Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P62988 (Ubiquitin)
 4,326 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A dominant insertional P-element mutation enhances position-effect variegation in Drosophila melanogaster. The mutation is homozygous, viable, and fertile and maps at 64E on the third chromosome. The corresponding gene was cloned by transposon tagging. Insertion of the transposon upstream of the open reading frame correlates with a strong reduction of transcript level. A transgene was constructed with the cDNA and found to have the effect opposite from that of the mutation, namely, to suppress variegation. Sequencing of the cDNA reveals a large open reading frame encoding a putative ubiquitin-specific protease (Ubp). Ubiquitin marks various proteins, frequently for proteasome-dependent degradation. Ubps can cleave the ubiquitin part from these proteins. We discuss the link established here between a deubiquitinating enzyme and epigenetic silencing processes.
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PMID:The dose of a putative ubiquitin-specific protease affects position-effect variegation in Drosophila melanogaster. 881 85

Degradation of many eukaryotic proteins requires their prior ligation to polyubiquitin chains, which target substrates to the 26S proteasome, an abundant cellular protease. We describe a yeast deubiquitinating enzyme, Ubp14, that specifically disassembles unanchored ('free') ubiquitin chains in vitro, a specificity shared by mammalian isopeptidase T. Correspondingly, deletion of the UBP14 gene from yeast cells results in a striking accumulation of free ubiquitin chains, which correlates with defects in ubiquitin-dependent proteolysis. Increasing the steady-state levels of ubiquitin chains in wild-type cells (by expressing a derivative of ubiquitin with an altered C-terminus) inhibits protein degradation to a degree comparable with that observed in ubp14delta cells. Inhibition of degradation is also seen when an active site mutant of Ubp14 is overproduced in vivo. Surprisingly, overproduction of wild-type Ubp14 can inhibit degradation of some proteins as well. Finally, Ubp14 and human isopeptidase T are shown to be functional homologs by complementation analysis. We propose that Ubp14 and isopeptidase T facilitate proteolysis in vivo by preventing unanchored ubiquitin chains from competitively inhibiting polyubiquitin-substrate binding to the 26S proteasome.
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PMID:In vivo disassembly of free polyubiquitin chains by yeast Ubp14 modulates rates of protein degradation by the proteasome. 930 25

The Drosophila fat facets gene encodes a deubiquitinating enzyme that regulates a cell communication pathway essential early during eye development to inhibit the determination of excess photoreceptors. Ubiquitin is a small polypeptide that tags proteins for degradation by a multisubunit proteolytic complex called the proteasome. The FAT FACETS protein is thought to be required to remove ubiquitin from a particular protein, thereby rescuing if from proteolysis. In order to identify the genes encoding the substrate of FAT FACETS and other components of the neural inhibition pathway, a mutagenesis screen for dominant enhancers of the fat facets mutant eye phenotype was performed. Several genes were identified, one of which is an excellent candidate for encoding a component of the pathway regulated by FAT FACETS. Three different eye phenotypes were observed when the fat facets mutants were dominantly enhanced by different mutations, suggesting that fat facets has other functions in addition to its critical role early in eye development.
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PMID:Mutagenesis screens for interacting genes reveal three roles for fat facets during Drosophila eye development. 933 74

Although cell differentiation usually involves synthesis of new proteins, little is known about the role of protein degradation. In eukaryotes, conjugation to ubiquitin polymers often targets a protein for destruction. This process is regulated by deubiquitinating enzymes, which can disassemble ubiquitin polymers or ubiquitin-substrate conjugates. We find that a deubiquitinating enzyme, UbpA, is required for Dictyostelium development. ubpA cells have normal protein profiles on gels, grow normally, and show normal responses to starvation such as differentiation and secretion of conditioned medium factor. However, ubpA cells have defective aggregation, chemotaxis, cAMP relay, and cell adhesion. These defects result from low expression of cAMP pulse-induced genes such as those encoding the cAR1 cAMP receptor, phosphodiesterase, and the gp80 adhesion protein. Treatment of ubpA cells with pulses of exogenous cAMP allows them to aggregate and express these genes like wild-type cells, but they still fail to develop fruiting bodies. Unlike wild type, ubpA cells accumulate ubiquitin-containing species that comigrate with ubiquitin polymers, suggesting a defect in polyubiquitin metabolism. UbpA has sequence similarity with yeast Ubp14, which disassembles free ubiquitin chains. Yeast ubp14 cells have a defect in proteolysis, due to excess ubiquitin chains competing for substrate binding to proteasomes. Cross-species complementation and enzyme specificity assays indicate that UbpA and Ubp14 are functional homologs. We suggest that specific developmental transitions in Dictyostelium require the degradation of specific proteins and that this process in turn requires the disassembly of polyubiquitin chains by UbpA.
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PMID:A deubiquitinating enzyme that disassembles free polyubiquitin chains is required for development but not growth in Dictyostelium. 978 28

Attachment of ubiquitin to cellular proteins frequently targets them to the 26S proteasome for degradation. In addition, ubiquitination of cell surface proteins stimulates their endocytosis and eventual degradation in the vacuole or lysosome. In the yeast Saccharomyces cerevisiae, ubiquitin is a long-lived protein, so it must be efficiently recycled from the proteolytic intermediates to which it becomes linked. We identified previously a yeast deubiquitinating enzyme, Doa4, that plays a central role in ubiquitin-dependent proteolysis by the proteasome. Biochemical and genetic data suggest that Doa4 action is closely linked to that of the proteasome. Here we provide evidence that Doa4 is required for recycling ubiquitin from ubiquitinated substrates targeted to the proteasome and, surprisingly, to the vacuole as well. In the doa4Delta mutant, ubiquitin is strongly depleted under certain conditions, most notably as cells approach stationary phase. Ubiquitin depletion precedes a striking loss of cell viability in stationary phase doa4Delta cells. This loss of viability and several other defects of doa4Delta cells are rescued by provision of additional ubiquitin. Ubiquitin becomes depleted in the mutant because it is degraded much more rapidly than in wild-type cells. Aberrant ubiquitin degradation can be partially suppressed by mutation of the proteasome or by inactivation of vacuolar proteolysis or endocytosis. We propose that Doa4 helps recycle ubiquitin from both proteasome-bound ubiquitinated intermediates and membrane proteins destined for destruction in the vacuole.
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PMID:The Doa4 deubiquitinating enzyme is required for ubiquitin homeostasis in yeast. 1043 14

The Drosophila fat facets gene encodes a deubiquitinating enzyme required during eye development to limit the number of photoreceptors in each facet to eight. Ubiquitin is a small polypeptide that targets proteins for degradation by the proteasome. Deubiquitinating enzymes cleave ubiquitin-protein bonds. In order to investigate the role of FAT FACETS in the ubiquitin pathway, genetic interactions between fat facets and the Drosophila UbcD1 gene were assessed. In addition, three yeast deubiquitinating enzyme genes were tested for their ability to substitute for fat facets in the developing Drosophila eye and for their effects on eye morphology. The results of these experiments support the hypothesis that FAT FACETS activity antagonizes that of the proteolytic machinery. The implications of these results for the specificity of FAF and yeast UBPs are discussed as well.
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PMID:Genetic analysis of the role of the drosophila fat facets gene in the ubiquitin pathway. 1057 Apr 63

Ubiquitin-specific processing proteases (UBPs) presently form the largest enzyme family in the ubiquitin system, characterized by a core region containing conserved motifs surrounded by divergent sequences, most commonly at the N-terminal end. The functions of these divergent sequences remain unclear. We identified two isoforms of a novel testis-specific UBP, UBP-t1 and UBP-t2, which contain identical core regions but distinct N termini, thereby permitting dissection of the functions of these two regions. Both isoforms were germ cell specific and developmentally regulated. Immunocytochemistry revealed that UBP-t1 was induced in step 16 to 19 spermatids while UBP-t2 was expressed in step 18 to 19 spermatids. Immunoelectron microscopy showed that UBP-t1 was found in the nucleus while UBP-t2 was extranuclear and was found in residual bodies. For the first time, we show that the differential subcellular localization was due to the distinct N-terminal sequences. When transfected into COS-7 cells, the core region was expressed throughout the cell but the UBP-t1 and UBP-t2 isoforms were concentrated in the nucleus and the perinuclear region, respectively. Fusions of each N-terminal end with green fluorescent protein yielded the same subcellular localization as the native proteins, indicating that the N-terminal ends were sufficient for determining differential localization. Interestingly, UBP-t2 colocalized with anti-gamma-tubulin immunoreactivity, indicating that like several other components of the ubiquitin system, a deubiquitinating enzyme is associated with the centrosome. Regulated expression and alternative N termini can confer specificity of UBP function by restricting its temporal and spatial loci of action.
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PMID:Divergent N-terminal sequences target an inducible testis deubiquitinating enzyme to distinct subcellular structures. 1093 31

