolved a series of protein quality-control checkpoints. When secretory pathway quality-control pathways fail, stress response pathways, such as the unfolded protein response (UPR), are induced. In addition, the ER, which is the initial hub of protein biogenesis in the secretory pathway, triages misfolded proteins by delivering substrates to the proteasome or to the lysosome/vacuole through ER-associated degradation (ERAD) or ER-phagy. Some misfolded proteins escape the ER and are instead selected for Golgi quality control. These substrates are targeted for degradation after retrieval to the ER or delivery to the lysosome/vacuole. Here, we discuss how these guardian pathways function, how their activities intersect upon induction of the UPR, and how decisions are made to dispose of misfolded proteins in the secretory pathway.

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