Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1997 Sep 15;16(18):5537–5549. doi: 10.1093/emboj/16.18.5537

Distinct roles of PP1 and PP2A-like phosphatases in control of microtubule dynamics during mitosis.

R Tournebize 1, S S Andersen 1, F Verde 1, M Dorée 1, E Karsenti 1, A A Hyman 1
PMCID: PMC1170186  PMID: 9312013

Abstract

Assembly of a mitotic spindle requires the accurate regulation of microtubule dynamics which is accomplished, at least in part, by phosphorylation-dephosphorylation reactions. Here we have investigated the role of serine-threonine phosphatases in the control of microtubule dynamics using specific inhibitors in Xenopus egg extracts. Type 2A phosphatases are required to maintain the short steady-state length of microtubules in mitosis by regulating the level of microtubule catastrophes, in part by controlling the the microtubule-destabilizing activity and phosphorylation of Op18/stathmin. Type 1 phosphatases are only required for control of microtubule dynamics during the transitions into and out of mitosis. Thus, although both type 2A and type 1 phosphatases are involved in the regulation of microtubule dynamics, they have distinct, non-overlapping roles.

Full Text

The Full Text of this article is available as a PDF (591.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Armstrong C. G., Mann D. J., Berndt N., Cohen P. T. Drosophila PPY, a novel male specific protein serine/threonine phosphatase localised in somatic cells of the testis. J Cell Sci. 1995 Nov;108(Pt 11):3367–3375. doi: 10.1242/jcs.108.11.3367. [DOI] [PubMed] [Google Scholar]
  2. Ault J. G., DeMarco A. J., Salmon E. D., Rieder C. L. Studies on the ejection properties of asters: astral microtubule turnover influences the oscillatory behavior and positioning of mono-oriented chromosomes. J Cell Sci. 1991 Aug;99(Pt 4):701–710. doi: 10.1242/jcs.99.4.701. [DOI] [PubMed] [Google Scholar]
  3. Axton J. M., Dombrádi V., Cohen P. T., Glover D. M. One of the protein phosphatase 1 isoenzymes in Drosophila is essential for mitosis. Cell. 1990 Oct 5;63(1):33–46. doi: 10.1016/0092-8674(90)90286-n. [DOI] [PubMed] [Google Scholar]
  4. Belmont L. D., Hyman A. A., Sawin K. E., Mitchison T. J. Real-time visualization of cell cycle-dependent changes in microtubule dynamics in cytoplasmic extracts. Cell. 1990 Aug 10;62(3):579–589. doi: 10.1016/0092-8674(90)90022-7. [DOI] [PubMed] [Google Scholar]
  5. Belmont L. D., Mitchison T. J. Identification of a protein that interacts with tubulin dimers and increases the catastrophe rate of microtubules. Cell. 1996 Feb 23;84(4):623–631. doi: 10.1016/s0092-8674(00)81037-5. [DOI] [PubMed] [Google Scholar]
  6. Black S., Andrews P. D., Sneddon A. A., Stark M. J. A regulated MET3-GLC7 gene fusion provides evidence of a mitotic role for Saccharomyces cerevisiae protein phosphatase 1. Yeast. 1995 Jun 30;11(8):747–759. doi: 10.1002/yea.320110806. [DOI] [PubMed] [Google Scholar]
  7. Booher R., Beach D. Involvement of a type 1 protein phosphatase encoded by bws1+ in fission yeast mitotic control. Cell. 1989 Jun 16;57(6):1009–1016. doi: 10.1016/0092-8674(89)90339-5. [DOI] [PubMed] [Google Scholar]
  8. Brattsand G., Marklund U., Nylander K., Roos G., Gullberg M. Cell-cycle-regulated phosphorylation of oncoprotein 18 on Ser16, Ser25 and Ser38. Eur J Biochem. 1994 Mar 1;220(2):359–368. doi: 10.1111/j.1432-1033.1994.tb18632.x. [DOI] [PubMed] [Google Scholar]
  9. Brewis N. D., Street A. J., Prescott A. R., Cohen P. T. PPX, a novel protein serine/threonine phosphatase localized to centrosomes. EMBO J. 1993 Mar;12(3):987–996. doi: 10.1002/j.1460-2075.1993.tb05739.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chen M. X., McPartlin A. E., Brown L., Chen Y. H., Barker H. M., Cohen P. T. A novel human protein serine/threonine phosphatase, which possesses four tetratricopeptide repeat motifs and localizes to the nucleus. EMBO J. 1994 Sep 15;13(18):4278–4290. doi: 10.1002/j.1460-2075.1994.tb06748.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Clarke P. R., Hoffmann I., Draetta G., Karsenti E. Dephosphorylation of cdc25-C by a type-2A protein phosphatase: specific regulation during the cell cycle in Xenopus egg extracts. Mol Biol Cell. 1993 Apr;4(4):397–411. doi: 10.1091/mbc.4.4.397. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Cohen P. The structure and regulation of protein phosphatases. Annu Rev Biochem. 1989;58:453–508. doi: 10.1146/annurev.bi.58.070189.002321. [DOI] [PubMed] [Google Scholar]
