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. 1991 Aug;10(8):2069–2075. doi: 10.1002/j.1460-2075.1991.tb07738.x

A cdc2-like kinase phosphorylates histone H1 in the amitotic macronucleus of Tetrahymena.

S Y Roth 1, M P Collini 1, G Draetta 1, D Beach 1, C D Allis 1
PMCID: PMC452890  PMID: 2065655

Abstract

Genetic and biochemical studies have shown that cdc2 protein kinase plays a pivotal role in a highly conserved mechanism controlling the entry of cells into mitosis. It is generally believed that one function of cdc2 kinase is to phosphorylate histone H1 which in turn promotes mitotic chromosome condensation. However, direct evidence linking H1 phosphorylation to mitotic chromatin condensation is limited and the exact cellular function(s) of H1 phosphorylation remains unclear. In this study, we show that mammalian cdc2 kinase phosphorylates H1 from the amitotic macronucleus of Tetrahymena with remarkable fidelity. Furthermore, we demonstrate that macronuclei from Tetrahymena contain a growth-associated H1 kinase activity which closely resembles cdc2 kinase from other eukaryotes. Using polyclonal antibodies raised against yeast p34cdc2, we have detected a 36 kd immunoactive polypeptide in macronuclei which binds to Suc1 (p13)-coated beads and closely follows H1 kinase activity. Since macronuclei divide without mitotic chromosome condensation, these data demonstrate that H1 phosphorylation by cdc2 kinase may be necessary, but is not sufficient to promote mitotic chromatin condensation. The fact that an activity which strongly resembles mammalian cdc2 kinase is active during cell growth in a nucleus which does not undergo mitosis and chromosome condensation suggests that other factors are needed for a true mitotic division to occur. These data also reinforce the notion that H1 phosphorylation has important functions outside mitosis both in Tetrahymena and in mammalian cells.

