title: A role for ubiquitinylation and the cytosolic proteasome in turnover of mitochondrial uncoupling protein 1 (UCP1).
abstract: In this study we show that mitochondrial uncoupling protein 1 (UCP1) in brown adipose tissue (BAT) and thymus mitochondria can be ubiquitinylated and degraded by the cytosolic proteasome. Using a ubiquitin conjugating system, we show that UCP1 can be ubiquitinylated in vitro. We demonstrate that UCP1 is ubiquitinylated in vivo using isolated mitochondria from brown adipose tissue, thymus and whole brown adipocytes. Using an in vitro ubiquitin conjugating-proteasome degradation system, we show that the cytosolic proteasome can degrade UCP1 at a rate commensurate with the half-life of UCP1 (i.e. 30-72h in brown adipocytes and ~3h, in thymocytes). In addition, we demonstrate that the cytoplasmic proteasome is required for UCP1 degradation from mitochondria that the process is inhibited by the proteasome inhibitor MG132 and that dissipation of the mitochondrial membrane potential inhibits degradation of UCP1. There also appears to be a greater amount of ubiquitinylated UCP1 associated with BAT mitochondria from cold-acclimated animals. We have also identified (using immunoprecipitation coupled with mass spectrometry) ubiquitinylated proteins with molecular masses greater than 32kDa, as being UCP1. We conclude that there is a role for ubiquitinylation and the cytosolic proteasome in turnover of mitochondrial UCP1. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012).
authors.affiliation: School of Biochemistry and Immunology, Trinity College Dublin, Dublin 2, Ireland.
authors.forename: Kieran J
authors.initials: KJ
authors.lastname: Clarke
authors.forename: Alison E
authors.initials: AE
authors.lastname: Adams
authors.forename: Lars H
authors.initials: LH
authors.lastname: Manzke
authors.forename: Terry W
authors.initials: TW
authors.lastname: Pearson
authors.forename: Christoph H
authors.initials: CH
authors.lastname: Borchers
authors.forename: Richard K
authors.initials: RK
authors.lastname: Porter
mesh_terms: D000064:Acclimatization / Q000187:drug effects / Q000502:physiology; D052437:Adipocytes, Brown / Q000201:enzymology; D002001:Adipose Tissue, Brown / Q000201:enzymology; D000818:Animals; D003080:Cold Temperature; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D007473:Ion Channels / Q000378:metabolism*; D007976:Leupeptins / Q000494:pharmacology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects / Q000502:physiology*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D061988:Proteasome Inhibitors / Q000494:pharmacology; D059748:Proteolysis; D051381:Rats; D017208:Rats, Wistar; D013950:Thymus Gland / Q000201:enzymology; D013997:Time Factors; D054875:Ubiquitination / Q000187:drug effects / Q000502:physiology*; D000071256:Uncoupling Protein 1
chemical_list: D015853:Cysteine Proteinase Inhibitors; D007473:Ion Channels; D007976:Leupeptins; D024101:Mitochondrial Proteins; D061988:Proteasome Inhibitors; C000604373:Ucp1 protein, rat; D000071256:Uncoupling Protein 1; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde