title: Recognition and cleavage of related to ubiquitin 1 (Rub1) and Rub1-ubiquitin chains by components of the ubiquitin-proteasome system.
abstract: Of all ubiquitin-like proteins, Rub1 (Nedd8 in mammals) is the closest kin of ubiquitin. We show via NMR that structurally, Rub1 and ubiquitin are fundamentally similar as well. Despite these profound similarities, the prevalence of Rub1/Nedd8 and of ubiquitin as modifiers of the proteome is starkly different, and their attachments to specific substrates perform different functions. Recently, some proteins, including p53, p73, EGFR, caspase-7, and Parkin, have been shown to be modified by both Rub1/Nedd8 and ubiquitin within cells. To understand whether and how it might be possible to distinguish among the same target protein modified by Rub1 or ubiquitin or both, we examined whether ubiquitin receptors can differentiate between Rub1 and ubiquitin. Surprisingly, Rub1 interacts with proteasome ubiquitin-shuttle proteins comparably to ubiquitin but binds more weakly to a proteasomal ubiquitin receptor Rpn10. We identified Rub1-ubiquitin heteromers in yeast and Nedd8-Ub heteromers in human cells. We validate that in human cells and in vitro, human Rub1 (Nedd8) forms chains with ubiquitin where it acts as a chain terminator. Interestingly, enzymatically assembled K48-linked Rub1-ubiquitin heterodimers are recognized by various proteasomal ubiquitin shuttles and receptors comparably to K48-linked ubiquitin homodimers. Furthermore, these heterologous chains are cleaved by COP9 signalosome or 26S proteasome. A derubylation function of the proteasome expands the repertoire of its enzymatic activities. In contrast, Rub1 conjugates may be somewhat resilient to the actions of other canonical deubiquitinating enzymes. Taken together, these findings suggest that once Rub1/Nedd8 is channeled into ubiquitin pathways, it is recognized essentially like ubiquitin.
authors.affiliation: Department of Chemistry and Biochemistry, Center for Biomolecular Structure and Organization, University of Maryland, College Park, MD 20742, USA.
authors.forename: Rajesh K
authors.initials: RK
authors.lastname: Singh
authors.forename: Sylvia
authors.initials: S
authors.lastname: Zerath
authors.forename: Oded
authors.initials: O
authors.lastname: Kleifeld
authors.forename: Martin
authors.initials: M
authors.lastname: Scheffner
authors.forename: Michael H
authors.initials: MH
authors.lastname: Glickman
authors.forename: David
authors.initials: D
authors.lastname: Fushman
mesh_terms: D000595:Amino Acid Sequence; D000075686:COP9 Signalosome Complex; D002460:Cell Line; D006801:Humans; D046912:Multiprotein Complexes / Q000378:metabolism; D000075687:NEDD8 Protein; D010447:Peptide Hydrolases / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D016601:RNA-Binding Proteins; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism*
chemical_list: D046912:Multiprotein Complexes; D000075687:NEDD8 Protein; C082860:NEDD8 protein, human; C498506:PSMD4 protein, human; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D014452:Ubiquitins; D044767:Ubiquitin-Protein Ligases; D010447:Peptide Hydrolases; D000075686:COP9 Signalosome Complex; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease