Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1997 Feb;17(2):571–583. doi: 10.1128/mcb.17.2.571

The human Myt1 kinase preferentially phosphorylates Cdc2 on threonine 14 and localizes to the endoplasmic reticulum and Golgi complex.

F Liu 1, J J Stanton 1, Z Wu 1, H Piwnica-Worms 1
PMCID: PMC231782  PMID: 9001210

Abstract

Entry into mitosis requires the activity of the Cdc2 kinase. Cdc2 associates with the B-type cyclins, and the Cdc2-cyclin B heterodimer is in turn regulated by phosphorylation. Phosphorylation of threonine 161 is required for the Cdc2-cyclin B complex to be catalytically active, whereas phosphorylation of threonine 14 and tyrosine 15 is inhibitory. Human kinases that catalyze the phosphorylation of threonine 161 and tyrosine 15 have been identified. Here we report the isolation of a novel human cDNA encoding a dual-specificity protein kinase (designated Myt1Hu) that preferentially phosphorylates Cdc2 on threonine 14 in a cyclin-dependent manner. Myt1Hu is 46% identical to Myt1Xe, a kinase recently characterized from Xenopus laevis. Myt1Hu localizes to the endoplasmic reticulum and Golgi complex in HeLa cells. A stretch of hydrophobic and uncharged amino acids located outside the catalytic domain of Myt1Hu is the likely membrane-targeting domain, as its deletion results in the localization of Myt1Hu primarily to the nucleus.

Full Text

The Full Text of this article is available as a PDF (616.9 KB).

Selected References

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

  1. Atherton-Fessler S., Hannig G., Piwnica-Worms H. Reversible tyrosine phosphorylation and cell cycle control. Semin Cell Biol. 1993 Dec;4(6):433–442. doi: 10.1006/scel.1993.1051. [DOI] [PubMed] [Google Scholar]
  2. Atherton-Fessler S., Liu F., Gabrielli B., Lee M. S., Peng C. Y., Piwnica-Worms H. Cell cycle regulation of the p34cdc2 inhibitory kinases. Mol Biol Cell. 1994 Sep;5(9):989–1001. doi: 10.1091/mbc.5.9.989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Atherton-Fessler S., Parker L. L., Geahlen R. L., Piwnica-Worms H. Mechanisms of p34cdc2 regulation. Mol Cell Biol. 1993 Mar;13(3):1675–1685. doi: 10.1128/mcb.13.3.1675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bailly E., Pines J., Hunter T., Bornens M. Cytoplasmic accumulation of cyclin B1 in human cells: association with a detergent-resistant compartment and with the centrosome. J Cell Sci. 1992 Mar;101(Pt 3):529–545. doi: 10.1242/jcs.101.3.529. [DOI] [PubMed] [Google Scholar]
  5. Balch W. E., Dunphy W. G., Braell W. A., Rothman J. E. Reconstitution of the transport of protein between successive compartments of the Golgi measured by the coupled incorporation of N-acetylglucosamine. Cell. 1984 Dec;39(2 Pt 1):405–416. doi: 10.1016/0092-8674(84)90019-9. [DOI] [PubMed] [Google Scholar]
  6. Baldin V., Ducommun B. Subcellular localisation of human wee1 kinase is regulated during the cell cycle. J Cell Sci. 1995 Jun;108(Pt 6):2425–2432. doi: 10.1242/jcs.108.6.2425. [DOI] [PubMed] [Google Scholar]
  7. Booher R. N., Deshaies R. J., Kirschner M. W. Properties of Saccharomyces cerevisiae wee1 and its differential regulation of p34CDC28 in response to G1 and G2 cyclins. EMBO J. 1993 Sep;12(9):3417–3426. doi: 10.1002/j.1460-2075.1993.tb06016.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Campbell S. D., Sprenger F., Edgar B. A., O'Farrell P. H. Drosophila Wee1 kinase rescues fission yeast from mitotic catastrophe and phosphorylates Drosophila Cdc2 in vitro. Mol Biol Cell. 1995 Oct;6(10):1333–1347. doi: 10.1091/mbc.6.10.1333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Den Haese G. J., Walworth N., Carr A. M., Gould K. L. The Wee1 protein kinase regulates T14 phosphorylation of fission yeast Cdc2. Mol Biol Cell. 1995 Apr;6(4):371–385. doi: 10.1091/mbc.6.4.371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Draetta G., Beach D. Activation of cdc2 protein kinase during mitosis in human cells: cell cycle-dependent phosphorylation and subunit rearrangement. Cell. 1988 Jul 1;54(1):17–26. doi: 10.1016/0092-8674(88)90175-4. [DOI] [PubMed] [Google Scholar]
  11. Draetta G., Piwnica-Worms H., Morrison D., Druker B., Roberts T., Beach D. Human cdc2 protein kinase is a major cell-cycle regulated tyrosine kinase substrate. Nature. 1988 Dec 22;336(6201):738–744. doi: 10.1038/336738a0. [DOI] [PubMed] [Google Scholar]
  12. Dunphy W. G., Kumagai A. The cdc25 protein contains an intrinsic phosphatase activity. Cell. 1991 Oct 4;67(1):189–196. doi: 10.1016/0092-8674(91)90582-j. [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. Frangioni J. V., Beahm P. H., Shifrin V., Jost C. A., Neel B. G. The nontransmembrane tyrosine phosphatase PTP-1B localizes to the endoplasmic reticulum via its 35 amino acid C-terminal sequence. Cell. 1992 Feb 7;68(3):545–560. doi: 10.1016/0092-8674(92)90190-n. [DOI] [PubMed] [Google Scholar]
  15. Frangioni J. V., Oda A., Smith M., Salzman E. W., Neel B. G. Calpain-catalyzed cleavage and subcellular relocation of protein phosphotyrosine phosphatase 1B (PTP-1B) in human platelets. EMBO J. 1993 Dec;12(12):4843–4856. doi: 10.1002/j.1460-2075.1993.tb06174.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gallant P., Nigg E. A. Identification of a novel vertebrate cyclin: cyclin B3 shares properties with both A- and B-type cyclins. EMBO J. 1994 Feb 1;13(3):595–605. doi: 10.1002/j.1460-2075.1994.tb06297.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gautier J., Solomon M. J., Booher R. N., Bazan J. F., Kirschner M. W. cdc25 is a specific tyrosine phosphatase that directly activates p34cdc2. Cell. 1991 Oct 4;67(1):197–211. doi: 10.1016/0092-8674(91)90583-k. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Gu Y., Rosenblatt J., Morgan D. O. Cell cycle regulation of CDK2 activity by phosphorylation of Thr160 and Tyr15. EMBO J. 1992 Nov;11(11):3995–4005. doi: 10.1002/j.1460-2075.1992.tb05493.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Heald R., McLoughlin M., McKeon F. Human wee1 maintains mitotic timing by protecting the nucleus from cytoplasmically activated Cdc2 kinase. Cell. 1993 Aug 13;74(3):463–474. doi: 10.1016/0092-8674(93)80048-j. [DOI] [PubMed] [Google Scholar]
  21. Honda R., Ohba Y., Yasuda H. The cell cycle regulator, human p50weel, is a tyrosine kinase and not a serine/tyrosine kinase. Biochem Biophys Res Commun. 1992 Aug 14;186(3):1333–1338. doi: 10.1016/s0006-291x(05)81552-9. [DOI] [PubMed] [Google Scholar]
  22. Igarashi M., Nagata A., Jinno S., Suto K., Okayama H. Wee1(+)-like gene in human cells. Nature. 1991 Sep 5;353(6339):80–83. doi: 10.1038/353080a0. [DOI] [PubMed] [Google Scholar]
  23. Jackman M., Firth M., Pines J. Human cyclins B1 and B2 are localized to strikingly different structures: B1 to microtubules, B2 primarily to the Golgi apparatus. EMBO J. 1995 Apr 18;14(8):1646–1654. doi: 10.1002/j.1460-2075.1995.tb07153.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kornbluth S., Sebastian B., Hunter T., Newport J. Membrane localization of the kinase which phosphorylates p34cdc2 on threonine 14. Mol Biol Cell. 1994 Mar;5(3):273–282. doi: 10.1091/mbc.5.3.273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Krek W., Nigg E. A. 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. doi: 10.1002/j.1460-2075.1991.tb07951.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lee M. S., Enoch T., Piwnica-Worms H. mik1+ encodes a tyrosine kinase that phosphorylates p34cdc2 on tyrosine 15. J Biol Chem. 1994 Dec 2;269(48):30530–30537. [PubMed] [Google Scholar]
  27. Lee M. S., Ogg S., Xu M., Parker L. L., Donoghue D. J., Maller J. L., Piwnica-Worms H. cdc25+ encodes a protein phosphatase that dephosphorylates p34cdc2. Mol Biol Cell. 1992 Jan;3(1):73–84. doi: 10.1091/mbc.3.1.73. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Matsuura I., Wang J. H. Demonstration of cyclin-dependent kinase inhibitory serine/threonine kinase in bovine thymus. J Biol Chem. 1996 Mar 8;271(10):5443–5450. doi: 10.1074/jbc.271.10.5443. [DOI] [PubMed] [Google Scholar]
  29. McGowan C. H., Russell P. Cell cycle regulation of human WEE1. EMBO J. 1995 May 15;14(10):2166–2175. doi: 10.1002/j.1460-2075.1995.tb07210.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. McGowan C. H., Russell P. Human Wee1 kinase inhibits cell division by phosphorylating p34cdc2 exclusively on Tyr15. EMBO J. 1993 Jan;12(1):75–85. doi: 10.1002/j.1460-2075.1993.tb05633.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Meijer L., Azzi L., Wang J. Y. Cyclin B targets p34cdc2 for tyrosine phosphorylation. EMBO J. 1991 Jun;10(6):1545–1554. doi: 10.1002/j.1460-2075.1991.tb07674.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Morgan D. O. Principles of CDK regulation. Nature. 1995 Mar 9;374(6518):131–134. doi: 10.1038/374131a0. [DOI] [PubMed] [Google Scholar]
  33. 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]
  34. Mueller P. R., Coleman T. R., Dunphy W. G. Cell cycle regulation of a Xenopus Wee1-like kinase. Mol Biol Cell. 1995 Jan;6(1):119–134. doi: 10.1091/mbc.6.1.119. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mueller P. R., Coleman T. R., Kumagai A., Dunphy W. G. Myt1: a membrane-associated inhibitory kinase that phosphorylates Cdc2 on both threonine-14 and tyrosine-15. Science. 1995 Oct 6;270(5233):86–90. doi: 10.1126/science.270.5233.86. [DOI] [PubMed] [Google Scholar]
  36. 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]
  37. Norbury C., Blow J., Nurse P. Regulatory phosphorylation of the p34cdc2 protein kinase in vertebrates. EMBO J. 1991 Nov;10(11):3321–3329. doi: 10.1002/j.1460-2075.1991.tb04896.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Norbury C., Nurse P. Animal cell cycles and their control. Annu Rev Biochem. 1992;61:441–470. doi: 10.1146/annurev.bi.61.070192.002301. [DOI] [PubMed] [Google Scholar]
  39. 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]
  40. Pardee A. B. G1 events and regulation of cell proliferation. Science. 1989 Nov 3;246(4930):603–608. doi: 10.1126/science.2683075. [DOI] [PubMed] [Google Scholar]
  41. Parker L. L., Atherton-Fessler S., Lee M. S., Ogg S., Falk J. L., Swenson K. I., Piwnica-Worms H. Cyclin promotes the tyrosine phosphorylation of p34cdc2 in a wee1+ dependent manner. EMBO J. 1991 May;10(5):1255–1263. doi: 10.1002/j.1460-2075.1991.tb08067.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Parker L. L., Atherton-Fessler S., Piwnica-Worms H. p107wee1 is a dual-specificity kinase that phosphorylates p34cdc2 on tyrosine 15. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2917–2921. doi: 10.1073/pnas.89.7.2917. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Parker L. L., Piwnica-Worms H. Inactivation of the p34cdc2-cyclin B complex by the human WEE1 tyrosine kinase. Science. 1992 Sep 25;257(5078):1955–1957. doi: 10.1126/science.1384126. [DOI] [PubMed] [Google Scholar]
  44. Parker L. L., Sylvestre P. J., Byrnes M. J., 3rd, Liu F., Piwnica-Worms H. Identification of a 95-kDa WEE1-like tyrosine kinase in HeLa cells. Proc Natl Acad Sci U S A. 1995 Oct 10;92(21):9638–9642. doi: 10.1073/pnas.92.21.9638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Parker L. L., Walter S. A., Young P. G., Piwnica-Worms H. Phosphorylation and inactivation of the mitotic inhibitor Wee1 by the nim1/cdr1 kinase. Nature. 1993 Jun 24;363(6431):736–738. doi: 10.1038/363736a0. [DOI] [PubMed] [Google Scholar]
  46. Pines J. Cyclins and cyclin-dependent kinases: take your partners. Trends Biochem Sci. 1993 Jun;18(6):195–197. doi: 10.1016/0968-0004(93)90185-p. [DOI] [PubMed] [Google Scholar]
  47. Pines J., Hunter T. The differential localization of human cyclins A and B is due to a cytoplasmic retention signal in cyclin B. EMBO J. 1994 Aug 15;13(16):3772–3781. doi: 10.1002/j.1460-2075.1994.tb06688.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. 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. doi: 10.1016/0092-8674(87)90458-2. [DOI] [PubMed] [Google Scholar]
  49. 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. doi: 10.1073/pnas.90.8.3521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Sherr C. J., Roberts J. M. Inhibitors of mammalian G1 cyclin-dependent kinases. Genes Dev. 1995 May 15;9(10):1149–1163. doi: 10.1101/gad.9.10.1149. [DOI] [PubMed] [Google Scholar]
  51. Solomon M. J. Activation of the various cyclin/cdc2 protein kinases. Curr Opin Cell Biol. 1993 Apr;5(2):180–186. doi: 10.1016/0955-0674(93)90100-5. [DOI] [PubMed] [Google Scholar]
  52. Solomon M. J., Lee T., Kirschner M. W. Role of phosphorylation in p34cdc2 activation: identification of an activating kinase. Mol Biol Cell. 1992 Jan;3(1):13–27. doi: 10.1091/mbc.3.1.13. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. 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]
  54. Terada Y., Tatsuka M., Jinno S., Okayama H. Requirement for tyrosine phosphorylation of Cdk4 in G1 arrest induced by ultraviolet irradiation. Nature. 1995 Jul 27;376(6538):358–362. doi: 10.1038/376358a0. [DOI] [PubMed] [Google Scholar]
  55. Terasaki M., Chen L. B., Fujiwara K. Microtubules and the endoplasmic reticulum are highly interdependent structures. J Cell Biol. 1986 Oct;103(4):1557–1568. doi: 10.1083/jcb.103.4.1557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Watanabe N., Broome M., Hunter T. Regulation of the human WEE1Hu CDK tyrosine 15-kinase during the cell cycle. EMBO J. 1995 May 1;14(9):1878–1891. doi: 10.1002/j.1460-2075.1995.tb07180.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. van der Geer P., Hunter T. Phosphopeptide mapping and phosphoamino acid analysis by electrophoresis and chromatography on thin-layer cellulose plates. Electrophoresis. 1994 Mar-Apr;15(3-4):544–554. doi: 10.1002/elps.1150150173. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES