title: p27(Kip1) ubiquitination and degradation is regulated by the SCF(Skp2) complex through phosphorylated Thr187 in p27.
abstract: Many tumorigenic processes affect cell-cycle progression by their effects on the levels of the cyclin-dependent kinase inhibitor p27(Kip1) [1,2]. The phosphorylation- and ubiquitination-dependent proteolysis of p27 is implicated in control of the G1-S transition in the cell cycle [3-6]. To determine the factors that control p27 stability, we established a cell-free extract assay that recapitulates the degradation of p27. Phosphorylation of p27 at Thr187 was essential for its degradation. Degradation was also dependent on SCF(Skp2), a protein complex implicated in targeting phosphorylated proteins for ubiquitination [7-10]. Immunodepletion of components of the complex - Cul-1, Skp1, or Skp2 - from the extract abolished p27 degradation, while addition of purified SCF(Skp2) to Skp2- depleted extract restored the capacity to degrade p27. A specific association was observed between Skp2 and a p27 carboxy-terminal peptide containing phosphorylated Thr187, but not between Skp2 and the non-phosphorylated peptide. Skp2-dependent associations between Skp1 or Cul-1 and the p27 phosphopeptide were also detected. Isolated SCF(Skp2) contained an E3 ubiquitin ligase activity towards p27. Our data thus suggest that SCF(Skp2) specifically targets p27 for degradation during cell-cycle progression.
authors.affiliation: Department of Genetics, Yale University School of Medicine, 333 Cedar Street, New Haven, Connecticut 06520, USA.
authors.forename: L M
authors.initials: LM
authors.lastname: Tsvetkov
authors.forename: K H
authors.initials: KH
authors.lastname: Yeh
authors.forename: S J
authors.initials: SJ
authors.lastname: Lee
authors.forename: H
authors.initials: H
authors.lastname: Sun
authors.forename: H
authors.initials: H
authors.lastname: Zhang
mesh_terms: D002453:Cell Cycle; D018797:Cell Cycle Proteins; D002474:Cell-Free System; D050760:Cyclin-Dependent Kinase Inhibitor p27; D006367:HeLa Cells; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000378:metabolism*; D010453:Peptide Synthases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011995:Recombination, Genetic; D044843:SKP Cullin F-Box Protein Ligases; D013912:Threonine / Q000737:chemistry / Q000378:metabolism; D025521:Tumor Suppressor Proteins; D014452:Ubiquitins / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D008869:Microtubule-Associated Proteins; D025521:Tumor Suppressor Proteins; D014452:Ubiquitins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D013912:Threonine; D044843:SKP Cullin F-Box Protein Ligases; D010453:Peptide Synthases