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Selected References

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

  1. Allan J., Hartman P. G., Crane-Robinson C., Aviles F. X. The structure of histone H1 and its location in chromatin. Nature. 1980 Dec 25;288(5792):675–679. doi: 10.1038/288675a0. [DOI] [PubMed] [Google Scholar]
  2. Allan J., Mitchell T., Harborne N., Bohm L., Crane-Robinson C. Roles of H1 domains in determining higher order chromatin structure and H1 location. J Mol Biol. 1986 Feb 20;187(4):591–601. doi: 10.1016/0022-2836(86)90337-2. [DOI] [PubMed] [Google Scholar]
  3. Allis C. D., Gorovsky M. A. Histone phosphorylation in macro- and micronuclei of Tetrahymena thermophila. Biochemistry. 1981 Jun 23;20(13):3828–3833. doi: 10.1021/bi00516a025. [DOI] [PubMed] [Google Scholar]
  4. Arion D., Meijer L., Brizuela L., Beach D. cdc2 is a component of the M phase-specific histone H1 kinase: evidence for identity with MPF. Cell. 1988 Oct 21;55(2):371–378. doi: 10.1016/0092-8674(88)90060-8. [DOI] [PubMed] [Google Scholar]
  5. Blow J. J., Nurse P. A cdc2-like protein is involved in the initiation of DNA replication in Xenopus egg extracts. Cell. 1990 Sep 7;62(5):855–862. doi: 10.1016/0092-8674(90)90261-c. [DOI] [PubMed] [Google Scholar]
  6. Booher R. N., Alfa C. E., Hyams J. S., Beach D. H. The fission yeast cdc2/cdc13/suc1 protein kinase: regulation of catalytic activity and nuclear localization. Cell. 1989 Aug 11;58(3):485–497. doi: 10.1016/0092-8674(89)90429-7. [DOI] [PubMed] [Google Scholar]
  7. Bradbury E. M., Inglis R. J., Matthews H. R. Control of cell division by very lysine rich histone (F1) phosphorylation. Nature. 1974 Feb 1;247(5439):257–261. doi: 10.1038/247257a0. [DOI] [PubMed] [Google Scholar]
  8. Bradbury E. M., Inglis R. J., Matthews H. R., Langan T. A. Molecular basis of control of mitotic cell division in eukaryotes. Nature. 1974 Jun 7;249(457):553–556. doi: 10.1038/249553a0. [DOI] [PubMed] [Google Scholar]
  9. Brizuela L., Draetta G., Beach D. Activation of human CDC2 protein as a histone H1 kinase is associated with complex formation with the p62 subunit. Proc Natl Acad Sci U S A. 1989 Jun;86(12):4362–4366. doi: 10.1073/pnas.86.12.4362. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Brizuela L., Draetta G., Beach D. p13suc1 acts in the fission yeast cell division cycle as a component of the p34cdc2 protein kinase. EMBO J. 1987 Nov;6(11):3507–3514. doi: 10.1002/j.1460-2075.1987.tb02676.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Chambers T. C., Langan T. A., Matthews H. R., Bradbury E. M. H1 histone kinases from nuclei of Physarum polycephalum. Biochemistry. 1983 Jan 4;22(1):30–37. doi: 10.1021/bi00270a005. [DOI] [PubMed] [Google Scholar]
  12. Churchill M. E., Suzuki M. 'SPKK' motifs prefer to bind to DNA at A/T-rich sites. EMBO J. 1989 Dec 20;8(13):4189–4195. doi: 10.1002/j.1460-2075.1989.tb08604.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Cyert M. S., Thorner J. Putting it on and taking it off: phosphoprotein phosphatase involvement in cell cycle regulation. Cell. 1989 Jun 16;57(6):891–893. doi: 10.1016/0092-8674(89)90325-5. [DOI] [PubMed] [Google Scholar]
  14. Dedon P. C., Soults J. A., Allis C. D., Gorovsky M. A. Formaldehyde cross-linking and immunoprecipitation demonstrate developmental changes in H1 association with transcriptionally active genes. Mol Cell Biol. 1991 Mar;11(3):1729–1733. doi: 10.1128/mcb.11.3.1729. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Draetta G., Brizuela L., Potashkin J., Beach D. Identification of p34 and p13, human homologs of the cell cycle regulators of fission yeast encoded by cdc2+ and suc1+. Cell. 1987 Jul 17;50(2):319–325. doi: 10.1016/0092-8674(87)90227-3. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Dunphy W. G., Newport J. W. Unraveling of mitotic control mechanisms. Cell. 1988 Dec 23;55(6):925–928. doi: 10.1016/0092-8674(88)90234-6. [DOI] [PubMed] [Google Scholar]
  18. Ericsson C., Grossbach U., Björkroth B., Daneholt B. Presence of histone H1 on an active Balbiani ring gene. Cell. 1990 Jan 12;60(1):73–83. doi: 10.1016/0092-8674(90)90717-s. [DOI] [PubMed] [Google Scholar]
  19. Glover C. V., Vavra K. J., Guttman S. D., Gorovsky M. A. Heat shock and deciliation induce phosphorylation of histone H1 in T. pyriformis. Cell. 1981 Jan;23(1):73–77. doi: 10.1016/0092-8674(81)90271-3. [DOI] [PubMed] [Google Scholar]
  20. Gorovsky M. A. Macro- and micronuclei of Tetrahymena pyriformis: a model system for studying the structure and function of eukaryotic nuclei. J Protozool. 1973 Feb;20(1):19–25. doi: 10.1111/j.1550-7408.1973.tb05995.x. [DOI] [PubMed] [Google Scholar]
  21. Gorovsky M. A., Yao M. C., Keevert J. B., Pleger G. L. Isolation of micro- and macronuclei of Tetrahymena pyriformis. Methods Cell Biol. 1975;9(0):311–327. doi: 10.1016/s0091-679x(08)60080-1. [DOI] [PubMed] [Google Scholar]
  22. Hartwell L. H., Weinert T. A. Checkpoints: controls that ensure the order of cell cycle events. Science. 1989 Nov 3;246(4930):629–634. doi: 10.1126/science.2683079. [DOI] [PubMed] [Google Scholar]
  23. Hayashi T., Hayashi H., Iwai K. Tetrahymena histone H1. Isolation and amino acid sequence lacking the central hydrophobic domain conserved in other H1 histones. J Biochem. 1987 Aug;102(2):369–376. doi: 10.1093/oxfordjournals.jbchem.a122063. [DOI] [PubMed] [Google Scholar]
  24. Hayles J., Beach D., Durkacz B., Nurse P. The fission yeast cell cycle control gene cdc2: isolation of a sequence suc1 that suppresses cdc2 mutant function. Mol Gen Genet. 1986 Feb;202(2):291–293. doi: 10.1007/BF00331653. [DOI] [PubMed] [Google Scholar]
  25. Hill C. S., Packman L. C., Thomas J. O. Phosphorylation at clustered -Ser-Pro-X-Lys/Arg- motifs in sperm-specific histones H1 and H2B. EMBO J. 1990 Mar;9(3):805–813. doi: 10.1002/j.1460-2075.1990.tb08177.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Hill R. J., Watt F., Wilson C. M., Fifis T., Underwood P. A., Tribbick G., Geysen H. M., Thomas J. O. Bands, interbands and puffs in native Drosophila polytene chromosomes are recognized by a monoclonal antibody to an epitope in the carboxy-terminal tail of histone H1. Chromosoma. 1989 Dec;98(6):411–421. doi: 10.1007/BF00292786. [DOI] [PubMed] [Google Scholar]
  27. Hindley J., Phear G., Stein M., Beach D. Sucl+ encodes a predicted 13-kilodalton protein that is essential for cell viability and is directly involved in the division cycle of Schizosaccharomyces pombe. Mol Cell Biol. 1987 Jan;7(1):504–511. doi: 10.1128/mcb.7.1.504. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Hohmann P. Phosphorylation of H1 histones. Mol Cell Biochem. 1983;57(1):81–92. doi: 10.1007/BF00223526. [DOI] [PubMed] [Google Scholar]
  29. Inglis R. J., Langan T. A., Matthews H. R., Hardie D. G., Bradbury E. M. Advance of mitosis by histone phosphokinase. Exp Cell Res. 1976 Feb;97(2):418–425. doi: 10.1016/0014-4827(76)90634-0. [DOI] [PubMed] [Google Scholar]
  30. 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]
  31. 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]
  32. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  33. Lake R. S. Further characterization of the F1-histone phosphokinase of metaphase-arrested animal cells. J Cell Biol. 1973 Aug;58(2):317–331. doi: 10.1083/jcb.58.2.317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Lake R. S., Salzman N. P. Occurrence and properties of a chromatin-associated F1-histone phosphokinase in mitotic Chinese hamster cells. Biochemistry. 1972 Dec 5;11(25):4817–4826. doi: 10.1021/bi00775a027. [DOI] [PubMed] [Google Scholar]
  35. Lamb N. J., Fernandez A., Watrin A., Labbé J. C., Cavadore J. C. Microinjection of p34cdc2 kinase induces marked changes in cell shape, cytoskeletal organization, and chromatin structure in mammalian fibroblasts. Cell. 1990 Jan 12;60(1):151–165. doi: 10.1016/0092-8674(90)90725-t. [DOI] [PubMed] [Google Scholar]
  36. 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]
  37. Lee M. G., Nurse P. Complementation used to clone a human homologue of the fission yeast cell cycle control gene cdc2. Nature. 1987 May 7;327(6117):31–35. doi: 10.1038/327031a0. [DOI] [PubMed] [Google Scholar]
  38. Lewin B. Driving the cell cycle: M phase kinase, its partners, and substrates. Cell. 1990 Jun 1;61(5):743–752. doi: 10.1016/0092-8674(90)90181-d. [DOI] [PubMed] [Google Scholar]
  39. Lohka M. J., Hayes M. K., Maller J. L. Purification of maturation-promoting factor, an intracellular regulator of early mitotic events. Proc Natl Acad Sci U S A. 1988 May;85(9):3009–3013. doi: 10.1073/pnas.85.9.3009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Matsumoto Y., Yasuda H., Mita S., Marunouchi T., Yamada M. Evidence for the involvement of H1 histone phosphorylation in chromosome condensation. Nature. 1980 Mar 13;284(5752):181–183. doi: 10.1038/284181a0. [DOI] [PubMed] [Google Scholar]