  13. Dogterom M., Félix M. A., Guet C. C., Leibler S. Influence of M-phase chromatin on the anisotropy of microtubule asters. J Cell Biol. 1996 Apr;133(1):125–140. doi: 10.1083/jcb.133.1.125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dohadwala M., da Cruz e Silva E. F., Hall F. L., Williams R. T., Carbonaro-Hall D. A., Nairn A. C., Greengard P., Berndt N. Phosphorylation and inactivation of protein phosphatase 1 by cyclin-dependent kinases. Proc Natl Acad Sci U S A. 1994 Jul 5;91(14):6408–6412. doi: 10.1073/pnas.91.14.6408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Evans D. R., Stark M. J. Mutations in the Saccharomyces cerevisiae type 2A protein phosphatase catalytic subunit reveal roles in cell wall integrity, actin cytoskeleton organization and mitosis. Genetics. 1997 Feb;145(2):227–241. doi: 10.1093/genetics/145.2.227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Felix M. A., Pines J., Hunt T., Karsenti E. A post-ribosomal supernatant from activated Xenopus eggs that displays post-translationally regulated oscillation of its cdc2+ mitotic kinase activity. EMBO J. 1989 Oct;8(10):3059–3069. doi: 10.1002/j.1460-2075.1989.tb08457.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fernandez A., Brautigan D. L., Lamb N. J. Protein phosphatase type 1 in mammalian cell mitosis: chromosomal localization and involvement in mitotic exit. J Cell Biol. 1992 Mar;116(6):1421–1430. doi: 10.1083/jcb.116.6.1421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Ferrigno P., Langan T. A., Cohen P. Protein phosphatase 2A1 is the major enzyme in vertebrate cell extracts that dephosphorylates several physiological substrates for cyclin-dependent protein kinases. Mol Biol Cell. 1993 Jul;4(7):669–677. doi: 10.1091/mbc.4.7.669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Glotzer M., Murray A. W., Kirschner M. W. Cyclin is degraded by the ubiquitin pathway. Nature. 1991 Jan 10;349(6305):132–138. doi: 10.1038/349132a0. [DOI] [PubMed] [Google Scholar]
  20. Gomes R., Karess R. E., Ohkura H., Glover D. M., Sunkel C. E. Abnormal anaphase resolution (aar): a locus required for progression through mitosis in Drosophila. J Cell Sci. 1993 Feb;104(Pt 2):583–593. doi: 10.1242/jcs.104.2.583. [DOI] [PubMed] [Google Scholar]
  21. Heald R., Tournebize R., Blank T., Sandaltzopoulos R., Becker P., Hyman A., Karsenti E. Self-organization of microtubules into bipolar spindles around artificial chromosomes in Xenopus egg extracts. Nature. 1996 Aug 1;382(6590):420–425. doi: 10.1038/382420a0. [DOI] [PubMed] [Google Scholar]
  22. Hisamoto N., Sugimoto K., Matsumoto K. The Glc7 type 1 protein phosphatase of Saccharomyces cerevisiae is required for cell cycle progression in G2/M. Mol Cell Biol. 1994 May;14(5):3158–3165. doi: 10.1128/mcb.14.5.3158. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Holloway S. L., Glotzer M., King R. W., Murray A. W. Anaphase is initiated by proteolysis rather than by the inactivation of maturation-promoting factor. Cell. 1993 Jul 2;73(7):1393–1402. doi: 10.1016/0092-8674(93)90364-v. [DOI] [PubMed] [Google Scholar]
  24. Honkanen R. E., Zwiller J., Daily S. L., Khatra B. S., Dukelow M., Boynton A. L. Identification, purification, and characterization of a novel serine/threonine protein phosphatase from bovine brain. J Biol Chem. 1991 Apr 5;266(10):6614–6619. [PubMed] [Google Scholar]
  25. Horwitz S. B., Shen H. J., He L., Dittmar P., Neef R., Chen J., Schubart U. K. The microtubule-destabilizing activity of metablastin (p19) is controlled by phosphorylation. J Biol Chem. 1997 Mar 28;272(13):8129–8132. doi: 10.1074/jbc.272.13.8129. [DOI] [PubMed] [Google Scholar]
