scholarly journals Tay–Sachs disease mutations in HEXA target the α chain of hexosaminidase A to endoplasmic reticulum–associated degradation

2016 ◽  
Vol 27 (24) ◽  
pp. 3813-3827 ◽  
Author(s):  
Devin Dersh ◽  
Yuichiro Iwamoto ◽  
Yair Argon
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Control Mechanisms ◽  
Permissive Temperature ◽  
Lysosomal Storage ◽  
Loss Of Function ◽  
Er Quality Control ◽  
Tay Sachs Disease ◽  
Hexosaminidase A ◽  
Α Chain

Loss of function of the enzyme β-hexosaminidase A (HexA) causes the lysosomal storage disorder Tay–Sachs disease (TSD). It has been proposed that mutations in the α chain of HexA can impair folding, enzyme assembly, and/or trafficking, yet there is surprisingly little known about the mechanisms of these potential routes of pathogenesis. We therefore investigated the biosynthesis and trafficking of TSD-associated HexA α mutants, seeking to identify relevant cellular quality control mechanisms. The α mutants E482K and G269S are defective in enzymatic activity, unprocessed by lysosomal proteases, and exhibit altered folding pathways compared with wild-type α. E482K is more severely misfolded than G269S, as observed by its aggregation and inability to associate with the HexA β chain. Importantly, both mutants are retrotranslocated from the endoplasmic reticulum (ER) to the cytosol and are degraded by the proteasome, indicating that they are cleared via ER-associated degradation (ERAD). Leveraging these discoveries, we observed that manipulating the cellular folding environment or ERAD pathways can alter the kinetics of mutant α degradation. Additionally, growth of patient fibroblasts at a permissive temperature or with chemical chaperones increases cellular Hex activity by improving mutant α folding. Therefore modulation of the ER quality control systems may be a potential therapeutic route for improving some forms of TSD.

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 A cross-kingdom conserved ER-phagy receptor maintains endoplasmic reticulum homeostasis during stress

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Madlen Stephani ◽  
Lorenzo Picchianti ◽  
Alexander Gajic ◽  
Rebecca Beveridge ◽  
Emilio Skarwan ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Mammalian Cells ◽  
Control Mechanisms ◽  
Receptor Complex ◽  
Er Quality Control ◽  
Proteotoxic Stress ◽  
Quality Control Mechanism ◽  
Passenger Proteins

Eukaryotes have evolved various quality control mechanisms to promote proteostasis in the endoplasmic reticulum (ER). Selective removal of certain ER domains via autophagy (termed as ER-phagy) has emerged as a major quality control mechanism. However, the degree to which ER-phagy is employed by other branches of ER-quality control remains largely elusive. Here, we identify a cytosolic protein, C53, that is specifically recruited to autophagosomes during ER-stress, in both plant and mammalian cells. C53 interacts with ATG8 via a distinct binding epitope, featuring a shuffled ATG8 interacting motif (sAIM). C53 senses proteotoxic stress in the ER lumen by forming a tripartite receptor complex with the ER-associated ufmylation ligase UFL1 and its membrane adaptor DDRGK1. The C53/UFL1/DDRGK1 receptor complex is activated by stalled ribosomes and induces the degradation of internal or passenger proteins in the ER. Consistently, the C53 receptor complex and ufmylation mutants are highly susceptible to ER stress. Thus, C53 forms an ancient quality control pathway that bridges selective autophagy with ribosome-associated quality control in the ER.

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.

Rapid Nonradioactive Tracer Method for Detecting Carriers of the Major Ashkenazi Jewish Tay-Sachs Disease Mutations

1992 ◽  
Vol 38 (11) ◽  
pp. 2249-2255 ◽  
Author(s):  
P M Strasberg ◽  
J T Clarke
Keyword(s):  
Mutation Analysis ◽  
Restriction Analysis ◽  
Type I ◽  
Ashkenazi Jewish ◽  
Lysosomal Storage ◽  
Ashkenazi Jews ◽  
Dot Blot ◽  
Chain Reactions ◽  
Tay Sachs Disease ◽  
Hexosaminidase A

Abstract Tay-Sachs disease (TSD, GM2 gangliosidosis, Type I) is an autosomal recessive lysosomal storage disease caused by deficiency of beta-hexosaminidase A (Hex A) resulting from mutations in the gene (HEXA) encoding the alpha-subunit of the enzyme. Three mutations, in exons 7 and 11 and at the exon 12-intron 12 junction, account for > 90% of alleles identified in obligate Ashkenazi Jewish carriers. Mutation analysis requires amplification of available DNA by separate polymerase chain reactions (PCRs) and either restriction digestion and gel electrophoresis or 32P-labeled allele-specific oligonucleotide (ASO) probes. We developed a simple, nonradioisotopic method for rapidly identifying TSD carriers by a triplex PCR reaction followed by dot-blot analysis, using three wild-type and three mutant ASOs end-labeled with digoxigenin-dUTP (dig-ASO). Hybridization was demonstrated immunologically by reaction with an anti-digoxigenin-alkaline phosphatase conjugate followed by colorimetric demonstration of phosphatase activity. The results of analyses by the dig-ASO method of 65 carriers identified by serum enzyme activity and of 6 high-risk fetuses in prenatal testing were the same as those obtained by more conventional restriction analysis. Dig-ASO testing correctly reclassified 10 individuals who had tested inconclusively on analysis for leukocyte beta-hexosaminidase A activity; 3 were identified as carriers and 7 as noncarriers. The simplicity of the assay and the avoidance of the radioisotopes make this a potentially useful method for TSD carrier detection by mutation analysis in Ashkenazi Jews from populations in whom the identity and frequencies of the common TSD mutations are known.

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 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 A cross-kingdom conserved ER-phagy receptor maintains endoplasmic reticulum homeostasis during stress

2020 ◽  
Author(s):  
Madlen Stephani ◽  
Lorenzo Picchianti ◽  
Alexander Gajic ◽  
Rebecca Beveridge ◽  
Emilio Skarwan ◽  
...  
Keyword(s):  
Quality Control ◽  
Er Stress ◽  
Mammalian Cells ◽  
Control Mechanisms ◽  
Receptor Complex ◽  
Er Quality Control ◽  
Proteotoxic Stress ◽  
Binding Epitope ◽  
Quality Control Mechanism ◽  
Passenger Proteins

SummaryEukaryotes have evolved various quality control mechanisms to promote proteostasis in the ER. Selective removal of certain ER domains via autophagy (termed as ER-phagy) has emerged as a major quality control mechanism. However, the degree to which ER-phagy is employed by other branches of ER-quality control remains largely elusive. Here, we identify a cytosolic protein, C53, that is specifically recruited to autophagosomes during ER-stress, in both plant and mammalian cells. C53 interacts with ATG8 via a distinct binding epitope, featuring a shuffled ATG8 interacting motif (sAIM). C53 senses proteotoxic stress in the ER lumen by forming a tripartite receptor complex with the ER-associated ufmylation ligase UFL1 and its membrane adaptor DDRGK1. The C53/UFL1/DDRGK1 receptor complex is activated by stalled ribosomes and induces the degradation of internal or passenger proteins in the ER. Consistently, the C53 receptor complex and ufmylation mutants are highly susceptible to ER stress. Thus, C53 forms an ancient quality control pathway that bridges selective autophagy with ribosome-associated quality control at the ER.

scholarly journals Cln6 mutants associated with neuronal ceroid lipofuscinosis are degraded in a proteasome-dependent manner

2009 ◽  
Vol 29 (3) ◽  
pp. 173-181 ◽  
Author(s):  
Kristina Oresic ◽  
Britta Mueller ◽  
Domenico Tortorella
Keyword(s):  
Quality Control ◽  
Protein Extraction ◽  
Neuronal Ceroid Lipofuscinosis ◽  
Point Mutations ◽  
Lysosomal Storage Diseases ◽  
Neuronal Ceroid Lipofuscinoses ◽  
Dependent Manner ◽  
Lysosomal Storage ◽  
Er Quality Control ◽  
Recessive Mutations

NCLs (neuronal ceroid lipofuscinoses), a group of inherited neurodegenerative lysosomal storage diseases that predominantly affect children, are the result of autosomal recessive mutations within one of the nine cln genes. The wild-type cln gene products are composed of membrane and soluble proteins that localize to the lysosome or the ER (endoplasmic reticulum). However, the destiny of the Cln variants has not been fully characterized. To explore a possible link between ER quality control and processing of Cln mutants, we investigated the fate of two NCL-related Cln6 mutants found in patient samples (Cln6G123D and Cln6M241T) in neuronal-derived human cells. The point mutations are predicted to be in the putative transmembrane domains and most probably generate misfolded membrane proteins that are subjected to ER quality control. Consistent with this paradigm, both mutants underwent rapid proteasome-mediated degradation and complexed with components of the ER extraction apparatus, Derlin-1 and p97. In addition, knockdown of SEL1L [sel-1 suppressor of lin-12-like (Caenorhabditis elegans)], a member of an E3 ubiquitin ligase complex involved in ER protein extraction, rescued significant amounts of Cln6G123D and Cln6M241T polypeptides. The results implicate ER quality control in the instability of the Cln variants that probably contributes to the development of NCL.

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