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. 1991 Oct 2;115(2):337–344. doi: 10.1083/jcb.115.2.337

Okadaic acid mimics a nuclear component required for cyclin B-cdc2 kinase microinjection to drive starfish oocytes into M phase

PMCID: PMC2289148  PMID: 1655804

Abstract

G2-arrested oocytes contain cdc2 kinase as an inactive cyclin B-cdc2 complex. When a small amount of highly purified and active cdc2 kinase, prepared from starfish oocytes at first meiotic metaphase, is microinjected into Xenopus oocytes, it induces activation of the inactive endogenous complex and, as a consequence, drives the recipient oocytes into M phase. In contrast, the microinjected kinase undergoes rapid inactivation in starfish oocytes, which remain arrested at G2. Endogenous cdc2 kinase becomes activated in both nucleated and enucleated starfish oocytes injected with cytoplasm taken from maturing oocytes at the time of nuclear envelope breakdown, but only cytoplasm taken from nucleated oocytes becomes able thereafter to release second recipient oocytes from G2 arrest, and thus contains M phase-promoting factor (MPF) activity. Both nucleated and enucleated starfish oocytes produce MPF activity when type 2A phosphatase is blocked by okadaic acid. If type 2A phosphatase is only partially inhibited, neither nucleated nor enucleated oocytes produce MPF activity, although both do so if purified cdc2 kinase is subsequently injected as a primer to activate the endogenous kinase. The nucleus of starfish oocytes contains an inhibitor of type 2A phosphatase, but neither active nor inactive cdc2 kinase. Microinjection of the content of a nucleus into the cytoplasm of G2-arrested starfish oocytes activates endogenous cdc2 kinase, produces MPF activity, and drives the recipient oocytes into M phase. Together, these results show that the MPF amplification loop is controlled, both positively and negatively, by cdc2 kinase and type 2A phosphatase, respectively. Activation of the MPF amplification loop in starfish requires a nuclear component to inhibit type 2A phosphatase in cytoplasm.

