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 proteasome is a multisubunit protease responsible for degrading proteins conjugated to ubiquitin. The 670-kDa core particle of the proteasome contains the proteolytic active sites, which face an interior chamber within the particle and are thus protected from the cytoplasm. The entry of substrates into this chamber is thought to be governed by the regulatory particle of the proteasome, which covers the presumed channels leading into the interior of the core particle. We have resolved native yeast proteasomes into two electrophoretic variants and have shown that these represent core particles capped with one or two regulatory particles. To determine the subunit composition of the regulatory particle, yeast proteasomes were purified and analyzed by gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Resolution of the individual polypeptides revealed 17 distinct proteins, whose identities were determined by amino acid sequence analysis. Six of the subunits have sequence features of ATPases (Rpt1 to Rpt6). Affinity chromatography was used to purify regulatory particles from various strains, each of which expressed one of the ATPases tagged with hexahistidine. In all cases, multiple untagged ATPases copurified, indicating that the ATPases assembled together into a heteromeric complex. Of the remaining 11 subunits that we have identified (Rpn1 to Rpn3 and Rpn5 to Rpn12), 8 are encoded by previously described genes and 3 are encoded by genes not previously characterized for yeasts. One of the previously unidentified subunits exhibits limited sequence similarity with deubiquitinating enzymes. Overall, regulatory particles from yeasts and mammals are remarkably similar, suggesting that the specific mechanistic features of the proteasome have been closely conserved over the course of evolution.
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PMID:The regulatory particle of the Saccharomyces cerevisiae proteasome. 958 56

The 19S regulatory particle of the yeast 26S proteasome consists of six related ATPases (Rpt proteins) and at least 11 non-ATPase proteins (Rpn proteins). RPN12 (formerly NIN1) encodes an Rpn component of the 19S regulatory particle and is essential for growth. To determine which subunit(s) of the 26S proteasome interact(s) with Rpn12, we attempted to screen for mutations that cause synthetic lethality in the presence of the rpn12-1 (formerly nin1-1) mutation. Among the candidates recovered was a new allele of RPT1 (formerly CIM5). This mutant allele was designated rpt1-2; on its own this mutation caused no phenotypic change, whereas the rpn12-1 rpt1-2 double mutant was lethal, suggesting a strong interaction between Rpn12 and Rpt1. The site of the rpt1-2 mutation was determined by DNA sequencing of the RPT1 locus retrieved from the mutant, and a single nucleotide alteration was found. This changes amino acid 446 of the RPT1 product from alanine to valine. The alanine residue is conserved in all Rpt proteins, except Rpt5, but no function has yet been assigned to the region that contains it. We propose that this region is necessary for Rpt1 to interact with Rpn12. The terminal phenotype of the rpn12-1 rpt1-2 double mutant was not cell cycle specific, suggesting that in the double mutant cells the function of the 26S proteasome is completely eliminated, thereby inducing multiple defects in cellular functions.
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PMID:Genetic evidence for interaction between components of the yeast 26S proteasome: combination of a mutation in RPN12 (a lid component gene) with mutations in RPT1 (an ATPase gene) causes synthetic lethality. 1050 46

Nob1p, which interacts with Nin1p/Rpn12, a subunit of the 19S regulatory particle (RP) of the yeast 26S proteasome, has been identified by two-hybrid screening. NOB1 was found to be an essential gene, encoding a protein of 459 amino acid residues. Nob1p was detected in growing cells but not in cells in the stationary phase. During the transition to the stationary phase, Nob1p was degraded, at least in part, by the 26S proteasome. Nob1p was found only in proteasomal fractions in a glycerol gradient centrifugation profile and immuno-coprecipitated with Rpt1, which is an ATPase component of the yeast proteasomes. These results suggest that association of Nob1p with the proteasomes is essential for the function of the proteasomes in growing cells.
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PMID:Nob1p, a new essential protein, associates with the 26S proteasome of growing saccharomyces cerevisiae cells. 1067 11

