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Query: UNIPROT:P62988 (Ubiquitin)
 4,326 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ubiquitination of integral membrane proteins is a common posttranslational modification used to mediate endocytosis and endocytic sorting of cell surface proteins in eukaryotic cells. Ubiquitin (Ub)-binding proteins (UBPs) regulate the stability, function, and localization of ubiquitinated cell surface proteins in the endocytic pathway. Here, I report that the immunoglobulin superfamily cell adhesion molecule L1 undergoes ubiquitination and dephosphorylation on the plasma membrane upon L1 antibody-induced clustering, which mimics L1-L1 homophilic binding, and that these modifications are critical for obtaining the maximal rate of internalization and trafficking to the lysosome, but not to the proteasome. Notably, L1 antibody-induced clustering leads to the association of ubiquitinated L1 with Rabex-5, a UBP and guanine nucleotide exchange factor for Rab5, via interaction with the motif interacting with Ub (MIU) domain, but not the A20-type zinc finger domain. This interaction specifically depends on the presence of an Ub moiety on lysine residues in L1. Rabex-5 expression accelerates the internalization rates of L1(WT) and L1(Y1176A), a tyrosine-based motif mutant, but not L1(K11R), an ubiquitination-deficient mutant, leading to the accumulation of ubiquitinated L1 on endosomes. In contrast, RNA interference-mediated knockdown of Rabex-5 impairs the internalizations of L1(WT) and L1(Y1176A), but not L1(K11R) from the plasma membrane. Overall, these results provide a novel mechanistic insight into how Rabex-5 regulates internalization and postendocytic trafficking of ubiquitinated L1 destined for lysosomal degradation.
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PMID:Rabex-5 protein regulates the endocytic trafficking pathway of ubiquitinated neural cell adhesion molecule L1. 2284 90

Ubiquitin (Ub) is able to form polymeric isopeptide-linked chains through condensation of any of its seven lysine (Lys) residues with the C-terminus of an adjacent Ub monomer. Electrospray ionisation mass spectrometry (ESI-MS) of commercial in vitro-generated Lys48-linked di-Ub (Lys48-Ub(2)) revealed a major population of cyclised dimer. The absence of a free C-terminus in this population was confirmed by an inability to bind the zinc finger ubiquitin-binding domain (ZnF-UBP) of USP5/isopeptidase-T. Endogenous Ub(2) purified from skeletal muscle and cultured mammalian cells was found to contain cyclic Lys48-Ub(2), demonstrating that cyclisation of poly-Ub can also occur in vivo.
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PMID:Cyclisation of Lys48-linked diubiquitin in vitro and in vivo. 2308 66

Unanchored polyubiquitin chains are emerging as important regulators of cellular physiology with diverse roles paralleling those of substrate-conjugated polyubiquitin. However tools able to discriminate unanchored polyubiquitin chains of different isopeptide linkages have not been reported. We describe the design of a linker-optimized ubiquitin-binding domain hybrid (t-UBD) containing two UBDs, a ZnF-UBP domain in tandem with a linkage-selective UBA domain, which exploits avidity effects to afford selective recognition of unanchored Lys48-linked polyubiquitin chains. Utilizing native MS to quantitatively probe binding affinities we confirm cooperative binding of the UBDs within the synthetic protein, and desired binding specificity for Lys48-linked ubiquitin dimers. Furthermore, MS/MS analyses indicate that the t-UBD, when applied as an affinity enrichment reagent, can be used to favor the purification of endogenous unanchored Lys48-linked polyubiquitin chains from mammalian cell extracts. Our study indicates that strategies for the rational design and engineering of polyubiquitin chain-selective binding in nonbiological polymers are possible, paving the way for the generation of reagents to probe unanchored polyubiquitin chains of different linkages and more broadly the 'ubiquitome'. All MS data have been deposited in the ProteomeXchange with identifier PXD004059 (http://proteomecentral.proteomexchange.org/dataset/PXD004059).
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PMID:Mass spectrometry insights into a tandem ubiquitin-binding domain hybrid engineered for the selective recognition of unanchored polyubiquitin. 2727 99

Influenza A virus (IAV) is an enveloped virus with a segmented single-stranded negative-strand RNA genome. In general, the role of virally encapsidated host cell proteins in the viral life cycle is unclear. The virion contains abundant ubiquitin molecules some of which have been identified as unanchored polyubiquitin chains. These ubiquitin chains have been postulated to play a role in recruiting histone deacetylase 6 (HDAC6) to the cytosolic surface of late endosomes (LEs), promoting IAV uncoating via aggresome processing-a cellular machinery that disposes of protein waste. HDAC6, a class II HDAC, is unusual because it resides mostly in the cytosol instead of the nucleus. It is a unique protein consisting of two catalytic domains (CDs) and a zinc-finger ubiquitin-binding domain (ZnF-UBP) close to its C-terminus. This ZnF-UBP recognizes the unconjugated ubiquitin C-terminus (di-Gly motif) with very high affinity. Biochemical analyses showed that free di-Gly motifs are present in the form of unanchored ubiquitin inside IAV virions. These motifs are exposed following low pH-triggered viral fusion at the LEs and attract HDAC6 transiently to the cytosolic surface of vesicles. The binding of the two components promotes viral uncoating via HDAC6 interaction with cellular motor proteins dynein and myosin II and the viral M1 capsid. The cellular mechanism involved is related to aggresome processing, a pathway that promotes degradation of misfolded protein aggregates. K63-linked ubiquitin chains are thought to be the trigger for aggresome processing, though it is still not clear whether such types of chains are prevalent within the IAV capsid. Here, we present methods using purified ZnF-UBP domain of HDAC6 to immunoprecipitate viral unanchored ubiquitin chains, which can then be used for further biochemical analyses of ubiquitin chain linkage.
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PMID:Purification of Unanchored Polyubiquitin Chains from Influenza Virions. 3015 81


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