  41. Moreno S., Nurse P. Substrates for p34cdc2: in vivo veritas? Cell. 1990 May 18;61(4):549–551. doi: 10.1016/0092-8674(90)90463-o. [DOI] [PubMed] [Google Scholar]
  42. 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]
  43. Mueller R. D., Yasuda H., Bradbury E. M. Phosphorylation of histone H1 through the cell cycle of Physarum polycephalum. 24 sites of phosphorylation at metaphase. J Biol Chem. 1985 Apr 25;260(8):5081–5086. [PubMed] [Google Scholar]
  44. Murray A. W., Kirschner M. W. Dominoes and clocks: the union of two views of the cell cycle. Science. 1989 Nov 3;246(4930):614–621. doi: 10.1126/science.2683077. [DOI] [PubMed] [Google Scholar]
  45. Nacheva G. A., Guschin D. Y., Preobrazhenskaya O. V., Karpov V. L., Ebralidse K. K., Mirzabekov A. D. Change in the pattern of histone binding to DNA upon transcriptional activation. Cell. 1989 Jul 14;58(1):27–36. doi: 10.1016/0092-8674(89)90399-1. [DOI] [PubMed] [Google Scholar]
  46. Pines J., Hunter T. p34cdc2: the S and M kinase? New Biol. 1990 May;2(5):389–401. [PubMed] [Google Scholar]
  47. Roth S. Y., Schulman I. G., Richman R., Cook R. G., Allis C. D. Characterization of phosphorylation sites in histone H1 in the amitotic macronucleus of Tetrahymena during different physiological states. J Cell Biol. 1988 Dec;107(6 Pt 2):2473–2482. doi: 10.1083/jcb.107.6.2473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Schulman I. G., Cook R. G., Richman R., Allis C. D. Tetrahymena contain two distinct and unusual high mobility group (HMG)-like proteins. J Cell Biol. 1987 Jun;104(6):1485–1494. doi: 10.1083/jcb.104.6.1485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Suzuki M. SPKK, a new nucleic acid-binding unit of protein found in histone. EMBO J. 1989 Mar;8(3):797–804. doi: 10.1002/j.1460-2075.1989.tb03440.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Suzuki M. SPXX, a frequent sequence motif in gene regulatory proteins. J Mol Biol. 1989 May 5;207(1):61–84. doi: 10.1016/0022-2836(89)90441-5. [DOI] [PubMed] [Google Scholar]
  51. Suzuki M., Sohma H., Yazawa M., Yagi K., Ebashi S. Histone H1 kinase specific to the SPKK motif. J Biochem. 1990 Sep;108(3):356–364. doi: 10.1093/oxfordjournals.jbchem.a123206. [DOI] [PubMed] [Google Scholar]
  52. Suzuki M. The heptad repeat in the largest subunit of RNA polymerase II binds by intercalating into DNA. Nature. 1990 Apr 5;344(6266):562–565. doi: 10.1038/344562a0. [DOI] [PubMed] [Google Scholar]
  53. Thoma F., Koller T., Klug A. Involvement of histone H1 in the organization of the nucleosome and of the salt-dependent superstructures of chromatin. J Cell Biol. 1979 Nov;83(2 Pt 1):403–427. doi: 10.1083/jcb.83.2.403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Weintraub H. Assembly and propagation of repressed and depressed chromosomal states. Cell. 1985 Oct;42(3):705–711. doi: 10.1016/0092-8674(85)90267-3. [DOI] [PubMed] [Google Scholar]
  55. Weintraub H. Histone-H1-dependent chromatin superstructures and the suppression of gene activity. Cell. 1984 Aug;38(1):17–27. doi: 10.1016/0092-8674(84)90522-1. [DOI] [PubMed] [Google Scholar]
  56. Wittenberg C., Sugimoto K., Reed S. I. G1-specific cyclins of S. cerevisiae: cell cycle periodicity, regulation by mating pheromone, and association with the p34CDC28 protein kinase. Cell. 1990 Jul 27;62(2):225–237. doi: 10.1016/0092-8674(90)90361-h. [DOI] [PubMed] [Google Scholar]
  57. Wolffe A. P. Dominant and specific repression of Xenopus oocyte 5S RNA genes and satellite I DNA by histone H1. EMBO J. 1989 Feb;8(2):527–537. doi: 10.1002/j.1460-2075.1989.tb03407.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Woodford T. A., Pardee A. B. Histone H1 kinase in exponential and synchronous populations of Chinese hamster fibroblasts. J Biol Chem. 1986 Apr 5;261(10):4669–4676. [PubMed] [Google Scholar]
  59. Wu M., Allis C. D., Richman R., Cook R. G., Gorovsky M. A. An intervening sequence in an unusual histone H1 gene of Tetrahymena thermophila. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8674–8678. doi: 10.1073/pnas.83.22.8674. [DOI] [PMC free article] [PubMed] [Google Scholar]

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