title: Phosphorylation of ubiquitin at Ser65 affects its polymerization, targets, and proteome-wide turnover.
abstract: Ubiquitylation is an essential post-translational modification that regulates numerous cellular processes, most notably protein degradation. Ubiquitin can itself be phosphorylated at nearly every serine, threonine, and tyrosine residue. However, the effect of this modification on ubiquitin function is largely unknown. Here, we characterized the effects of phosphorylation of yeast ubiquitin at serine 65 in vivo and in vitro. We find this post-translational modification to be regulated under oxidative stress, occurring concomitantly with the restructuring of the ubiquitin landscape into a highly polymeric state. Phosphomimetic mutation of S65 recapitulates the oxidative stress phenotype, causing a dramatic accumulation of ubiquitylated proteins and a proteome-wide reduction of protein turnover rates. Importantly, this mutation impacts ubiquitin chain disassembly, chain linkage distribution, ubiquitin interactions, and substrate targeting. These results demonstrate that phosphorylation is an additional mode of ubiquitin regulation with broad implications in cellular physiology.
authors.affiliation: Department of Genome Sciences, University of Washington, Seattle, WA, USA.
authors.forename: Danielle L
authors.initials: DL
authors.lastname: Swaney
authors.affiliation: Department of Genome Sciences, University of Washington, Seattle, WA, USA.
authors.forename: Ricard A
authors.initials: RA
authors.lastname: Rodríguez-Mias
authors.affiliation: Department of Genome Sciences, University of Washington, Seattle, WA, USA jvillen@u.washington.edu.
authors.forename: Judit
authors.initials: J
authors.lastname: Villén
mesh_terms: D002470:Cell Survival; D013058:Mass Spectrometry; D009154:Mutation; D018384:Oxidative Stress; D010766:Phosphorylation; D058105:Polymerization; D054730:Protein Interaction Domains and Motifs; D011499:Protein Processing, Post-Translational; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013312:Stress, Physiological / Q000235:genetics; D025801:Ubiquitin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination / Q000235:genetics
chemical_list: D020543:Proteome; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin
keywords: oxidative stress; phosphorylation; protein turnover; proteomics; ubiquitin