title: A quantitative atlas of mitotic phosphorylation.
abstract: The eukaryotic cell division cycle is characterized by a sequence of orderly and highly regulated events resulting in the duplication and separation of all cellular material into two newly formed daughter cells. Protein phosphorylation by cyclin-dependent kinases (CDKs) drives this cycle. To gain further insight into how phosphorylation regulates the cell cycle, we sought to identify proteins whose phosphorylation is cell cycle regulated. Using stable isotope labeling along with a two-step strategy for phosphopeptide enrichment and high mass accuracy mass spectrometry, we examined protein phosphorylation in a human cell line arrested in the G(1) and mitotic phases of the cell cycle. We report the identification of >14,000 different phosphorylation events, more than half of which, to our knowledge, have not been described in the literature, along with relative quantitative data for the majority of these sites. We observed >1,000 proteins with increased phosphorylation in mitosis including many known cell cycle regulators. The majority of sites on regulated phosphopeptides lie in [S/T]P motifs, the minimum required sequence for CDKs, suggesting that many of the proteins may be CDK substrates. Analysis of non-proline site-containing phosphopeptides identified two unique motifs that suggest there are at least two undiscovered mitotic kinases.
authors.affiliation: Department of Cell Biology, Harvard University Medical School, 240 Longwood Avenue, Boston, MA 02115, USA.
authors.forename: Noah
authors.initials: N
authors.lastname: Dephoure
authors.forename: Chunshui
authors.initials: C
authors.lastname: Zhou
authors.forename: Judit
authors.initials: J
authors.lastname: Villén
authors.forename: Sean A
authors.initials: SA
authors.lastname: Beausoleil
authors.forename: Corey E
authors.initials: CE
authors.lastname: Bakalarski
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Elledge
authors.forename: Steven P
authors.initials: SP
authors.lastname: Gygi
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000378:metabolism*; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D013058:Mass Spectrometry; D008938:Mitosis / Q000502:physiology*; D008969:Molecular Sequence Data; D010748:Phosphopeptides / Q000378:metabolism*; D010766:Phosphorylation; D040901:Proteomics
chemical_list: D018797:Cell Cycle Proteins; D010748:Phosphopeptides; D018844:Cyclin-Dependent Kinases