Cdc25A is a novel phosphatase functioning early in the cell cycle.

S Jinno

K Suto

A Nagata

M Igarashi

Y Kanaoka

H Nojima

H Okayama

Abstract

The cdc25+ tyrosine phosphatase is a key mitotic inducer of the fission yeast Schizosaccharomyces pombe, controlling the timing of the initiation of mitosis. Mammals contain at least three cdc25+ homologues called cdc25A, cdc25B and cdc25C. In this study we investigate the biological function of cdc25A. Although very potent in rescuing the S.pombe cdc25 mutant, cdc25A is less structurally related to the S.pombe enzyme. Northern and Western blotting detection reveals that unlike cdc25B, cdc25C and cdc2, cdc25A is predominantly expressed in late G1. Moreover, immunodepletion of cdc25A in rat cells by microinjection of a specific antibody effectively blocks their cell cycle progression from G1 into the S phase, as determined by laser scanning single cell cytometry. These results indicate that cdc25A is not a mitotic regulator but a novel phosphatase that plays a crucial role in the start of the cell cycle. In view of its strong ability to activate cdc2 kinase and its specific expression in late G1, cdc2-related kinases functioning early in the cell cycle may be targets for this phosphatase.

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Selected References

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Assoian RK, Komoriya A, Meyers CA, Miller DM, Sporn MB. Transforming growth factor-beta in human platelets. Identification of a major storage site, purification, and characterization. J Biol Chem. 1983 Jun 10;258(11):7155–7160. [PubMed] [Google Scholar]
Booher RN, Alfa CE, Hyams JS, Beach DH. The fission yeast cdc2/cdc13/suc1 protein kinase: regulation of catalytic activity and nuclear localization. Cell. 1989 Aug 11;58(3):485–497. [PubMed] [Google Scholar]
Chen C, Okayama H. High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol. 1987 Aug;7(8):2745–2752.[PMC free article] [PubMed] [Google Scholar]
Dulić V, Lees E, Reed SI. Association of human cyclin E with a periodic G1-S phase protein kinase. Science. 1992 Sep 25;257(5078):1958–1961. [PubMed] [Google Scholar]
Elledge SJ, Spottswood MR. A new human p34 protein kinase, CDK2, identified by complementation of a cdc28 mutation in Saccharomyces cerevisiae, is a homolog of Xenopus Eg1. EMBO J. 1991 Sep;10(9):2653–2659.[PMC free article] [PubMed] [Google Scholar]
Enoch T, Nurse P. Mutation of fission yeast cell cycle control genes abolishes dependence of mitosis on DNA replication. Cell. 1990 Feb 23;60(4):665–673. [PubMed] [Google Scholar]
Fang F, Newport JW. Evidence that the G1-S and G2-M transitions are controlled by different cdc2 proteins in higher eukaryotes. Cell. 1991 Aug 23;66(4):731–742. [PubMed] [Google Scholar]
Forsburg SL, Nurse P. Cell cycle regulation in the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe. Annu Rev Cell Biol. 1991;7:227–256. [PubMed] [Google Scholar]
Galaktionov K, Beach D. Specific activation of cdc25 tyrosine phosphatases by B-type cyclins: evidence for multiple roles of mitotic cyclins. Cell. 1991 Dec 20;67(6):1181–1194. [PubMed] [Google Scholar]
Gautier J, Solomon MJ, Booher RN, Bazan JF, Kirschner MW. cdc25 is a specific tyrosine phosphatase that directly activates p34cdc2. Cell. 1991 Oct 4;67(1):197–211. [PubMed] [Google Scholar]
Gould KL, Moreno S, Tonks NK, Nurse P. Complementation of the mitotic activator, p80cdc25, by a human protein-tyrosine phosphatase. Science. 1990 Dec 14;250(4987):1573–1576. [PubMed] [Google Scholar]
Gu Y, Rosenblatt J, Morgan DO. Cell cycle regulation of CDK2 activity by phosphorylation of Thr160 and Tyr15. EMBO J. 1992 Nov;11(11):3995–4005.[PMC free article] [PubMed] [Google Scholar]
Hagan I, Hayles J, Nurse P. Cloning and sequencing of the cyclin-related cdc13+ gene and a cytological study of its role in fission yeast mitosis. J Cell Sci. 1988 Dec;91(Pt 4):587–595. [PubMed] [Google Scholar]
Hoffmann I, Clarke PR, Marcote MJ, Karsenti E, Draetta G. Phosphorylation and activation of human cdc25-C by cdc2--cyclin B and its involvement in the self-amplification of MPF at mitosis. EMBO J. 1993 Jan;12(1):53–63.[PMC free article] [PubMed] [Google Scholar]
Honda R, Ohba Y, Nagata A, Okayama H, Yasuda H. Dephosphorylation of human p34cdc2 kinase on both Thr-14 and Tyr-15 by human cdc25B phosphatase. FEBS Lett. 1993 Mar 8;318(3):331–334. [PubMed] [Google Scholar]
Igarashi M, Nagata A, Jinno S, Suto K, Okayama H. Wee1(+)-like gene in human cells. Nature. 1991 Sep 5;353(6339):80–83. [PubMed] [Google Scholar]
Inoue H, Nojima H, Okayama H. High efficiency transformation of Escherichia coli with plasmids. Gene. 1990 Nov 30;96(1):23–28. [PubMed] [Google Scholar]
Koff A, Cross F, Fisher A, Schumacher J, Leguellec K, Philippe M, Roberts JM. Human cyclin E, a new cyclin that interacts with two members of the CDC2 gene family. Cell. 1991 Sep 20;66(6):1217–1228. [PubMed] [Google Scholar]
Koff A, Giordano A, Desai D, Yamashita K, Harper JW, Elledge S, Nishimoto T, Morgan DO, Franza BR, Roberts JM. Formation and activation of a cyclin E-cdk2 complex during the G1 phase of the human cell cycle. Science. 1992 Sep 18;257(5077):1689–1694. [PubMed] [Google Scholar]
Krek W, Nigg EA. Differential phosphorylation of vertebrate p34cdc2 kinase at the G1/S and G2/M transitions of the cell cycle: identification of major phosphorylation sites. EMBO J. 1991 Feb;10(2):305–316.[PMC free article] [PubMed] [Google Scholar]
Lew DJ, Dulić V, Reed SI. Isolation of three novel human cyclins by rescue of G1 cyclin (Cln) function in yeast. Cell. 1991 Sep 20;66(6):1197–1206. [PubMed] [Google Scholar]
Lundgren K, Walworth N, Booher R, Dembski M, Kirschner M, Beach D. mik1 and wee1 cooperate in the inhibitory tyrosine phosphorylation of cdc2. Cell. 1991 Mar 22;64(6):1111–1122. [PubMed] [Google Scholar]
Matsushime H, Roussel MF, Ashmun RA, Sherr CJ. Colony-stimulating factor 1 regulates novel cyclins during the G1 phase of the cell cycle. Cell. 1991 May 17;65(4):701–713. [PubMed] [Google Scholar]
Matsushime H, Ewen ME, Strom DK, Kato JY, Hanks SK, Roussel MF, Sherr CJ. Identification and properties of an atypical catalytic subunit (p34PSK-J3/cdk4) for mammalian D type G1 cyclins. Cell. 1992 Oct 16;71(2):323–334. [PubMed] [Google Scholar]
Meyerson M, Enders GH, Wu CL, Su LK, Gorka C, Nelson C, Harlow E, Tsai LH. A family of human cdc2-related protein kinases. EMBO J. 1992 Aug;11(8):2909–2917.[PMC free article] [PubMed] [Google Scholar]
Nagata A, Igarashi M, Jinno S, Suto K, Okayama H. An additional homolog of the fission yeast cdc25+ gene occurs in humans and is highly expressed in some cancer cells. New Biol. 1991 Oct;3(10):959–968. [PubMed] [Google Scholar]
