scholarly journals Endoplasmic Reticulum Quality Control of Oligomeric Membrane Proteins: Topogenic Determinants Involved in the Degradation of the Unassembled Na,K-ATPase α Subunit and in Its Stabilization by β Subunit Assembly

2000 ◽  
Vol 11 (5) ◽  
pp. 1657-1672 ◽  
Author(s):  
Pascal Béguin ◽  
Udo Hasler ◽  
Olivier Staub ◽  
Käthi Geering
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Membrane Insertion ◽  
Membrane Domains ◽  
Related Factors ◽  
Er Quality Control ◽  
Α Subunit ◽  
Subunit Assembly ◽  
Α Subunits

The molecular nature of determinants that mediate degradation of unassembled, polytopic subunits of oligomeric membrane proteins and their stabilization after partner subunit assembly is largely unknown. Expressing truncated Na,K-ATPase α subunits alone or together with β subunits, we find that in unassembled α subunits neither the four N-terminal transmembrane segments acting as efficient alternating signal anchor–stop transfer sequences nor the large, central cytoplasmic loop exposes any degradation signal, whereas poor membrane insertion efficiency of C-terminal membrane domains M5, M7, and M9 coincides with the transient exposure of degradation signals to the cytoplasmic side. β assembly with an α domain comprising at least D902 up to Y910 in the extracytoplasmic M7/M8 loop is necessary to stabilize Na,K-ATPase α subunits by favoring M7/M8 membrane pair formation and by protecting a degradation signal recognized from the endoplasmic reticulum (ER) lumenal side. Thus our results suggest that ER degradation of Na,K-ATPase α subunits is 1) mainly mediated by folding defects caused by inefficient membrane insertion of certain membrane domains, 2) a multistep process, which involves proteolytic and/or chaperone components acting from the ER lumenal side in addition to cytosolic, proteasome-related factors, and 3) prevented by partner subunit assembly because of direct protection and retrieval of degradation signals from the cytoplasm to the ER lumenal side. These results likely represent a paradigm for the ER quality control of unassembled, polytopic subunits of oligomeric membrane proteins.

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 Endoplasmic Reticulum Quality Control of Unassembled Iron Transporter Depends on Rer1p-mediated Retrieval from the Golgi

2004 ◽  
Vol 15 (3) ◽  
pp. 1417-1424 ◽  
Author(s):  
Miyuki Sato ◽  
Ken Sato ◽  
Akihiko Nakano
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Model System ◽  
Er Quality Control ◽  
Iron Transporter ◽  
Er Retention ◽  
Endoplasmic Reticulum Quality Control ◽  
Sorting Receptor

Endoplasmic reticulum (ER) quality control is a conserved process by which misfolded or unassembled proteins are selectively retained in the endoplasmic reticulum (ER). Failure in oligomerization of multisubunit membrane proteins is one of the events that triggers ER quality control. The transmembrane domains (TMDs) of unassembled subunits are determinants of ER retention in many cases, although the mechanism of the TMD-mediated sorting of unassembled subunits remains elusive. We studied a yeast iron transporter complex on the cell surface as a new model system for ER quality control. When Fet3p, a transmembrane subunit, is not assembled with the other membrane subunit, Ftr1p, unassembled Fet3p is exclusively localized to the ER at steady state. The TMD of Fet3p contains a determinant for this process. However, pulse-chase analysis and in vitro budding assays indicate that unassembled Fet3p rapidly escapes from the ER. Furthermore, Rer1p, a retrieval receptor for ER-resident membrane proteins in the Golgi, is responsible for the TMD-dependent ER retrieval of unassembled Fet3p. These findings provide clear evidence that the ER quality control of unassembled membrane proteins can be achieved by retrieval from the Golgi and that Rer1p serves as a specific sorting receptor in this process.

scholarly journals Modularity of the Hrd1 ERAD complex underlies its diverse client range

