title: Dual phosphorylation controls Cdc25 phosphatases and mitotic entry.
abstract: Negative regulation of the Cdc25C protein phosphatase by phosphorylation on Ser 216, the 14-3-3-binding site, is an important regulatory mechanism used by cells to block mitotic entry under normal conditions and after DNA damage. During mitosis, Cdc25C is not phosphorylated on Ser 216 and ionizing radiation (IR) does not induce either phosphorylation of Ser 216, or binding to 14-3-3. Here, we show that Cdc25C is phosphorylated on Ser 214 during mitosis, which in turn prevents phosphorylation of Ser 216. Mutation of Ser 214 to Ala reconstitutes Ser 216 phosphorylation and 14-3-3 binding during mitosis. Introduction of exogenous Cdc25C(S214A) into HeLa cells depleted of endogenous Cdc25C results in a substantial delay to mitotic entry. This effect was fully reversed in a S214A/S216A double-mutant, implying that the inhibitory effect of S214A mutant was entirely dependent on Ser 216 phosphorylation. A similar regulatory mechanism may also apply to another mitotic phosphatase, Cdc25B, as well as mitotic phosphatases of other species, including Xenopus laevis. We propose that this pathway ensures that Cdc2 remains active once mitosis is initiated and is a key control mechanism for maintaining the proper order of cell-cycle transitions.
authors.affiliation: Gene Response Section, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Dmitry V
authors.initials: DV
authors.lastname: Bulavin
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Higashimoto
authors.forename: Zoya N
authors.initials: ZN
authors.lastname: Demidenko
authors.forename: Sarah
authors.initials: S
authors.lastname: Meek
authors.forename: Paul
authors.initials: P
authors.lastname: Graves
authors.forename: Crissy
authors.initials: C
authors.lastname: Phillips
authors.forename: Hui
authors.initials: H
authors.lastname: Zhao
authors.forename: Sally A
authors.initials: SA
authors.lastname: Moody
authors.forename: Ettore
authors.initials: E
authors.lastname: Appella
authors.forename: Helen
authors.initials: H
authors.lastname: Piwnica-Worms
authors.forename: Albert J
authors.initials: AJ
authors.lastname: Fornace
mesh_terms: D048948:14-3-3 Proteins; D000409:Alanine / Q000378:metabolism; D019943:Amino Acid Substitution; D000970:Antineoplastic Agents / Q000494:pharmacology; D004249:DNA Damage; D016195:G2 Phase / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis; D016297:Mutagenesis, Site-Directed; D015739:Nocodazole / Q000494:pharmacology; D010766:Phosphorylation; D017354:Point Mutation; D011485:Protein Binding; D011839:Radiation, Ionizing; D012694:Serine / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000737:chemistry; D020687:cdc25 Phosphatases / Q000235:genetics / Q000378:metabolism*
chemical_list: D048948:14-3-3 Proteins; D000970:Antineoplastic Agents; D012694:Serine; D014446:Tyrosine 3-Monooxygenase; D020687:cdc25 Phosphatases; D000409:Alanine; D015739:Nocodazole