Ninomiya-Tsuji J, Nomoto S, Yasuda H, Reed SI, Matsumoto K. Cloning of a human cDNA encoding a CDC2-related kinase by complementation of a budding yeast cdc28 mutation. Proc Natl Acad Sci U S A. 1991 Oct 15;88(20):9006–9010.[PMC free article] [PubMed] [Google Scholar]
Okayama H, Kawaichi M, Brownstein M, Lee F, Yokota T, Arai K. High-efficiency cloning of full-length cDNA; construction and screening of cDNA expression libraries for mammalian cells. Methods Enzymol. 1987;154:3–28. [PubMed] [Google Scholar]
Okazaki K, Okazaki N, Kume K, Jinno S, Tanaka K, Okayama H. High-frequency transformation method and library transducing vectors for cloning mammalian cDNAs by trans-complementation of Schizosaccharomyces pombe. Nucleic Acids Res. 1990 Nov 25;18(22):6485–6489.[PMC free article] [PubMed] [Google Scholar]
Parker LL, Piwnica-Worms H. Inactivation of the p34cdc2-cyclin B complex by the human WEE1 tyrosine kinase. Science. 1992 Sep 25;257(5078):1955–1957. [PubMed] [Google Scholar]
Rosenblatt J, Gu Y, Morgan DO. Human cyclin-dependent kinase 2 is activated during the S and G2 phases of the cell cycle and associates with cyclin A. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2824–2828.[PMC free article] [PubMed] [Google Scholar]
Russell P, Nurse P. cdc25+ functions as an inducer in the mitotic control of fission yeast. Cell. 1986 Apr 11;45(1):145–153. [PubMed] [Google Scholar]
Russell P, Nurse P. Negative regulation of mitosis by wee1+, a gene encoding a protein kinase homolog. Cell. 1987 May 22;49(4):559–567. [PubMed] [Google Scholar]
Sadhu K, Reed SI, Richardson H, Russell P. Human homolog of fission yeast cdc25 mitotic inducer is predominantly expressed in G2. Proc Natl Acad Sci U S A. 1990 Jul;87(13):5139–5143.[PMC free article] [PubMed] [Google Scholar]
Sebastian B, Kakizuka A, Hunter T. Cdc25M2 activation of cyclin-dependent kinases by dephosphorylation of threonine-14 and tyrosine-15. Proc Natl Acad Sci U S A. 1993 Apr 15;90(8):3521–3524.[PMC free article] [PubMed] [Google Scholar]
Cyclin in fission yeast. Cell. 1988 Sep 9;54(6):738–740. [PubMed] [Google Scholar]
Solomon MJ, Glotzer M, Lee TH, Philippe M, Kirschner MW. Cyclin activation of p34cdc2. Cell. 1990 Nov 30;63(5):1013–1024. [PubMed] [Google Scholar]
Strausfeld U, Labbé JC, Fesquet D, Cavadore JC, Picard A, Sadhu K, Russell P, Dorée M. Dephosphorylation and activation of a p34cdc2/cyclin B complex in vitro by human CDC25 protein. Nature. 1991 May 16;351(6323):242–245. [PubMed] [Google Scholar]
Th'ng JP, Wright PS, Hamaguchi J, Lee MG, Norbury CJ, Nurse P, Bradbury EM. The FT210 cell line is a mouse G2 phase mutant with a temperature-sensitive CDC2 gene product. Cell. 1990 Oct 19;63(2):313–324. [PubMed] [Google Scholar]
Vindeløv LL, Christensen IJ, Nissen NI. A detergent-trypsin method for the preparation of nuclei for flow cytometric DNA analysis. Cytometry. 1983 Mar;3(5):323–327. [PubMed] [Google Scholar]
Xiong Y, Connolly T, Futcher B, Beach D. Human D-type cyclin. Cell. 1991 May 17;65(4):691–699. [PubMed] [Google Scholar]
Yamanashi Y, Fukui Y, Wongsasant B, Kinoshita Y, Ichimori Y, Toyoshima K, Yamamoto T. Activation of Src-like protein-tyrosine kinase Lyn and its association with phosphatidylinositol 3-kinase upon B-cell antigen receptor-mediated signaling. Proc Natl Acad Sci U S A. 1992 Feb 1;89(3):1118–1122.[PMC free article] [PubMed] [Google Scholar]

Department of Molecular Genetics, Osaka University, Japan.

Abstract