title: The Rsp5 ubiquitin ligase is coupled to and antagonized by the Ubp2 deubiquitinating enzyme.
abstract: Saccharomyces cerevisiae Rsp5 is an essential HECT ubiquitin ligase involved in several biological processes. To gain further insight into regulation of this enzyme, we identified proteins that copurified with epitope-tagged Rsp5. Ubp2, a deubiquitinating enzyme, was a prominent copurifying protein. Rup1, a previously uncharacterized UBA domain protein, was required for binding of Rsp5 to Ubp2 both in vitro and in vivo. Overexpression of Ubp2 or Rup1 in the rsp5-1 mutant elicited a strong growth defect, while overexpression of a catalytically inactive Ubp2 mutant or Rup1 deleted of the UBA domain did not, suggesting an antagonistic relationship between Rsp5 and the Ubp2/Rup1 complex. Consistent with this model, rsp5-1 temperature sensitivity was suppressed by either ubp2Delta or rup1Delta mutations. Ubp2 reversed Rsp5-catalyzed substrate ubiquitination in vitro, and Rsp5 and Ubp2 preferentially assembled and disassembled, respectively, K63-linked polyubiquitin chains. Together, these results indicate that Rsp5 activity is modulated by being physically coupled to the Rup1/Ubp2 deubiquitinating enzyme complex, representing a novel mode of regulation for an HECT ubiquitin ligase.
authors.affiliation: Institute for Cellular and Molecular Biology, Section of Molecular Genetics and Microbiology, University of Texas at Austin, Austin, TX 78712, USA.
authors.forename: Younghoon
authors.initials: Y
authors.lastname: Kee
authors.forename: Nancy
authors.initials: N
authors.lastname: Lyon
authors.forename: Jon M
authors.initials: JM
authors.lastname: Huibregtse
mesh_terms: D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport; D004789:Enzyme Activation; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D043743:Ubiquitin-Protein Ligase Complexes / Q000235:genetics / Q000378:metabolism*
chemical_list: D056827:Endosomal Sorting Complexes Required for Transport; D029701:Saccharomyces cerevisiae Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D010450:Endopeptidases; C059237:ubiquitin-Nalpha-protein hydrolase; C469287:RSP5 protein, S cerevisiae