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SKP Cullin F-Box Protein LigasesF-Box ProteinsCullin ProteinsUbiquitin-Protein LigasesLigasesS-Phase Kinase-Associated ProteinsLuteoviridaebeta-Transducin Repeat-Containing ProteinsCell Cycle ProteinsF-Box MotifsDEAD Box Protein 20DNA LigasesDEAD-box RNA HelicasesUbiquitinationMolecular Sequence DataAmino Acid SequenceRNA HelicasesProtein BindingUbiquitinUbiquitinsTranscription FactorsArabidopsis ProteinsUbiquitin-Conjugating EnzymesForkhead Transcription FactorsHMGB1 ProteinUbiquitin-Protein Ligase ComplexesDNA-Binding ProteinsTATA BoxPolynucleotide LigasesSequence Homology, Amino AcidProtein Structure, TertiaryMultiprotein ComplexesRNA NucleotidyltransferasesBase SequenceMinichromosome Maintenance 1 ProteinHigh Mobility Group ProteinsProteasome Endopeptidase ComplexPeptide SynthasesMutationMADS Domain ProteinsTwo-Hybrid System TechniquesSaccharomyces cerevisiae ProteinsCarrier ProteinsRING Finger DomainsCoenzyme A LigasesApc11 Subunit, Anaphase-Promoting Complex-CyclosomeBinding SitesAnaphase-Promoting Complex-CyclosomeCell LineRats, Inbred F344ProteolysisNuclear ProteinsHeLa CellsSequence AlignmentAmino Acid MotifsRNA Ligase (ATP)Saccharomyces cerevisiaeSuppressor of Cytokine Signaling ProteinsSignal TransductionRNA InterferenceHMGB ProteinsCloning, MolecularRepressor ProteinsRNA, MessengerUbiquitin-Activating EnzymesConserved SequenceTranscription, GeneticPolyubiquitinArabidopsisGene Expression RegulationF FactorPeptide HydrolasesPromoter Regions, GeneticRNA-Binding ProteinsCyclin-Dependent Kinase Inhibitor p27Cell CycleRecombinant ProteinsSubstrate SpecificityApc2 Subunit, Anaphase-Promoting Complex-CyclosomeDrosophila ProteinsRecombinant Fusion ProteinsModels, MolecularF2-IsoprostanesEndosomal Sorting Complexes Required for TransportCell NucleusRibonucleoprotein, U2 Small NuclearGene Expression Regulation, PlantHepatocyte Nuclear Factor 3-alphaRNA, Small InterferingTransfectionProtein Processing, Post-TranslationalPhenotypeSOXC Transcription FactorsFungal ProteinsSex-Determining Region Y ProteinImmunoprecipitationDNAPseudohypoaldosteronismAdenosine TriphosphatasesPlant Proteins

Inducible degradation of IkappaBalpha by the proteasome requires interaction with the F-box protein h-betaTrCP. (1/624)

Activation of NF-kappaB transcription factors requires phosphorylation and ubiquitin-proteasome-dependent degradation of IkappaB proteins. We provide evidence that a human F-box protein, h-betaTrCP, a component of Skp1-Cullin-F-box protein (SCF) complexes, a new class of E3 ubiquitin ligases, is essential for inducible degradation of IkappaBalpha. betaTrCP associates with Ser32-Ser36 phosphorylated, but not with unmodified IkappaBalpha or Ser32-Ser36 phosphorylation-deficient mutants. Expression of a F-box-deleted betaTrCP inhibits IkappaBalpha degradation, promotes accumulation of phosphorylated Ser32-Ser36 IkappaBalpha, and prevents NF-kappaB-dependent transcription. Our findings indicate that betaTrCP is the adaptor protein required for IkappaBalpha recognition by the SCFbetaTrCP E3 complex that ubiquitinates IkappaBalpha and makes it a substrate for the proteasome.  (+info)

Rbx1, a component of the VHL tumor suppressor complex and SCF ubiquitin ligase. (2/624)

The von Hippel-Lindau (VHL) tumor suppressor gene is mutated in most human kidney cancers. The VHL protein is part of a complex that includes Elongin B, Elongin C, and Cullin-2, proteins associated with transcriptional elongation and ubiquitination. Here it is shown that the endogenous VHL complex in rat liver also includes Rbx1, an evolutionarily conserved protein that contains a RING-H2 fingerlike motif and that interacts with Cullins. The yeast homolog of Rbx1 is a subunit and potent activator of the Cdc53-containing SCFCdc4 ubiquitin ligase required for ubiquitination of the cyclin-dependent kinase inhibitor Sic1 and for the G1 to S cell cycle transition. These findings provide a further link between VHL and the cellular ubiquitination machinery.  (+info)

Reconstitution of G1 cyclin ubiquitination with complexes containing SCFGrr1 and Rbx1. (3/624)

Control of cyclin levels is critical for proper cell cycle regulation. In yeast, the stability of the G1 cyclin Cln1 is controlled by phosphorylation-dependent ubiquitination. Here it is shown that this reaction can be reconstituted in vitro with an SCF E3 ubiquitin ligase complex. Phosphorylated Cln1 was ubiquitinated by SCF (Skp1-Cdc53-F-box protein) complexes containing the F-box protein Grr1, Rbx1, and the E2 Cdc34. Rbx1 promotes association of Cdc34 with Cdc53 and stimulates Cdc34 auto-ubiquitination in the context of Cdc53 or SCF complexes. Rbx1, which is also a component of the von Hippel-Lindau tumor suppressor complex, may define a previously unrecognized class of E3-associated proteins.  (+info)

An F-box protein, FWD1, mediates ubiquitin-dependent proteolysis of beta-catenin. (4/624)

beta-catenin plays an essential role in the Wingless/Wnt signaling cascade and is a component of the cadherin cell adhesion complex. Deregulation of beta-catenin accumulation as a result of mutations in adenomatous polyposis coli (APC) tumor suppressor protein is believed to initiate colorectal neoplasia. beta-catenin levels are regulated by the ubiquitin-dependent proteolysis system and beta-catenin ubiquitination is preceded by phosphorylation of its N-terminal region by the glycogen synthase kinase-3beta (GSK-3beta)/Axin kinase complex. Here we show that FWD1 (the mouse homologue of Slimb/betaTrCP), an F-box/WD40-repeat protein, specifically formed a multi-molecular complex with beta-catenin, Axin, GSK-3beta and APC. Mutations at the signal-induced phosphorylation site of beta-catenin inhibited its association with FWD1. FWD1 facilitated ubiquitination and promoted degradation of beta-catenin, resulting in reduced cytoplasmic beta-catenin levels. In contrast, a dominant-negative mutant form of FWD1 inhibited the ubiquitination process and stabilized beta-catenin. These results suggest that the Skp1/Cullin/F-box protein FWD1 (SCFFWD1)-ubiquitin ligase complex is involved in beta-catenin ubiquitination and that FWD1 serves as an intracellular receptor for phosphorylated beta-catenin. FWD1 also links the phosphorylation machinery to the ubiquitin-proteasome pathway to ensure prompt and efficient proteolysis of beta-catenin in response to external signals. SCFFWD1 may be critical for tumor development and suppression through regulation of beta-catenin protein stability.  (+info)

Recruitment of a ROC1-CUL1 ubiquitin ligase by Skp1 and HOS to catalyze the ubiquitination of I kappa B alpha. (5/624)

Activation of the transcription factor NF-kappa B in response to proinflammatory stimuli requires the phosphorylation-triggered and ubiquitin-dependent degradation of the NF-kappa B inhibitor, I kappa B alpha. Here, we show the in vitro reconstitution of the phosphorylation-dependent ubiquitination of I kappa B alpha with purified components. ROC1, a novel SCF-associated protein, is recruited by cullin 1 to form a quatemary SCFHOS-ROC1 holenzyme (with Skp1 and the beta-TRCP homolog HOS). SCFHOS-ROC1 binds IKK beta-phosphorylated I kappa B alpha and catalyzes its ubiquitination in the presence of ubiquitin, E1, and Cdc34. ROC1 plays a unique role in the ubiquitination reaction by heterodimerizing with cullin 1 to catalyze ubiquitin polymerization.  (+info)

ROC1, a homolog of APC11, represents a family of cullin partners with an associated ubiquitin ligase activity. (6/624)

We have identified two highly conserved RING finger proteins, ROC1 and ROC2, that are homologous to APC11, a subunit of the anaphase-promoting complex. ROC1 and ROC2 commonly interact with all cullins while APC11 specifically interacts with APC2, a cullin-related APC subunit. YeastROC1 encodes an essential gene whose reduced expression resulted in multiple, elongated buds and accumulation of Sic1p and Cln2p. ROC1 and APC11 immunocomplexes can catalyze isopeptide ligations to form polyubiquitin chains in an E1- and E2-dependent manner. ROC1 mutations completely abolished their ligase activity without noticeable changes in associated proteins. Ubiquitination of phosphorylated I kappa B alpha can be catalyzed by the ROC1 immunocomplex in vitro. Hence, combinations of ROC/APC11 and cullin proteins proteins potentially constitute a wide variety of ubiquitin ligases.  (+info)

HOS, a human homolog of Slimb, forms an SCF complex with Skp1 and Cullin1 and targets the phosphorylation-dependent degradation of IkappaB and beta-catenin. (7/624)

SCF E3 ubiquitin ligases mediate ubiquitination and proteasome-dependent degradation of phosphorylated substrates. We identified a human F-box/WD40 repeats protein (HOS), which is homologous to Slimb/h betaTrCP. Being a part of SCF complex with Skp1 and Cullin1, HOS specifically interacted with the phosphorylated IkappaB and beta-catenin, targeting these proteins for proteasome-dependent degradation in vivo. This targeting required Cullin1 as expression of a mutant Cullin1 abrogated the degradation of IkappaB and of beta-catenin. Mutant HOS which lacks the F-box blocked TNF alpha-induced degradation of IkappaB as well as GSK3beta-mediated degradation of beta-catenin. This mutant also inhibited NF-kappaB transactivation and increased the beta-catenin-dependent transcription activity of Tcf. These results demonstrate that SCF(HOS) E3 ubiquitin ligase regulate both NF-kappaB and beta-catenin signaling pathways.  (+info)

p27(Kip1) ubiquitination and degradation is regulated by the SCF(Skp2) complex through phosphorylated Thr187 in p27. (8/624)

Many tumorigenic processes affect cell-cycle progression by their effects on the levels of the cyclin-dependent kinase inhibitor p27(Kip1) [1,2]. The phosphorylation- and ubiquitination-dependent proteolysis of p27 is implicated in control of the G1-S transition in the cell cycle [3-6]. To determine the factors that control p27 stability, we established a cell-free extract assay that recapitulates the degradation of p27. Phosphorylation of p27 at Thr187 was essential for its degradation. Degradation was also dependent on SCF(Skp2), a protein complex implicated in targeting phosphorylated proteins for ubiquitination [7-10]. Immunodepletion of components of the complex - Cul-1, Skp1, or Skp2 - from the extract abolished p27 degradation, while addition of purified SCF(Skp2) to Skp2- depleted extract restored the capacity to degrade p27. A specific association was observed between Skp2 and a p27 carboxy-terminal peptide containing phosphorylated Thr187, but not between Skp2 and the non-phosphorylated peptide. Skp2-dependent associations between Skp1 or Cul-1 and the p27 phosphopeptide were also detected. Isolated SCF(Skp2) contained an E3 ubiquitin ligase activity towards p27. Our data thus suggest that SCF(Skp2) specifically targets p27 for degradation during cell-cycle progression.  (+info)