Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.25.1 (proteasome)
28,817 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The endoplasmic reticulum (ER) glycoprotein HMG-CoA reductase (HMGR) catalyzes the rate-limiting step in sterols biosynthesis. Mammalian HMGR is ubiquitinated and degraded by the proteasome when sterols accumulate in cells, representing the best example for metabolically controlled ER-associated degradation (ERAD). This regulated degradation involves the short-lived ER protein Insig-1. Here, we investigated the dislocation of these ERAD substrates to the cytosol en route to proteasomal degradation. We show that the tagged HMGR membrane region, HMG(350)-HA, the endogenous HMGR, and Insig-1-Myc, all polytopic membrane proteins, dislocate to the cytosol as intact full-length polypeptides. Dislocation of HMG(350)-HA and Insig-1-Myc requires metabolic energy and involves the AAA-ATPase p97/VCP. Sterols stimulate HMG(350)-HA and HMGR release to the cytosol concurrent with removal of their N-glycan by cytosolic peptide:N-glycanase. Sterols neither accelerate dislocation nor stimulate deglycosylation of ubiquitination-defective HMG(350)-HA((K89 + 248R)) mutant. Dislocation of HMG(350)-HA depends on Insig-1-Myc, whose dislocation and degradation are sterol independent. Coimmunoprecipitation experiments demonstrate sterol-stimulated association between HMG(350)-HA and Insig-1-Myc. Sterols do not enhance binding to Insig-1-Myc of HMG(350)-HA mutated in its sterol-sensing domain or of HMG(350)-HA((K89 + 248R)). Wild-type HMG(350)-HA and Insig-1-Myc coimmunoprecipitate from the soluble fraction only when both proteins were coexpressed in the same cell, indicating their encounter before or during dislocation, raising the possibility that they are dislocated as a tightly bound complex.
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PMID:Dislocation of HMG-CoA reductase and Insig-1, two polytopic endoplasmic reticulum proteins, en route to proteasomal degradation. 1945 99

Inositol 1,4,5-trisphosphate (IP(3)) receptors are endoplasmic reticulum (ER) membrane calcium channels that, upon activation, become substrates for the ER-associated degradation (ERAD) pathway. While it is clear that IP(3) receptors are polyubiquitinated and are transferred to the proteasome by a p97-based complex, currently very little is known about the proteins that initially select activated IP(3) receptors for ERAD. Here, we have transfected HeLa cells to stably express m3 muscarinic receptors to allow for the study of IP(3) receptor ERAD in this cell type, and show that IP(3) receptors are polyubiquitinated and then degraded by the proteasome in response to carbachol, a muscarinic agonist. In seeking to identify proteins that mediate IP(3) receptor ERAD we found that both SPFH1 and SPFH2 (also known as erlin 1 and erlin 2), which exist as a hetero-oligomeric complex, rapidly associate with IP(3) receptors in a manner that precedes polyubiquitination and the association of p97. Suppression of SPFH1 and SPFH2 expression by RNA interference markedly inhibited carbachol-induced IP(3) receptor polyubiquitination and degradation, but did not affect carbachol-induced calcium mobilization or IkappaBalpha processing, indicating that the SPFH1/2 complex is a key player in IP(3) receptor ERAD, acting at a step after IP(3) receptor activation, but prior to IP(3) receptor polyubiquitination. Suppression of SPFH1 and SPFH2 expression had only slight effects on the turnover of some exogenous model ERAD substrates, and had no effect on sterol-induced ERAD of endogenous 3-hydroxy-3-methylglutaryl-CoA reductase. Overall, these studies show that m3 receptor-expressing HeLa cells are a valuable system for studying IP(3) receptor ERAD, and suggest that the SPFH1/2 complex is a factor that selectively mediates the ERAD of activated IP(3) receptors.
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PMID:SPFH1 and SPFH2 mediate the ubiquitination and degradation of inositol 1,4,5-trisphosphate receptors in muscarinic receptor-expressing HeLa cells. 1975 72

p97 (CDC-48 in Caenorhabditis elegans) is a ubiquitin-selective AAA (ATPases associated with diverse cellular activities) chaperone and its key function is to disassemble protein complexes. p97 functions in diverse cellular processes including endoplasmic reticulum (ER)-associated degradation, membrane fusion, and meiotic and mitotic progression. However, its cellular functions in development have not yet been clarified. Here, we present data that p97 is involved in the switch from spermatogenesis to oogenesis in the germline of the C. elegans hermaphrodite. We found that the cdc-48.1 deletion mutant produced less sperm than the wild type and thus showed a decreased brood size. The cdc-48.1 mutation suppressed the sperm-overproducing phenotypes of fbf-1 and fem-3(gf) mutants. In addition, the p97/CDC-48-UFD-1-NPL-4 complex interacted with the E3 ubiquitin ligase CUL-2 complex via NPL-4 binding to Elongin C. Furthermore, TRA-1A, which is the terminal effector of the sex determination pathway and is regulated by CUL-2-mediated proteolysis, accumulated in the cdc-48.1 mutant. Proteasome activity was also required for the brood size determination and sperm-oocyte switch. Our results demonstrate that the C. elegans p97/CDC-48-UFD-1-NPL-4 complex controls the sperm-oocyte switch by regulating CUL-2-mediated TRA-1A proteasome degradation.
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PMID:Caenorhabditis elegans p97 controls germline-specific sex determination by controlling the TRA-1 level in a CUL-2-dependent manner. 1977 60

The multifunctional AAA-ATPase p97 is one of the most abundant and conserved proteins in eukaryotic cells. The p97/Npl4/Ufd1 complex dislocates proteins that fail the protein quality control in the endoplasmic reticulum to the cytosol where they are subject to degradation by the ubiquitin/proteasome system. Substrate dislocation depends on the unfoldase activity of p97. Interestingly, p97 is also involved in the degradation of specific soluble proteasome substrates but the exact mode of action of p97 in this process is unclear. Here, we show that both the central pore and ATPase activity of p97 are necessary for the degradation of cytosolic ubiquitin-fusion substrates. Addition of a flexible extended C-terminal peptide to the substrate relieves the requirement for p97. Deletion mapping reveals a conserved length dependency of 20 residues for the peptide, which allows p97-independent degradation to occur. Our results suggest that initiation of unfolding may be more complex than previously anticipated and that the 19S regulatory complex of the proteasome can require preprocessing of highly folded, ubiquitylated substrates by the p97(Ufd1/Npl4) complex. Our data provide an explanation for the observation that p97 is only essential for a subpopulation of soluble substrates and predict that a common characteristic of soluble p97-dependent substrates is the lack of an initiation site to facilitate unfolding by the 26S proteasome.
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PMID:A conserved unfoldase activity for the p97 AAA-ATPase in proteasomal degradation. 1978 90

Autosomal dominant mutations in the SFTPC gene are associated with idiopathic pulmonary fibrosis, a progressive lethal interstitial lung disease. Mutations that cause misfolding of the encoded proprotein surfactant protein C (SP-C) trigger endoplasmic reticulum (ER)-associated degradation, a pathway that segregates terminally misfolded substrate for retrotranslocation to the cytosol and degradation by proteasome. Microarray screens for genes involved in SP-C ER-associated degradation identified MKS3/TMEM67, a locus previously linked to the ciliopathy Meckel-Gruber syndrome. In this study, MKS3 was identified as a membrane glycoprotein predominantly localized to the ER. Expression of MKS3 was up-regulated by genetic or pharmacological inducers of ER stress. The ER lumenal domain of MKS3 interacted with a complex that included mutant SP-C and associated chaperones, whereas the region predicted to encode the transmembrane domains of MKS3 interacted with cytosolic p97. Deletion of the transmembrane and cytosolic domains abrogated interaction of MKS3 with p97 and resulted in accumulation of mutant SP-C proprotein; knockdown of MKS3 also inhibited degradation of mutant SP-C. These results support a model in which MKS3 links the ER lumenal quality control machinery with the cytosolic degradation apparatus.
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PMID:Meckel-Gruber syndrome protein MKS3 is required for endoplasmic reticulum-associated degradation of surfactant protein C. 1981 49

Unwanted proteins in the endoplasmic reticulum (ER) are exported into the cytoplasm and degraded by the proteasome through the ER-associated protein degradation pathway (ERAD). Disturbances in ERAD are linked to ER stress, which has been implicated in the pathogenesis of several human diseases. However, the composition and organization of ERAD complexes in human cells is still poorly understood. In this paper, we describe a trimeric complex that we propose functions in ERAD. Knockdown of erasin, a platform for p97/VCP and ubiquilin binding, or knockdown of ubiquilin in human cells slowed degradation of two classical ERAD substrates. In Caenorhabditis elegans, ubiquilin and erasin are ER stress-response genes that are regulated by the ire-1 branch of the unfolded protein response pathway. Loss of ubiquilin or erasin resulted in activation of ER stress, increased accumulation of polyubiquitinated proteins, and shortened lifespan in worms. Our results strongly support a role for this complex in ERAD and in the regulation of ER stress.
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PMID:Ubiquilin and p97/VCP bind erasin, forming a complex involved in ERAD. 1982 69

Ubiquitinated proteins can alternatively be delivered directly to the proteasome or via p97/VCP (valosin-containing protein). Whereas the proteasome degrades ubiquitinated proteins, the homohexameric ATPase p97/VCP seems to control the ubiquitination status of recruited substrates. The COP9 signalosome (CSN) is also involved in the ubiquitin/proteasome system (UPS) as exemplified by regulating the neddylation of ubiquitin E3 ligases. Here, we show that p97/VCP colocalizes and directly interacts with subunit 5 of the CSN (CSN5) in vivo and is associated with the entire CSN complex in an ATP-dependent manner. Furthermore, we provide evidence that the CSN and in particular the isopeptidase activity of its subunit CSN5 as well as the associated deubiquitinase USP15 are required for proper processing of polyubiquitinated substrates bound to p97/VCP. Moreover, we show that in addition to NEDD8, CSN5 binds to oligoubiquitin chains in vitro. Therefore, CSN and p97/VCP could form an ATP-dependent complex that resembles the 19 S proteasome regulatory particle and serves as a key mediator between ubiquitination and degradation pathways.
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PMID:COP9 signalosome interacts ATP-dependently with p97/valosin-containing protein (VCP) and controls the ubiquitination status of proteins bound to p97/VCP. 1982 4

PLAA (ortholog of yeast Doa1/Ufd3, also know as human PLAP or phospholipase A2-activating protein) has been implicated in a variety of disparate biological processes that involve the ubiquitin system. It is linked to the maintenance of ubiquitin levels, but the mechanism by which it accomplishes this is unclear. The C-terminal PUL (PLAP, Ufd3p, and Lub1p) domain of PLAA binds p97, an AAA ATPase, which among other functions helps transfer ubiquitinated proteins to the proteasome for degradation. In yeast, loss of Doa1 is suppressed by altering p97/Cdc48 function indicating that physical interaction between PLAA and p97 is functionally important. Although the overall regions of interaction between these proteins are known, the structural basis has been unavailable. We solved the high resolution crystal structure of the p97-PLAA complex showing that the PUL domain forms a 6-mer Armadillo-containing domain. Its N-terminal extension folds back onto the inner curvature forming a deep ridge that is positively charged with residues that are phylogenetically conserved. The C terminus of p97 binds in this ridge, where the side chain of p97-Tyr(805), implicated in phosphorylation-dependent regulation, is buried. Expressed in doa1Delta null cells, point mutants of the yeast ortholog Doa1 that disrupt this interaction display slightly reduced ubiquitin levels, but unlike doa1Delta null cells, showed only some of the growth phenotypes. These data suggest that the p97-PLAA interaction is important for a subset of PLAA-dependent biological processes and provides a framework to better understand the role of these complex molecules in the ubiquitin system.
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PMID:Structure and function of the PLAA/Ufd3-p97/Cdc48 complex. 1988 78

The human type II AAA+ protein p97 participates in various cellular activities, presumably through its involvement in the ubiquitin-proteasome degradation pathway. Mutations in p97 have been implicated in patients with inclusion-body myopathy associated with Paget's disease of the bone and frontotemporal dementia (IBMPFD). In this work, three mutant p97 N-D1 fragments, R86A, R95G and R155H, were crystallized in the presence of ATPgammaS with PEG 3350 as a main precipitant, yielding two different crystal forms. The R155H mutant crystal belonged to space group R3, with unit-cell parameters in the hexagonal setting of a = b = 134.2, c = 182.9 angstrom, and was merohedrally twinned, with an estimated twin fraction of 0.34. The crystals of the R86A and R95G mutants belonged to space group P1, with similar unit-cell parameters of a = 90.89, b = 102.6, c = 107.2 angstrom, alpha = 97.5, beta = 90.6, gamma = 91.5 degrees and a = 92.76, b = 103.7, c = 107.7 angstrom , alpha = 97.7, beta = 91.9, gamma = 89.7 degrees, respectively.
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PMID:Purification, crystallization and preliminary X-ray diffraction analysis of disease-related mutants of p97. 1992 42

UBXD1 is a recently identified adaptor for p97, a highly abundant and conserved member of the AAA family of ATPase that plays pivotal roles in a multitude of cellular processes involving the ubiquitin-proteasome pathway. Very little is known about the biochemical, cellular, and molecular functions of UBXD1. Here we report the generation of two mouse monoclonal antibodies, 5C3-1 and 2F8-24, that recognize UBXD1 using Western blotting, immunoprecipitation, and immunofluorescence.
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PMID:Generation and characterization of novel monoclonal antibodies recognizing UBXD1. 2002 8


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