Reverse genetic analysis was performed on the Caenorhabditis elegans 26S proteasome subunit genes by double-stranded RNA-mediated interference (RNAi). Embryonic and post-embryonic lethality was caused by interference of all of the eight tested 20S core subunits and all of the 19S regulatory particle subunits except for Ce-Rpn9, Ce-Rpn10, and Ce-Rpn12, where RNAi caused no abnormality. However, synthetic suppression of Ce-Rpn10 and Ce-Rpn12 was lethal, whereas neither the combination of Ce-Rpn9 with Ce-Rpn10 nor with Ce-Rpn12 resulted in abnormalities in RNAi. These results indicate that the 26S proteasome is indispensable for embryogenesis and post-embryonic development, although Ce-Rpn9, Ce-Rpn10, and Ce-Rpn12 are not essential, at least under the conditions used. Ce-Rpn10 and Ce-Rpn12 are considered to compensate for the suppression of each other.
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PMID:Reverse genetic analysis of the Caenorhabditis elegans 26S proteasome subunits by RNA interference. 1243 14

Conjugation of proteins to ubiquitin plays a central role for a number of cellular processes including endocytosis, DNA repair and degradation by the 26S proteasome. However, ubiquitination is reversible as a number of deubiquitinating enzymes mediate the disassembly of ubiquitin-protein conjugates. Some deubiquitinating enzymes are associated with the 26S proteasome contributing to and regulating the particle's activity. Here, we characterise fission yeast Uch2 and Ubp6, two proteasome associated deubiquitinating enzymes. The human orthologues of these enzymes are known as Uch37 and Usp14, respectively. We report that the subunit Uch2/Uch37 is the major deubiquitinating enzyme associated with the fission yeast 26S proteasome. In contrast, the activity of Ubp6 appears to play a more regulatory and/or structural role involving the proteasome subunits Mts1/Rpn9, Mts2/Rpt2 and Mts3/Rpn12, as Ubp6 becomes essential when activity of these subunits is compromised by conditional mutations. Finally, when the genes encoding Uch2/Uch37 and Ubp6 are disrupted, the cells are viable without showing obvious signs of impaired ubiquitin-dependent proteolysis, indicating that other deubiquitinating enzymes may remedy for the redundancy of these enzymes.
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PMID:Uch2/Uch37 is the major deubiquitinating enzyme associated with the 26S proteasome in fission yeast. 1553 39

Human DSS1 associates with BRCA2, a tumour suppressor protein required for efficient recombinational DNA repair, but the biochemical function of DSS1 is not known. Orthologues of DSS1 are found in organisms such as budding yeast and fission yeast that do not have BRCA2-related proteins, indicating that DSS1 has a physiological role independent of BRCA2. The DSS1 orthologue in Saccharomyces cerevisiae has been shown to associate with the 26 S proteasome and, in the present paper, we report that in the distantly related fission yeast Schizosaccharomyces pombe, Dss1 associates with the 19 S RP (regulatory particle) of the 26 S proteasome. A role for S. pombe Dss1 in proteasome function is supported by three lines of evidence. First, overexpression of two components of the 19 S RP, namely Pad1/Rpn11 and Mts3/Rpn12, rescued the temperature-sensitive growth defect of the dss1 mutant. Secondly, the dss1 mutant showed phenotypes indicative of a defect in proteasome function: growth of the dss1 mutant was inhibited by low concentrations of L-canavanine, an amino acid analogue, and cells of the dss1 mutant accumulated high molecular mass poly-ubiquitylated proteins. Thirdly, synthetic growth defects were found when the dss1 mutation was combined with mutations in other proteasome subunit genes. These findings show that DSS1 has an evolutionarily conserved role as a regulator of proteasome function and suggest that DSS1 may provide a link between BRCA2 and ubiquitin-mediated proteolysis in human cells.
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PMID:Fission yeast Dss1 associates with the proteasome and is required for efficient ubiquitin-dependent proteolysis. 1614 16

