title: SKP2 associates with p130 and accelerates p130 ubiquitylation and degradation in human cells.
abstract: p130 is a member of the retinoblastoma family of pocket proteins, which includes pRB and p107. Unlike pRB and p107, p130 protein levels decrease dramatically following its hyperphosphorylation starting in the mid-G1 phase of the cell cycle. However, the mechanism leading to p130 downregulation is unknown. We have found that the proteasome inhibitor, lactacystin, inhibited p130 downregulation in T98G cells progressing through the G1/S transition and S phase and that p130 is multiubiquitylated in 293 cells. We have previously shown that ectopic expression of both cyclin D and E induces phosphorylation and downregulation of p130. Since the SKP1/Cul1/SKP2 E3 ubiquitin ligase complex mediates ubiquitylation of substrates previously phosphorylated by cyclin-dependent kinases, we investigated the potential role of this ubiquitin ligase in mediating p130 downregulation. We found that p130 interacts with SKP1, Cul-1 and SKP2 in human 293 cells. We also found that ectopic coexpression of SKP2 and p130 leads to dose-dependent downregulation of p130, reduces p130 protein half-life and induces p130 ubiquitylation in these cells. Moreover, adenoviral-mediated expression of SKP2 accelerates downregulation of endogenous hyperphosphorylated p130 in mitogen-stimulated T98G cells and primary WI38 fibroblasts. We conclude that p130 is a substrate of the SCF(SKP2) ubiquitin ligase and this E3 ligase regulates p130 abundance during the cell cycle.
authors.affiliation: Fels Institute for Cancer Research and Molecular Biology and Department of Biochemistry, Temple University School of Medicine, 3307 North Broad Street, Philadelphia, PA 19140, USA.
authors.forename: Sabyasachi
authors.initials: S
authors.lastname: Bhattacharya
authors.forename: Judit
authors.initials: J
authors.lastname: Garriga
authors.forename: Joaquim
authors.initials: J
authors.lastname: Calbó
authors.forename: Thomas
authors.initials: T
authors.lastname: Yong
authors.forename: Dale S
authors.initials: DS
authors.lastname: Haines
authors.forename: Xavier
authors.initials: X
authors.lastname: Graña
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000378:metabolism; D016193:G1 Phase / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D009097:Multienzyme Complexes / Q000378:metabolism; D010453:Peptide Synthases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D011506:Proteins; D050717:Retinoblastoma-Like Protein p130; D016196:S Phase / Q000502:physiology; D044786:S-Phase Kinase-Associated Proteins; D044843:SKP Cullin F-Box Protein Ligases; D025801:Ubiquitin / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D009097:Multienzyme Complexes; D010750:Phosphoproteins; D011506:Proteins; C493837:RBL2 protein, human; D050717:Retinoblastoma-Like Protein p130; D044786:S-Phase Kinase-Associated Proteins; D025801:Ubiquitin; D044843:SKP Cullin F-Box Protein Ligases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D010453:Peptide Synthases