Ubiquitin-dependent proteolysis is catalyzed by the 26S proteasome, a dynamic complex of 32 different proteins whose mode of assembly and mechanism of action are poorly understood, in part due to the difficulties encountered in purifying the intact complex. Here we describe a one-step affinity method for purifying intact 26S proteasomes, 19S regulatory caps, and 20S core particles from budding yeast cells. Affinity-purified 26S proteasomes hydrolyze both model peptides and the ubiquitinated Cdk inhibitor Sic1. Affinity purifications performed in the absence of ATP or presence of the poorly hydrolyzable analog ATP-gamma-S unexpectedly revealed that a large number of proteins, including subunits of the skp1-cullin-F-box protein ligase (SCF) and anaphase-promoting complex (APC) ubiquitin ligases, copurify with the 19S cap. To identify these proteasome-interacting proteins, we used a recently developed method that enables the direct analysis of the composition of large protein complexes (DALPC) by mass spectrometry. Using DALPC, we identified more than 24 putative proteasome-interacting proteins, including Ylr421c (Daq1), which we demonstrate to be a new subunit of the budding yeast 19S cap, and Ygr232w (Nas6), which is homologous to a subunit of the mammalian 19S cap (PA700 complex). Additional PIPs include the heat shock proteins Hsp70 and Hsp82, the deubiquitinating enzyme Ubp6, and proteins involved in transcriptional control, mitosis, tubulin assembly, RNA metabolism, and signal transduction. Our data demonstrate that nucleotide hydrolysis modulates the association of many proteins with the 26S proteasome, and validate DALPC as a powerful tool for rapidly identifying stoichiometric and substoichiometric components of large protein assemblies.
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PMID:Proteasomal proteomics: identification of nucleotide-sensitive proteasome-interacting proteins by mass spectrometric analysis of affinity-purified proteasomes. 1102 46

The Drosophila Fat facets protein is a deubiquitinating enzyme required for patterning the developing compound eye. Ubiquitin, a 76-amino-acid polypeptide, serves as a tag to direct proteins to the proteasome, a protein degradation complex. Deubiquitinating enzymes are a large group of proteins that cleave ubiquitin-protein bonds. Fat facets belongs to a class of deubiquitinating enzymes called Ubps that share a conserved catalytic domain. Fat facets is unique among them in its large size and also because Fat facets is thought to deubiquitinate a specific substrate thereby preventing its proteolysis. Here we asked which portions of the Fat facets protein are essential for its function. P-element constructs that express partial Fat facets proteins were tested for function. In addition, the DNA sequences of 12 mutant fat facets alleles were determined. Finally, regions of amino acid sequence similarity in 18 Drosophila Ubps revealed by the Genome Project were identified. The results indicate functions for specific conserved amino acids in the catalytic region of Fat facets and also indicate that regions of the protein both N- and C-terminal to the catalytic region are required for Fat facets function.
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PMID:In vivo Structure/Function analysis of the Drosophila fat facets deubiquitinating enzyme gene. 1110 77

The human isopeptidase T (isoT) is a zinc-binding deubiquitinating enzyme involved in the disassembly of free K48-linked polyubiquitin chains into ubiquitin monomers. The catalytic site of this enzyme is thought to be composed of Cys335, Asp435, His786 and His795. These four residues were site-directed mutagenized. None of the mutants were able to cleave a peptide-linked ubiquitin dimer. Similarly, C335S, D435N and H795N mutants had virtually no activity against a K48-linked isopeptide ubiquitin dimer, which is an isoT-specific substrate that mimics the K48-linked polyubiquitin chains. On the other hand, the H786N mutant retained a partial activity toward the K48-linked substrate, suggesting that the His786 residue might not be part of the catalytic site. None of the mutations significantly affected the capacity of isoT to bind ubiquitin and zinc. Thus, the catalytic site of UBPs could resemble that of other cysteine proteases, which contain one Cys, one Asp and one His.
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PMID:Further characterization of the putative human isopeptidase T catalytic site. 1243 95


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