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Selected References

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  1. Aitken A., Cohen P. Isolation and characterisation of active fragments of protein phosphatase inhibitor-1 from rabbit skeletal muscle. FEBS Lett. 1982 Oct 4;147(1):54–58. doi: 10.1016/0014-5793(82)81010-7. [DOI] [PubMed] [Google Scholar]
  2. Beavo J. A., Bechtel P. J., Krebs E. G. Preparation of homogeneous cyclic AMP-dependent protein kinase(s) and its subunits from rabbit skeletal muscle. Methods Enzymol. 1974;38:299–308. doi: 10.1016/0076-6879(74)38046-9. [DOI] [PubMed] [Google Scholar]
  3. Belenguer P., Caizergues-Ferrer M., Labbé J. C., Dorée M., Amalric F. Mitosis-specific phosphorylation of nucleolin by p34cdc2 protein kinase. Mol Cell Biol. 1990 Jul;10(7):3607–3618. doi: 10.1128/mcb.10.7.3607. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chou Y. H., Bischoff J. R., Beach D., Goldman R. D. Intermediate filament reorganization during mitosis is mediated by p34cdc2 phosphorylation of vimentin. Cell. 1990 Sep 21;62(6):1063–1071. doi: 10.1016/0092-8674(90)90384-q. [DOI] [PubMed] [Google Scholar]
  5. Cohen P., Holmes C. F., Tsukitani Y. Okadaic acid: a new probe for the study of cellular regulation. Trends Biochem Sci. 1990 Mar;15(3):98–102. doi: 10.1016/0968-0004(90)90192-e. [DOI] [PubMed] [Google Scholar]
  6. Cormier P., Mulner-Lorillon O., Belle R. In vivo progesterone regulation of protein phosphatase activity in Xenopus oocytes. Dev Biol. 1990 Jun;139(2):427–431. doi: 10.1016/0012-1606(90)90311-6. [DOI] [PubMed] [Google Scholar]
  7. Cyert M. S., Kirschner M. W. Regulation of MPF activity in vitro. Cell. 1988 Apr 22;53(2):185–195. doi: 10.1016/0092-8674(88)90380-7. [DOI] [PubMed] [Google Scholar]
  8. Dabauvalle M. C., Doree M., Bravo R., Karsenti E. Role of nuclear material in the early cell cycle of Xenopus embryos. Cell. 1988 Feb 26;52(4):525–533. doi: 10.1016/0092-8674(88)90465-5. [DOI] [PubMed] [Google Scholar]
  9. Dorée M. Control of M-phase by maturation-promoting factor. Curr Opin Cell Biol. 1990 Apr;2(2):269–273. doi: 10.1016/0955-0674(90)90018-a. [DOI] [PubMed] [Google Scholar]
  10. Dorée M., Labbé J. C., Picard A. M phase-promoting factor: its identification as the M phase-specific H1 histone kinase and its activation by dephosphorylation. J Cell Sci Suppl. 1989;12:39–51. doi: 10.1242/jcs.1989.supplement_12.5. [DOI] [PubMed] [Google Scholar]
  11. Dorée M. Protein synthesis is not involved in initiation or amplification of the maturation-promoting factor (MPF) in starfish oocytes. Exp Cell Res. 1982 May;139(1):127–133. doi: 10.1016/0014-4827(82)90326-3. [DOI] [PubMed] [Google Scholar]
  12. Dunphy W. G., Newport J. W. Fission yeast p13 blocks mitotic activation and tyrosine dephosphorylation of the Xenopus cdc2 protein kinase. Cell. 1989 Jul 14;58(1):181–191. doi: 10.1016/0092-8674(89)90414-5. [DOI] [PubMed] [Google Scholar]
  13. Featherstone C., Russell P. Fission yeast p107wee1 mitotic inhibitor is a tyrosine/serine kinase. Nature. 1991 Feb 28;349(6312):808–811. doi: 10.1038/349808a0. [DOI] [PubMed] [Google Scholar]
  14. Gautier J., Maller J. L. Cyclin B in Xenopus oocytes: implications for the mechanism of pre-MPF activation. EMBO J. 1991 Jan;10(1):177–182. doi: 10.1002/j.1460-2075.1991.tb07934.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gautier J., Matsukawa T., Nurse P., Maller J. Dephosphorylation and activation of Xenopus p34cdc2 protein kinase during the cell cycle. Nature. 1989 Jun 22;339(6226):626–629. doi: 10.1038/339626a0. [DOI] [PubMed] [Google Scholar]
  16. Gerhart J., Wu M., Kirschner M. Cell cycle dynamics of an M-phase-specific cytoplasmic factor in Xenopus laevis oocytes and eggs. J Cell Biol. 1984 Apr;98(4):1247–1255. doi: 10.1083/jcb.98.4.1247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Goris J., Hermann J., Hendrix P., Ozon R., Merlevede W. Okadaic acid, a specific protein phosphatase inhibitor, induces maturation and MPF formation in Xenopus laevis oocytes. FEBS Lett. 1989 Mar 13;245(1-2):91–94. doi: 10.1016/0014-5793(89)80198-x. [DOI] [PubMed] [Google Scholar]
