title: Mitotic phosphorylation of Cdc25B Ser321 disrupts 14-3-3 binding to the high affinity Ser323 site.
abstract: Cdc25B is a key regulator of entry into mitosis, and its activity and localization are regulated by binding of the 14-3-3 dimer. There are three 14-3-3 binding sites on Cdc25B, with Ser(323) being the highest affinity binding and is highly homologous to the Ser(216) 14-3-3 binding site on Cdc25C. Loss of 14-3-3 binding to Ser(323) increases cyclin/Cdk substrate access to the catalytic site, thereby increasing its activity. It also affects the localization of Cdc25B. Thus, phosphorylation and 14-3-3 binding to this site is essential for down-regulating Cdc25B activity, blocking its mitosis promoting function. The question of how this inhibitory signal is relieved to allow Cdc25B activation and entry into mitosis is yet to be resolved. Here, we show that Ser(323) phosphorylation is maintained into mitosis, but phosphorylation of Ser(321) disrupts 14-3-3 binding to Ser(323), mimicking the effect of inhibiting Ser(323) phosphorylation on both Cdc25B activity and localization. The unphosphorylated Ser(321) appears to have a role in stabilizing 14-3-3 binding to Ser(323), and loss of the Ser hydroxyl group appears to be sufficient to significantly reduce 14-3-3 binding. A consequence of loss of 14-3-3 binding is dephosphorylation of Ser(323). Ser(321) is phosphorylated in mitosis by Cdk1. The mitotic phosphorylation of Ser(321) acts to maintain full activation of Cdc25B by disrupting 14-3-3 binding to Ser(323) and enhancing the dephosphorylation of Ser(323) to block 14-3-3 binding to this site.
authors.affiliation: University of Queensland Diamantina Institute, Princess Alexandra Hospital, Brisbane, Queensland 4102, Australia.
authors.forename: Puji
authors.initials: P
authors.lastname: Astuti
authors.forename: Rose
authors.initials: R
authors.lastname: Boutros
authors.forename: Bernard
authors.initials: B
authors.lastname: Ducommun
authors.forename: Brian
authors.initials: B
authors.lastname: Gabrielli
mesh_terms: D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000502:physiology; D015536:Down-Regulation / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D010766:Phosphorylation / Q000502:physiology; D011485:Protein Binding / Q000502:physiology; D021381:Protein Transport / Q000502:physiology; D012694:Serine / Q000235:genetics / Q000378:metabolism; D020687:cdc25 Phosphatases / Q000235:genetics / Q000378:metabolism*
chemical_list: D048948:14-3-3 Proteins; D012694:Serine; C517008:CDC25B protein, human; D020687:cdc25 Phosphatases
references.citation: Mol Biol Cell. 2009 Apr;20(8):2186-95
references.pmid: 19244340
references.citation: Oncogene. 2001 Feb 22;20(8):921-32
references.pmid: 11314027
references.citation: J Cell Sci. 1996 May;109 ( Pt 5):1081-93
references.pmid: 8743955
references.citation: Nat Rev Cancer. 2007 Jul;7(7):495-507
references.pmid: 17568790
references.citation: Mol Cell. 2004 Sep 10;15(5):799-811
references.pmid: 15350223
references.citation: J Cell Biol. 2005 Oct 10;171(1):35-45
references.pmid: 16216921
references.citation: Oncogene. 2001 Jul 19;20(32):4393-401
references.pmid: 11466620
references.citation: Oncogene. 2008 Jul 17;27(31):4261-8
references.pmid: 18372919
references.citation: Mol Biol Cell. 2006 Apr;17(4):1779-89
references.pmid: 16467385
references.citation: J Proteome Res. 2008 Mar;7(3):1264-73
references.pmid: 18237113
references.citation: J Cell Sci. 2004 Jun 15;117(Pt 14):3011-20
references.pmid: 15173315
references.citation: J Biol Chem. 2003 Aug 1;278(31):28580-7
references.pmid: 12764136
references.citation: J Cell Sci. 1998 Aug;111 ( Pt 16):2445-53
references.pmid: 9683638
references.citation: J Biol Chem. 2009 Dec 4;284(49):33781-8
references.pmid: 19801682
references.citation: Semin Cancer Biol. 2006 Jun;16(3):173-82
references.pmid: 16678437
references.citation: Nat Cell Biol. 2003 Jun;5(6):545-51
references.pmid: 12766774
references.citation: Proc Natl Acad Sci U S A. 2004 Nov 23;101(47):16437-41
references.pmid: 15534213
references.citation: J Biol Chem. 2002 May 3;277(18):16102-15
references.pmid: 11821419
references.citation: J Biol Chem. 1997 Dec 26;272(52):32731-4
references.pmid: 9407044
references.citation: Proc Natl Acad Sci U S A. 2006 Jul 11;103(28):10660-5
references.pmid: 16818887
references.citation: J Biol Chem. 1997 Nov 7;272(45):28607-14
references.pmid: 9353326
references.citation: Mol Cell Biol. 1994 Mar;14(3):1594-602
references.pmid: 8114697
references.citation: J Biol Chem. 1996 Mar 1;271(9):5118-24
references.pmid: 8617791
references.citation: Biochemistry. 2005 Dec 20;44(50):16563-73
references.pmid: 16342947
references.citation: J Cell Biol. 1999 Aug 9;146(3):573-84
references.pmid: 10444066
references.citation: Mol Cancer Ther. 2006 Jun;5(6):1446-51
references.pmid: 16818502
references.citation: Biochem J. 1993 Dec 15;296 ( Pt 3):843-9
references.pmid: 8280084
references.citation: Mol Biol Cell. 1994 Sep;5(9):989-1001
references.pmid: 7841526
references.citation: Biochem Biophys Res Commun. 1999 Jul 5;260(2):510-5
references.pmid: 10403798
references.citation: Oncogene. 2003 Jan 16;22(2):220-32
references.pmid: 12527891
references.citation: Oncogene. 1997 May 22;14(20):2485-95
references.pmid: 9188863
references.citation: J Cell Sci. 2006 Oct 15;119(Pt 20):4269-75
references.pmid: 17003105
references.citation: J Cell Sci. 2004 Oct 1;117(Pt 21):4979-90
references.pmid: 15456846