title: Phosphorylation of AIB1 at mitosis is regulated by CDK1/CYCLIN B.
abstract: BACKGROUND
  Although the AIB1 oncogene has an important role during the early phase of the cell cycle as a coactivator of E2F1, little is known about its function during mitosis.
  
  
  RESULTS
  Mitotic cells isolated by nocodazole treatment as well as by shake-off revealed a post-translational modification occurring in AIB1 specifically during mitosis. This modification was sensitive to the treatment with phosphatase, suggesting its modification by phosphorylation. Using specific inhibitors and in vitro kinase assays we demonstrate that AIB1 is phosphorylated on Ser728 and Ser867 by Cdk1/cyclin B at the onset of mitosis and remains phosphorylated until exit from M phase. Differences in the sensitivity to phosphatase inhibitors suggest that PP1 mediates dephosphorylation of AIB1 at the end of mitosis. The phosphorylation of AIB1 during mitosis was not associated with ubiquitylation or degradation, as confirmed by western blotting and flow cytometry analysis. In addition, luciferase reporter assays showed that this phosphorylation did not alter the transcriptional properties of AIB1. Importantly, fluorescence microscopy and sub-cellular fractionation showed that AIB1 phosphorylation correlated with the exclusion from the condensed chromatin, thus preventing access to the promoters of AIB1-dependent genes. Phospho-specific antibodies developed against Ser728 further demonstrated the presence of phosphorylated AIB1 only in mitotic cells where it was localized preferentially in the periphery of the cell.
  
  
  CONCLUSIONS
  Collectively, our results describe a new mechanism for the regulation of AIB1 during mitosis, whereby phosphorylation of AIB1 by Cdk1 correlates with the subcellular redistribution of AIB1 from a chromatin-associated state in interphase to a more peripheral localization during mitosis. At the exit of mitosis, AIB1 is dephosphorylated, presumably by PP1. This exclusion from chromatin during mitosis may represent a mechanism for governing the transcriptional activity of AIB1.
authors.affiliation: Laboratory of Cellular and Molecular Biology, Centro de Investigación Príncipe Felipe, Valencia, Spain.
authors.forename: Macarena
authors.initials: M
authors.lastname: Ferrero
authors.forename: Juan
authors.initials: J
authors.lastname: Ferragud
authors.forename: Leonardo
authors.initials: L
authors.lastname: Orlando
authors.forename: Luz
authors.initials: L
authors.lastname: Valero
authors.forename: Manuel
authors.initials: M
authors.lastname: Sánchez del Pino
authors.forename: Rosa
authors.initials: R
authors.lastname: Farràs
authors.forename: Jaime
authors.initials: J
authors.lastname: Font de Mora
mesh_terms: D000818:Animals; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D019556:COS Cells; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D002843:Chromatin / Q000378:metabolism; D019925:Cyclin A / Q000378:metabolism; D019926:Cyclin B / Q000378:metabolism*; D050687:E2F1 Transcription Factor / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods; D006367:HeLa Cells; D006801:Humans; D008387:Marine Toxins; D008938:Mitosis; D008954:Models, Biological; D056921:Nuclear Receptor Coactivator 3 / Q000378:metabolism*; D019319:Okadaic Acid / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational
chemical_list: D002843:Chromatin; D019925:Cyclin A; D019926:Cyclin B; D050687:E2F1 Transcription Factor; D008387:Marine Toxins; D010080:Oxazoles; D019319:Okadaic Acid; C059041:calyculin A; C538866:NCOA3 protein, human; D056921:Nuclear Receptor Coactivator 3; D016203:CDC2 Protein Kinase