The 26S proteasome is a highly conserved multisubunit protease that degrades ubiquitinated proteins in eukaryotic cells. It comprises a 20S core particle and two 19S regulatory particles that are further divided into the lid and base complexes. The lid is a nine subunits complex that is structurally related to the COP9 signalosome and the eukaryotic initiation factor 3. Although the assembly pathway of the 20S and the base are well described, that of the lid is still unclear. In this study, we dissected the lid assembly using yeast lid mutant cells, rpn7-3, Delta rpn9, and rpn12-1. Using mass spectrometry, we identified a number of lid subassemblies, such as Rpn3-Rpn7 pair and a lid-like complex lacking Rpn12, in the mutants. Our analysis suggests that the assembly of the lid is a highly ordered and multi-step process; first, Rpn5, 6, 8, 9, and 11 are assembled to form a core module, then a second module, consisting of Rpn3, 7, and Sem1, is attached, followed by the incorporation of Rpn12 to form the lid complex.
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PMID:Dissection of the assembly pathway of the proteasome lid in Saccharomyces cerevisiae. 2047 55

The 26S proteasome, the central eukaryotic protease, comprises a core particle capped by a 19S regulatory particle (RP). The RP is divisible into base and lid subcomplexes. Lid biogenesis and incorporation into the RP remain poorly understood. We report several lid intermediates, including the free Rpn12 subunit and a lid particle (LP) containing the remaining eight subunits, LP2. Rpn12 binds LP2 in vitro, and each requires the other for assembly into 26S proteasomes. Stable Rpn12 incorporation depends on all other lid subunits, indicating that Rpn12 distinguishes LP2 from smaller lid subcomplexes. The highly conserved C terminus of Rpn12 bridges the lid and base, mediating both stable binding to LP2 and lid-base joining. Our data suggest a hierarchical assembly mechanism where Rpn12 binds LP2 only upon correct assembly of all other lid subunits, and the Rpn12 tail then helps drive lid-base joining. Rpn12 incorporation thus links proper lid assembly to subsequent assembly steps.
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PMID:Incorporation of the Rpn12 subunit couples completion of proteasome regulatory particle lid assembly to lid-base joining. 2219 64

It has recently become clear that components of the proteasome are recruited to sites of gene transcription. Prevailing evidence suggests that the transcriptionally relevant form of the proteasome is a subcomplex of 19S base proteins, which functions as an ATP-dependent chaperone that influences transcriptional processes. Despite this notion, compelling evidence for a transcription-dedicated 19S base complex is lacking, and 20S proteasome subunits have been shown to associate with chromatin in some contexts. To gain insight into the form of the proteasome that is recruited to chromatin, we assembled a panel of highly specific antibodies that recognize native yeast proteasome subunits in chromatin immunoprecipitation assays. Using these reagents, we show that components from the three major subassemblies of the proteasome--19S lid, 19S base, and 20S core--associate with the activated GAL10 gene in yeast in a virtually indistinguishable manner. We find that proteasome subunits Rpt1, Rpt4, Rpn8, Rpn12, Pre6, and Pre10 are recruited to GAL10 rapidly upon galactose induction. These subunits associate with the entire transcribed portion of GAL10, display near-identical patterns of distribution, and dissociate from chromatin rapidly once transcription is shut down. We also find that proteasome subunits are enriched at telomeres and at genes transcribed by RNA polymerase III. Our data suggest that the transcriptionally relevant form of the proteasome is the canonical 26S complex.
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PMID:Similar temporal and spatial recruitment of native 19S and 20S proteasome subunits to transcriptionally active chromatin. 2247 42

The ubiquitin-proteasome system targets selected proteins for degradation by the 26S proteasome. Rpn12 is an essential component of the 19S regulatory particle and plays a role in recruiting the extrinsic ubiquitin receptor Rpn10. In the present paper we report the crystal structure of Rpn12, a proteasomal PCI-domain-containing protein. The structure helps to define a core structural motif for the PCI domain and identifies potential sites through which Rpn12 might form protein-protein interactions. We demonstrate that mutating residues at one of these sites impairs Rpn12 binding to Rpn10 in vitro and reduces Rpn10 incorporation into proteasomes in vivo.
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PMID:Structural and functional characterization of Rpn12 identifies residues required for Rpn10 proteasome incorporation. 2290 49


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