2010 ◽  
Vol 188 (5) ◽  
pp. 707-716 ◽  
Author(s):  
Kazue Kanehara ◽  
Wei Xie ◽  
Davis T.W. Ng
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Protein Folding ◽  
Membrane Proteins ◽  
Secretory Protein ◽  
Modular Organization ◽  
Control Mechanisms ◽  
Core Complex ◽  
Er Quality Control ◽  
Folding State

Secretory protein folding is monitored by endoplasmic reticulum (ER) quality control mechanisms. Misfolded proteins are retained and targeted to ER-associated degradation (ERAD) pathways. At their core are E3 ubiquitin ligases, which organize factors that recognize, ubiquitinate, and translocate substrates. Of these, we report that the Hrd1 complex manages three distinct substrate classes. A core complex is required for all classes and is sufficient for some membrane proteins. The accessory factors Usa1p and Der1p adapt the complex to process luminal substrates. Their integration is sufficient to process molecules bearing glycan-independent degradation signals. The presence of Yos9p extends the substrate range by mediating the recognition of glycan-based degradation signals. This modular organization enables the Hrd1 complex to recognize topologically diverse substrates. The Hrd1 system does not directly evaluate the folding state of polypeptides. Instead, it does so indirectly, by recognizing specific embedded signals displayed upon misfolding.

scholarly journals Single, context-specific glycans can target misfolded glycoproteins for ER-associated degradation

2005 ◽  
Vol 169 (1) ◽  
pp. 73-82 ◽  
Author(s):  
Eric D. Spear ◽  
Davis T.W. Ng
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Secretory Protein ◽  
Carboxypeptidase Y ◽  
Er Quality Control ◽  
Systematic Analysis ◽  
Proteinase A ◽  
Context Specific ◽  
The Relationship ◽  
Erad Pathway

The endoplasmic reticulum (ER) maintains an environment essential for secretory protein folding. Consequently, the premature transport of polypeptides would be harmful to the cell. To avert this scenario, mechanisms collectively termed “ER quality control” prevent the transport of nascent polypeptides until they properly fold. Irreversibly misfolded molecules are sorted for disposal by the ER-associated degradation (ERAD) pathway. To better understand the relationship between quality control and ERAD, we studied a new misfolded variant of carboxypeptidase Y (CPY). The molecule was recognized and retained by ER quality control but failed to enter the ERAD pathway. Systematic analysis revealed that a single, specific N-linked glycan of CPY was required for sorting into the pathway. The determinant is dependent on the putative lectin-like receptor Htm1/Mnl1p. The discovery of a similar signal in misfolded proteinase A supported the generality of the mechanism. These studies show that specific signals embedded in glycoproteins can direct their degradation if they fail to fold.

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 Endoplasmic reticulum chaperones participate in human cytomegalovirus US2-mediated degradation of class I major histocompatibility complex molecules

2008 ◽  
Vol 89 (5) ◽  
pp. 1122-1130 ◽  
Author(s):  
Kristina Oresic ◽  
Domenico Tortorella
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Major Histocompatibility Complex ◽  
Human Cytomegalovirus ◽  
Class I ◽  
Cell Surface Expression ◽  
Major Histocompatibility ◽  
Er Quality Control ◽  
Heavy Chains ◽  
Histocompatibility Complex

Inhibition of cell-surface expression of major histocompatibility complex class I molecules by human cytomegalovirus (HCMV, a β-herpesvirus) promotes escape from recognition by CD8+ cytotoxic T cells. The HCMV US2 and US11 gene products induce class I downregulation during the early phase of HCMV infection by facilitating the degradation of class I heavy chains. The HCMV proteins promote the transport of the class I heavy chains across the endoplasmic reticulum (ER) membrane into the cytosol by a process referred to as ‘dislocation’, which is then followed by proteasome degradation. This process has striking similarities to the degradation of misfolded ER proteins mediated by ER quality control. Even though the major steps of the dislocation reaction have been characterized, the cellular proteins, specifically the ER chaperones involved in targeting class I for dislocation, have not been fully delineated. To elucidate the chaperones involved in HCMV-mediated class I dislocation, we utilized a chimeric class I heavy chain with an affinity tag at its carboxy terminus. Interestingly, US2 but not US11 continued to target the class I chimera for destruction, suggesting a structural limitation for US11-mediated degradation. Association studies in US2 cells and in cells that express a US2 mutant, US2–186HA, revealed that class I specifically interacts with calnexin, BiP and calreticulin. These findings demonstrate that US2-mediated class I destruction utilizes specific chaperones to facilitate class I dislocation. The data suggest a more general model in which the chaperones that mediate protein folding may also function during ER quality control to eliminate aberrant ER proteins.

scholarly journals Exposure of bipartite hydrophobic signal triggers nuclear quality control of Ndc10 at the endoplasmic reticulum/nuclear envelope

2011 ◽  
Vol 22 (24) ◽  
pp. 4726-4739 ◽  
Author(s):  
Noa Furth ◽  
Or Gertman ◽  
Ayala Shiber ◽  
Omri S. Alfassy ◽  
Itamar Cohen ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Protein Folding ◽  
Nuclear Envelope ◽  
Reverse Genetics ◽  
Point Mutations ◽  
Hydrophobic Surface ◽  
Structural Features ◽  
Er Quality Control ◽  
C Terminus

Proper functioning of the protein-folding quality control network depends on the network's ability to discern diverse structural perturbations to the native states of its protein substrates. Despite the centrality of the detection of misfolded states to cell home­ostasis, very little is known about the exact sequence and structural features that mark a protein as being misfolded. To investigate these features, we studied the requirements for the degradation of the yeast kinetochore protein Ndc10p. Mutant Ndc10p is a substrate of a protein-folding quality control pathway mediated by the E3 ubiquitin (Ub) ligase Doa10p at the endoplasmic reticulum (ER)/nuclear envelope membrane. Analysis of Ndc10p mutant derivatives, employing a reverse genetics approach, identified an autonomous quality control–associated degradation motif near the C-terminus of the protein. This motif is composed of two indispensable hydrophobic elements: a hydrophobic surface of an amphipathic helix and a loosely structured hydrophobic C-terminal tail. Site-specific point mutations expose these elements, triggering ubiquitin-mediated and HSP70 chaperone–dependent degradation of Ndc10p. These findings substantiate the ability of the ER quality control system to recognize subtle perturbation(s) in the native structure of a nuclear protein.

scholarly journals Analysis of Quality Control Substrates in Distinct Cellular Compartments Reveals a Unique Role for Rpn4p in Tolerating Misfolded Membrane Proteins

2009 ◽  
Vol 20 (3) ◽  
pp. 1006-1019 ◽  
Author(s):  
Meredith Boyle Metzger ◽  
Susan Michaelis
Keyword(s):  
Quality Control ◽  
Membrane Proteins ◽  
Transcriptional Response ◽  
Transcriptional Responses ◽  
Er Quality Control ◽  
Yeast Viability ◽  
Severe Impairment ◽  
Proteasomal Genes ◽  
The Unfolded Protein Response ◽  
Cellular Compartments

ER quality control (ERQC) prevents the exit of misfolded secretory and membrane proteins from the ER. A critical aspect of ERQC is a transcriptional response called the unfolded protein response (UPR), which up-regulates genes that enable cells to cope with misfolded, ER-retained proteins. In this study, we compare the transcriptional responses in yeast resulting from the acute expression of misfolded proteins residing in three different cellular compartments (the ER lumen, membrane, and cytosol), and find that each elicits a distinct transcriptional response. The classical UPR response, here-designated UPR-L, is induced by the ER lumenal misfolded protein, CPY*. The UPR-Cyto response is induced by the cytosolic protein, VHL-L158P, and is characterized by a rapid, transient induction of cytosolic chaperones similar to the heat-shock response. In contrast, the misfolded membrane protein with a cystolic lesion, Ste6p*, elicits a unique response designated UPR-M/C, characterized by the modest induction of >20 genes regulated by Rpn4p, an activator of proteasomal genes. Independently, we identified several genes required for yeast viability during UPR-M/C stress, but not UPR-L or UPR-Cyto stress. Among these is RPN4, highlighting the importance of the Rpn4p-dependent response in tolerating UPR-M/C stress. Further analysis suggests the requirement for Rpn4p reflects severe impairment of the proteasome by UPR-M/C stress.

scholarly journals Brucella effector hijacks endoplasmic reticulum quality control machinery to prevent premature egress

2019 ◽  
Author(s):  
Jean-Baptiste Luizet ◽  
Julie Raymond ◽  
Thais Lourdes Santos Lacerda ◽  
Magali Bonici ◽  
Frédérique Lembo ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Er Quality Control ◽  
Unfolded Protein ◽  
Type Iv ◽  
Endoplasmic Reticulum Quality Control ◽  
Fine Regulation ◽  
Infected Cells ◽  
The Unfolded Protein Response ◽  
Protein Response

AbstractPerturbation of endoplasmic reticulum (ER) functions can have critical consequences for cellular homeostasis. An elaborate surveillance system known as ER quality control (ERQC) ensures that only correctly assembled proteins reach their destination. Persistence of misfolded or improperly matured proteins upregulates the unfolded protein response (UPR) to cope with stress, activates ER associated degradation (ERAD) for delivery to proteasomes for degradation. We have identified a Brucella abortus type IV secretion system effector called BspL that targets Herp, a key component of ERQC and is able to augment ERAD. Modulation of ERQC by BspL results in tight control of the kinetics of autophagic Brucella-containing vacuole formation, preventing premature bacterial egress from infected cells. This study highlights how bacterial pathogens may hijack ERAD components for fine regulation of their intracellular trafficking.

scholarly journals Characterization of constitutive ER-phagy of excess membrane proteins

PLoS Genetics ◽  
2020 ◽  
Vol 16 (12) ◽  
pp. e1009255
Author(s):  
Zhanna Lipatova ◽  
Valeriya Gyurkovska ◽  
Sarah F. Zhao ◽  
Nava Segev
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Human Health ◽  
Mammalian Cells ◽  
Normal Growth ◽  
Integral Membrane Proteins ◽  
Yeast Cells ◽  
Cellular Proteins

Thirty percent of all cellular proteins are inserted into the endoplasmic reticulum (ER), which spans throughout the cytoplasm. Two well-established stress-induced pathways ensure quality control (QC) at the ER: ER-phagy and ER-associated degradation (ERAD), which shuttle cargo for degradation to the lysosome and proteasome, respectively. In contrast, not much is known about constitutive ER-phagy. We have previously reported that excess of integral-membrane proteins is delivered from the ER to the lysosome via autophagy during normal growth of yeast cells. Whereas endogenously expressed ER resident proteins serve as cargos at a basal level, this level can be induced by overexpression of membrane proteins that are not ER residents. Here, we characterize this pathway as constitutive ER-phagy. Constitutive and stress-induced ER-phagy share the basic macro-autophagy machinery including the conserved Atgs and Ypt1 GTPase. However, induction of stress-induced autophagy is not needed for constitutive ER-phagy to occur. Moreover, the selective receptors needed for starvation-induced ER-phagy, Atg39 and Atg40, are not required for constitutive ER-phagy and neither these receptors nor their cargos are delivered through it to the vacuole. As for ERAD, while constitutive ER-phagy recognizes cargo different from that recognized by ERAD, these two ER-QC pathways can partially substitute for each other. Because accumulation of membrane proteins is associated with disease, and constitutive ER-phagy players are conserved from yeast to mammalian cells, this process could be critical for human health.

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