  18. Gould K. L., Nurse P. Tyrosine phosphorylation of the fission yeast cdc2+ protein kinase regulates entry into mitosis. Nature. 1989 Nov 2;342(6245):39–45. doi: 10.1038/342039a0. [DOI] [PubMed] [Google Scholar]
  19. Hermann J., Bellé R., Tso J., Ozon R. Stabilization of the maturation promoting factor (MPF) from Xenopus laevis oocytes. Protection against calcium ions. Cell Differ. 1983 Oct;13(2):143–148. doi: 10.1016/0045-6039(83)90106-9. [DOI] [PubMed] [Google Scholar]
  20. Hiramoto Y. A method of microinjection. Exp Cell Res. 1974 Aug;87(2):403–406. doi: 10.1016/0014-4827(74)90503-5. [DOI] [PubMed] [Google Scholar]
  21. Johnson R. T., Rao P. N. Mammalian cell fusion: induction of premature chromosome condensation in interphase nuclei. Nature. 1970 May 23;226(5247):717–722. doi: 10.1038/226717a0. [DOI] [PubMed] [Google Scholar]
  22. Kinoshita N., Ohkura H., Yanagida M. Distinct, essential roles of type 1 and 2A protein phosphatases in the control of the fission yeast cell division cycle. Cell. 1990 Oct 19;63(2):405–415. doi: 10.1016/0092-8674(90)90173-c. [DOI] [PubMed] [Google Scholar]
  23. Kishimoto T., Hirai S., Kanatani H. Role of germinal vesicle material in producing maturation-promoting factor in starfish oocyte. Dev Biol. 1981 Jan 15;81(1):177–181. doi: 10.1016/0012-1606(81)90360-2. [DOI] [PubMed] [Google Scholar]
  24. Kishimoto T., Kuriyama R., Kondo H., Kanatani H. Generality of the action of various maturation-promoting factors. Exp Cell Res. 1982 Jan;137(1):121–126. doi: 10.1016/0014-4827(82)90014-3. [DOI] [PubMed] [Google Scholar]
  25. Kuang J., Penkala J. E., Wright D. A., Saunders G. F., Rao P. N. A novel M phase-specific H1 kinase recognized by the mitosis-specific monoclonal antibody MPM-2. Dev Biol. 1991 Mar;144(1):54–64. doi: 10.1016/0012-1606(91)90478-l. [DOI] [PubMed] [Google Scholar]
  26. Kumagai A., Dunphy W. G. The cdc25 protein controls tyrosine dephosphorylation of the cdc2 protein in a cell-free system. Cell. 1991 Mar 8;64(5):903–914. doi: 10.1016/0092-8674(91)90315-p. [DOI] [PubMed] [Google Scholar]
  27. Labbe J. C., Lee M. G., Nurse P., Picard A., Doree M. Activation at M-phase of a protein kinase encoded by a starfish homologue of the cell cycle control gene cdc2+. Nature. 1988 Sep 15;335(6187):251–254. doi: 10.1038/335251a0. [DOI] [PubMed] [Google Scholar]
  28. Labbe J. C., Picard A., Peaucellier G., Cavadore J. C., Nurse P., Doree M. Purification of MPF from starfish: identification as the H1 histone kinase p34cdc2 and a possible mechanism for its periodic activation. Cell. 1989 Apr 21;57(2):253–263. doi: 10.1016/0092-8674(89)90963-x. [DOI] [PubMed] [Google Scholar]
  29. Labbé J. C., Capony J. P., Caput D., Cavadore J. C., Derancourt J., Kaghad M., Lelias J. M., Picard A., Dorée M. MPF from starfish oocytes at first meiotic metaphase is a heterodimer containing one molecule of cdc2 and one molecule of cyclin B. EMBO J. 1989 Oct;8(10):3053–3058. doi: 10.1002/j.1460-2075.1989.tb08456.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Labbé J. C., Picard A., Karsenti E., Dorée M. An M-phase-specific protein kinase of Xenopus oocytes: partial purification and possible mechanism of its periodic activation. Dev Biol. 1988 May;127(1):157–169. doi: 10.1016/0012-1606(88)90197-2. [DOI] [PubMed] [Google Scholar]
  31. Langan T. A., Gautier J., Lohka M., Hollingsworth R., Moreno S., Nurse P., Maller J., Sclafani R. A. Mammalian growth-associated H1 histone kinase: a homolog of cdc2+/CDC28 protein kinases controlling mitotic entry in yeast and frog cells. Mol Cell Biol. 1989 Sep;9(9):3860–3868. doi: 10.1128/mcb.9.9.3860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Lee T. H., Solomon M. J., Mumby M. C., Kirschner M. W. INH, a negative regulator of MPF, is a form of protein phosphatase 2A. Cell. 1991 Jan 25;64(2):415–423. doi: 10.1016/0092-8674(91)90649-j. [DOI] [PubMed] [Google Scholar]
  33. 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. doi: 10.1016/0092-8674(91)90266-2. [DOI] [PubMed] [Google Scholar]
  34. Masui Y., Markert C. L. Cytoplasmic control of nuclear behavior during meiotic maturation of frog oocytes. J Exp Zool. 1971 Jun;177(2):129–145. doi: 10.1002/jez.1401770202. [DOI] [PubMed] [Google Scholar]
  35. Meikrantz W., Suprynowicz F. A., Halleck M. S., Schlegel R. A. Identification of mitosis-specific p65 dimer as a component of human M phase-promoting factor. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9600–9604. doi: 10.1073/pnas.87.24.9600. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Morla A. O., Draetta G., Beach D., Wang J. Y. Reversible tyrosine phosphorylation of cdc2: dephosphorylation accompanies activation during entry into mitosis. Cell. 1989 Jul 14;58(1):193–203. doi: 10.1016/0092-8674(89)90415-7. [DOI] [PubMed] [Google Scholar]
  37. Nurse P. Universal control mechanism regulating onset of M-phase. Nature. 1990 Apr 5;344(6266):503–508. doi: 10.1038/344503a0. [DOI] [PubMed] [Google Scholar]
  38. Peter M., Nakagawa J., Dorée M., Labbé J. C., Nigg E. A. Identification of major nucleolar proteins as candidate mitotic substrates of cdc2 kinase. Cell. 1990 Mar 9;60(5):791–801. doi: 10.1016/0092-8674(90)90093-t. [DOI] [PubMed] [Google Scholar]
  39. Peter M., Nakagawa J., Dorée M., Labbé J. C., Nigg E. A. In vitro disassembly of the nuclear lamina and M phase-specific phosphorylation of lamins by cdc2 kinase. Cell. 1990 May 18;61(4):591–602. doi: 10.1016/0092-8674(90)90471-p. [DOI] [PubMed] [Google Scholar]
  40. Picard A., Capony J. P., Brautigan D. L., Dorée M. Involvement of protein phosphatases 1 and 2A in the control of M phase-promoting factor activity in starfish. J Cell Biol. 1989 Dec;109(6 Pt 2):3347–3354. doi: 10.1083/jcb.109.6.3347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Picard A., Doree M. The role of the germinal vesicle in producing maturation-promoting factor (MPF) as revealed by the removal and transplantation of nuclear material in starfish oocytes. Dev Biol. 1984 Aug;104(2):357–365. doi: 10.1016/0012-1606(84)90091-5. [DOI] [PubMed] [Google Scholar]
  42. Picard A., Harricane M. C., Labbe J. C., Doree M. Germinal vesicle components are not required for the cell-cycle oscillator of the early starfish embryo. Dev Biol. 1988 Jul;128(1):121–128. doi: 10.1016/0012-1606(88)90273-4. [DOI] [PubMed] [Google Scholar]
  43. Pondaven P., Meijer L., Beach D. Activation of M-phase-specific histone H1 kinase by modification of the phosphorylation of its p34cdc2 and cyclin components. Genes Dev. 1990 Jan;4(1):9–17. doi: 10.1101/gad.4.1.9. [DOI] [PubMed] [Google Scholar]
  44. Rime H., Ozon R. Protein phosphatases are involved in the in vivo activation of histone H1 kinase in mouse oocyte. Dev Biol. 1990 Sep;141(1):115–122. doi: 10.1016/0012-1606(90)90106-s. [DOI] [PubMed] [Google Scholar]
  45. Skoblina M. N., Pivnitsky K. K., Kondratieva O. T. The role of germinal vesicle in maturation of Pleurodeles waltlii oocytes induced by steroids. Cell Differ. 1984 Jun;14(2):153–157. doi: 10.1016/0045-6039(84)90040-x. [DOI] [PubMed] [Google Scholar]
  46. Smith L. D., Ecker R. E. Role of the oocyte nucleus in physiological maturation in Rana pipiens. Dev Biol. 1969 Mar;19(3):281–309. doi: 10.1016/0012-1606(69)90065-7. [DOI] [PubMed] [Google Scholar]
  47. Strausfeld U., Labbé J. C., Fesquet D., Cavadore J. C., 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. doi: 10.1038/351242a0. [DOI] [PubMed] [Google Scholar]
  48. Tuomikoski T., Felix M. A., Dorée M., Gruenberg J. Inhibition of endocytic vesicle fusion in vitro by the cell-cycle control protein kinase cdc2. Nature. 1989 Dec 21;342(6252):942–945. doi: 10.1038/342942a0. [DOI] [PubMed] [Google Scholar]
  49. Wasserman W. J., Masui Y. Effects of cyclohexamide on a cytoplasmic factor initiating meiotic naturation in Xenopus oocytes. Exp Cell Res. 1975 Mar 15;91(2):381–388. doi: 10.1016/0014-4827(75)90118-4. [DOI] [PubMed] [Google Scholar]
  50. Yamashiro S., Yamakita Y., Hosoya H., Matsumura F. Phosphorylation of non-muscle caldesmon by p34cdc2 kinase during mitosis. Nature. 1991 Jan 10;349(6305):169–172. doi: 10.1038/349169a0. [DOI] [PubMed] [Google Scholar]
  51. Yamashita S., Maller J. L. Identification of an activator required for elevation of maturation-promoting factor (MPF) activity by gamma-S-ATP. J Cell Biol. 1990 May;110(5):1583–1588. doi: 10.1083/jcb.110.5.1583. [DOI] [PMC free article] [PubMed] [Google Scholar]

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