  26. Hyman A., Drechsel D., Kellogg D., Salser S., Sawin K., Steffen P., Wordeman L., Mitchison T. Preparation of modified tubulins. Methods Enzymol. 1991;196:478–485. doi: 10.1016/0076-6879(91)96041-o. [DOI] [PubMed] [Google Scholar]
  27. Kinoshita N., Yamano H., Niwa H., Yoshida T., Yanagida M. Negative regulation of mitosis by the fission yeast protein phosphatase ppa2. Genes Dev. 1993 Jun;7(6):1059–1071. doi: 10.1101/gad.7.6.1059. [DOI] [PubMed] [Google Scholar]
  28. Larsson N., Melander H., Marklund U., Osterman O., Gullberg M. G2/M transition requires multisite phosphorylation of oncoprotein 18 by two distinct protein kinase systems. J Biol Chem. 1995 Jun 9;270(23):14175–14183. doi: 10.1074/jbc.270.23.14175. [DOI] [PubMed] [Google Scholar]
  29. Lee T. H., Turck C., Kirschner M. W. Inhibition of cdc2 activation by INH/PP2A. Mol Biol Cell. 1994 Mar;5(3):323–338. doi: 10.1091/mbc.5.3.323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lorca T., Cruzalegui F. H., Fesquet D., Cavadore J. C., Méry J., Means A., Dorée M. Calmodulin-dependent protein kinase II mediates inactivation of MPF and CSF upon fertilization of Xenopus eggs. Nature. 1993 Nov 18;366(6452):270–273. doi: 10.1038/366270a0. [DOI] [PubMed] [Google Scholar]
  31. MAQSOOD M., BIOL M. I. Radioisotopes in medicine. Pak J Health. 1957 Jan;6(4):193–203. [PubMed] [Google Scholar]
  32. Marklund U., Larsson N., Gradin H. M., Brattsand G., Gullberg M. Oncoprotein 18 is a phosphorylation-responsive regulator of microtubule dynamics. EMBO J. 1996 Oct 1;15(19):5290–5298. [PMC free article] [PubMed] [Google Scholar]
  33. Minshull J., Sun H., Tonks N. K., Murray A. W. A MAP kinase-dependent spindle assembly checkpoint in Xenopus egg extracts. Cell. 1994 Nov 4;79(3):475–486. doi: 10.1016/0092-8674(94)90256-9. [DOI] [PubMed] [Google Scholar]
  34. Murray A. W., Solomon M. J., Kirschner M. W. The role of cyclin synthesis and degradation in the control of maturation promoting factor activity. Nature. 1989 May 25;339(6222):280–286. doi: 10.1038/339280a0. [DOI] [PubMed] [Google Scholar]
  35. Nebreda A. R., Hunt T. The c-mos proto-oncogene protein kinase turns on and maintains the activity of MAP kinase, but not MPF, in cell-free extracts of Xenopus oocytes and eggs. EMBO J. 1993 May;12(5):1979–1986. doi: 10.1002/j.1460-2075.1993.tb05847.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Ohkura H., Kinoshita N., Miyatani S., Toda T., Yanagida M. The fission yeast dis2+ gene required for chromosome disjoining encodes one of two putative type 1 protein phosphatases. Cell. 1989 Jun 16;57(6):997–1007. doi: 10.1016/0092-8674(89)90338-3. [DOI] [PubMed] [Google Scholar]
  37. Parsons S. F., Salmon E. D. Microtubule assembly in clarified Xenopus egg extracts. Cell Motil Cytoskeleton. 1997;36(1):1–11. doi: 10.1002/(SICI)1097-0169(1997)36:1<1::AID-CM1>3.0.CO;2-E. [DOI] [PubMed] [Google Scholar]
  38. 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]
  39. Picard A., Labbé J. C., Barakat H., Cavadore J. C., Dorée M. Okadaic acid mimics a nuclear component required for cyclin B-cdc2 kinase microinjection to drive starfish oocytes into M phase. J Cell Biol. 1991 Oct;115(2):337–344. doi: 10.1083/jcb.115.2.337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sawin K. E., Mitchison T. J. Mitotic spindle assembly by two different pathways in vitro. J Cell Biol. 1991 Mar;112(5):925–940. doi: 10.1083/jcb.112.5.925. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Shamu C. E., Murray A. W. Sister chromatid separation in frog egg extracts requires DNA topoisomerase II activity during anaphase. J Cell Biol. 1992 Jun;117(5):921–934. doi: 10.1083/jcb.117.5.921. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Shenolikar S. Protein serine/threonine phosphatases--new avenues for cell regulation. Annu Rev Cell Biol. 1994;10:55–86. doi: 10.1146/annurev.cb.10.110194.000415. [DOI] [PubMed] [Google Scholar]
  43. Shibuya E. K., Polverino A. J., Chang E., Wigler M., Ruderman J. V. Oncogenic ras triggers the activation of 42-kDa mitogen-activated protein kinase in extracts of quiescent Xenopus oocytes. Proc Natl Acad Sci U S A. 1992 Oct 15;89(20):9831–9835. doi: 10.1073/pnas.89.20.9831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Snaith H. A., Armstrong C. G., Guo Y., Kaiser K., Cohen P. T. Deficiency of protein phosphatase 2A uncouples the nuclear and centrosome cycles and prevents attachment of microtubules to the kinetochore in Drosophila microtubule star (mts) embryos. J Cell Sci. 1996 Dec;109(Pt 13):3001–3012. doi: 10.1242/jcs.109.13.3001. [DOI] [PubMed] [Google Scholar]
  45. Sobel A. Stathmin: a relay phosphoprotein for multiple signal transduction? Trends Biochem Sci. 1991 Aug;16(8):301–305. doi: 10.1016/0968-0004(91)90123-d. [DOI] [PubMed] [Google Scholar]
  46. Sontag E., Nunbhakdi-Craig V., Bloom G. S., Mumby M. C. A novel pool of protein phosphatase 2A is associated with microtubules and is regulated during the cell cycle. J Cell Biol. 1995 Mar;128(6):1131–1144. doi: 10.1083/jcb.128.6.1131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Surana U., Amon A., Dowzer C., McGrew J., Byers B., Nasmyth K. Destruction of the CDC28/CLB mitotic kinase is not required for the metaphase to anaphase transition in budding yeast. EMBO J. 1993 May;12(5):1969–1978. doi: 10.1002/j.1460-2075.1993.tb05846.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Van Dolah F. M., Ramsdell J. S. Okadaic acid inhibits a protein phosphatase activity involved in formation of the mitotic spindle of GH4 rat pituitary cells. J Cell Physiol. 1992 Jul;152(1):190–198. doi: 10.1002/jcp.1041520124. [DOI] [PubMed] [Google Scholar]
  49. Verde F., Dogterom M., Stelzer E., Karsenti E., Leibler S. Control of microtubule dynamics and length by cyclin A- and cyclin B-dependent kinases in Xenopus egg extracts. J Cell Biol. 1992 Sep;118(5):1097–1108. doi: 10.1083/jcb.118.5.1097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Verde F., Labbé J. C., Dorée M., Karsenti E. Regulation of microtubule dynamics by cdc2 protein kinase in cell-free extracts of Xenopus eggs. Nature. 1990 Jan 18;343(6255):233–238. doi: 10.1038/343233a0. [DOI] [PubMed] [Google Scholar]
  51. Walczak C. E., Mitchison T. J., Desai A. XKCM1: a Xenopus kinesin-related protein that regulates microtubule dynamics during mitotic spindle assembly. Cell. 1996 Jan 12;84(1):37–47. doi: 10.1016/s0092-8674(00)80991-5. [DOI] [PubMed] [Google Scholar]
  52. Walker R. A., O'Brien E. T., Pryer N. K., Soboeiro M. F., Voter W. A., Erickson H. P., Salmon E. D. Dynamic instability of individual microtubules analyzed by video light microscopy: rate constants and transition frequencies. J Cell Biol. 1988 Oct;107(4):1437–1448. doi: 10.1083/jcb.107.4.1437. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Wells W. A. The spindle-assembly checkpoint: aiming for a perfect mitosis, every time. Trends Cell Biol. 1996 Jun;6(6):228–234. doi: 10.1016/0962-8924(96)10018-0. [DOI] [PubMed] [Google Scholar]
  54. Yamano H., Ishii K., Yanagida M. Phosphorylation of dis2 protein phosphatase at the C-terminal cdc2 consensus and its potential role in cell cycle regulation. EMBO J. 1994 Nov 15;13(22):5310–5318. doi: 10.1002/j.1460-2075.1994.tb06865.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Zhai Y., Kronebusch P. J., Simon P. M., Borisy G. G. Microtubule dynamics at the G2/M transition: abrupt breakdown of cytoplasmic microtubules at nuclear envelope breakdown and implications for spindle morphogenesis. J Cell Biol. 1996 Oct;135(1):201–214. doi: 10.1083/jcb.135.1.201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Zhang D., Nicklas R. B. The impact of chromosomes and centrosomes on spindle assembly as observed in living cells. J Cell Biol. 1995 Jun;129(5):1287–1300. doi: 10.1083/jcb.129.5.1287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. de Pennart H., Verlhac M. H., Cibert C., Santa Maria A., Maro B. Okadaic acid induces spindle lengthening and disrupts the interaction of microtubules with the kinetochores in metaphase II-arrested mouse oocytes. Dev Biol. 1993 May;157(1):170–181. doi: 10.1006/dbio.1993.1121. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES