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. 1994 Nov 1;13(21):5203–5211. doi: 10.1002/j.1460-2075.1994.tb06851.x

A zinc finger protein, essential for chromosome segregation, constitutes a putative DNA binding subunit of the Saccharomyces cerevisiae kinetochore complex, Cbf3.

J Lechner 1
PMCID: PMC395469  PMID: 7957085

Abstract

A multisubunit protein complex, Cbf3, is a component of the Saccharomyces cerevisiae kinetochore. Cbf3 was recently shown to be essential for chromosome segregation in vivo and for movement of centromere DNA (CEN) along microtubules in vitro. Cbf3 contains three proteins, Cbf3a, Cbf3b and Cbf3c. Here the characterization of Cbf3b is described. Cbf3b contains an N-terminal Zn2Cys6 type zinc finger domain, a C-terminal acidic domain and a putative coiled coil dimerization domain. Cbf3b is essential for growth. Mutations within the zinc finger domain result in cells that exhibit a G2-M cell cycle delay and increased chromosome loss in each mitotic cell division. Therefore, Cbf3b has an essential function in chromosome segregation and the zinc finger domain executes part of this function presumably by providing the specific interaction between Cbf3 and CEN. Finally, data are provided to show that Cbf3c is encoded by CTF13, a gene that had been cloned recently by complementing a temperature sensitive mutant that exhibits chromosome loss as a result of a defective centromere.

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Selected References

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

  1. Baker R. E., Fitzgerald-Hayes M., O'Brien T. C. Purification of the yeast centromere binding protein CP1 and a mutational analysis of its binding site. J Biol Chem. 1989 Jun 25;264(18):10843–10850. [PubMed] [Google Scholar]
  2. Baker R. E., Masison D. C. Isolation of the gene encoding the Saccharomyces cerevisiae centromere-binding protein CP1. Mol Cell Biol. 1990 Jun;10(6):2458–2467. doi: 10.1128/mcb.10.6.2458. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bloom K. The centromere frontier: kinetochore components, microtubule-based motility, and the CEN-value paradox. Cell. 1993 May 21;73(4):621–624. doi: 10.1016/0092-8674(93)90242-i. [DOI] [PubMed] [Google Scholar]
  4. Bram R. J., Kornberg R. D. Isolation of a Saccharomyces cerevisiae centromere DNA-binding protein, its human homolog, and its possible role as a transcription factor. Mol Cell Biol. 1987 Jan;7(1):403–409. doi: 10.1128/mcb.7.1.403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brinkley B. R. Centromeres and kinetochores: integrated domains on eukaryotic chromosomes. Curr Opin Cell Biol. 1990 Jun;2(3):446–452. doi: 10.1016/0955-0674(90)90126-y. [DOI] [PubMed] [Google Scholar]
  6. Cai M. J., Davis R. W. Purification of a yeast centromere-binding protein that is able to distinguish single base-pair mutations in its recognition site. Mol Cell Biol. 1989 Jun;9(6):2544–2550. doi: 10.1128/mcb.9.6.2544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cai M., Davis R. W. Yeast centromere binding protein CBF1, of the helix-loop-helix protein family, is required for chromosome stability and methionine prototrophy. Cell. 1990 May 4;61(3):437–446. doi: 10.1016/0092-8674(90)90525-j. [DOI] [PubMed] [Google Scholar]
  8. Clarke L., Carbon J. Genomic substitutions of centromeres in Saccharomyces cerevisiae. Nature. 1983 Sep 1;305(5929):23–28. doi: 10.1038/305023a0. [DOI] [PubMed] [Google Scholar]
  9. Clarke L., Carbon J. The structure and function of yeast centromeres. Annu Rev Genet. 1985;19:29–55. doi: 10.1146/annurev.ge.19.120185.000333. [DOI] [PubMed] [Google Scholar]
  10. Clarke L. Centromeres of budding and fission yeasts. Trends Genet. 1990 May;6(5):150–154. doi: 10.1016/0168-9525(90)90149-z. [DOI] [PubMed] [Google Scholar]
  11. Cumberledge S., Carbon J. Mutational analysis of meiotic and mitotic centromere function in Saccharomyces cerevisiae. Genetics. 1987 Oct;117(2):203–212. doi: 10.1093/genetics/117.2.203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Doheny K. F., Sorger P. K., Hyman A. A., Tugendreich S., Spencer F., Hieter P. Identification of essential components of the S. cerevisiae kinetochore. Cell. 1993 May 21;73(4):761–774. doi: 10.1016/0092-8674(93)90255-O. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gaudet A., Fitzgerald-Hayes M. Alterations in the adenine-plus-thymine-rich region of CEN3 affect centromere function in Saccharomyces cerevisiae. Mol Cell Biol. 1987 Jan;7(1):68–75. doi: 10.1128/mcb.7.1.68. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Goh P. Y., Kilmartin J. V. NDC10: a gene involved in chromosome segregation in Saccharomyces cerevisiae. J Cell Biol. 1993 May;121(3):503–512. doi: 10.1083/jcb.121.3.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. He Z., Brinton B. T., Greenblatt J., Hassell J. A., Ingles C. J. The transactivator proteins VP16 and GAL4 bind replication factor A. Cell. 1993 Jun 18;73(6):1223–1232. doi: 10.1016/0092-8674(93)90650-f. [DOI] [PubMed] [Google Scholar]
  16. Hegemann J. H., Fleig U. N. The centromere of budding yeast. Bioessays. 1993 Jul;15(7):451–460. doi: 10.1002/bies.950150704. [DOI] [PubMed] [Google Scholar]
  17. Hegemann J. H., Shero J. H., Cottarel G., Philippsen P., Hieter P. Mutational analysis of centromere DNA from chromosome VI of Saccharomyces cerevisiae. Mol Cell Biol. 1988 Jun;8(6):2523–2535. doi: 10.1128/mcb.8.6.2523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hyman A. A., Middleton K., Centola M., Mitchison T. J., Carbon J. Microtubule-motor activity of a yeast centromere-binding protein complex. Nature. 1992 Oct 8;359(6395):533–536. doi: 10.1038/359533a0. [DOI] [PubMed] [Google Scholar]
  19. Hyman A. A., Mitchison T. J. Two different microtubule-based motor activities with opposite polarities in kinetochores. Nature. 1991 May 16;351(6323):206–211. doi: 10.1038/351206a0. [DOI] [PubMed] [Google Scholar]
  20. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jehn B., Niedenthal R., Hegemann J. H. In vivo analysis of the Saccharomyces cerevisiae centromere CDEIII sequence: requirements for mitotic chromosome segregation. Mol Cell Biol. 1991 Oct;11(10):5212–5221. doi: 10.1128/mcb.11.10.5212. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jiang W. D., Philippsen P. Purification of a protein binding to the CDEI subregion of Saccharomyces cerevisiae centromere DNA. Mol Cell Biol. 1989 Dec;9(12):5585–5593. doi: 10.1128/mcb.9.12.5585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jiang W., Lechner J., Carbon J. Isolation and characterization of a gene (CBF2) specifying a protein component of the budding yeast kinetochore. J Cell Biol. 1993 May;121(3):513–519. doi: 10.1083/jcb.121.3.513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jiang W., Middleton K., Yoon H. J., Fouquet C., Carbon J. An essential yeast protein, CBF5p, binds in vitro to centromeres and microtubules. Mol Cell Biol. 1993 Aug;13(8):4884–4893. doi: 10.1128/mcb.13.8.4884. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Johnston M., Dover J. Mutational analysis of the GAL4-encoded transcriptional activator protein of Saccharomyces cerevisiae. Genetics. 1988 Sep;120(1):63–74. doi: 10.1093/genetics/120.1.63. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Keegan L., Gill G., Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein. Science. 1986 Feb 14;231(4739):699–704. doi: 10.1126/science.3080805. [DOI] [PubMed] [Google Scholar]
  27. Kingsbury J., Koshland D. Centromere-dependent binding of yeast minichromosomes to microtubules in vitro. Cell. 1991 Aug 9;66(3):483–495. doi: 10.1016/0092-8674(81)90012-x. [DOI] [PubMed] [Google Scholar]
  28. Kraulis P. J., Raine A. R., Gadhavi P. L., Laue E. D. Structure of the DNA-binding domain of zinc GAL4. Nature. 1992 Apr 2;356(6368):448–450. doi: 10.1038/356448a0. [DOI] [PubMed] [Google Scholar]
  29. Lechner J., Carbon J. A 240 kd multisubunit protein complex, CBF3, is a major component of the budding yeast centromere. Cell. 1991 Feb 22;64(4):717–725. doi: 10.1016/0092-8674(91)90501-o. [DOI] [PubMed] [Google Scholar]
  30. Lechner J., Sumper M. The primary structure of a procaryotic glycoprotein. Cloning and sequencing of the cell surface glycoprotein gene of halobacteria. J Biol Chem. 1987 Jul 15;262(20):9724–9729. [PubMed] [Google Scholar]
  31. Li R., Botchan M. R. The acidic transcriptional activation domains of VP16 and p53 bind the cellular replication protein A and stimulate in vitro BPV-1 DNA replication. Cell. 1993 Jun 18;73(6):1207–1221. doi: 10.1016/0092-8674(93)90649-b. [DOI] [PubMed] [Google Scholar]
  32. Ma J., Ptashne M. The carboxy-terminal 30 amino acids of GAL4 are recognized by GAL80. Cell. 1987 Jul 3;50(1):137–142. doi: 10.1016/0092-8674(87)90670-2. [DOI] [PubMed] [Google Scholar]
  33. Marmorstein R., Carey M., Ptashne M., Harrison S. C. DNA recognition by GAL4: structure of a protein-DNA complex. Nature. 1992 Apr 2;356(6368):408–414. doi: 10.1038/356408a0. [DOI] [PubMed] [Google Scholar]
  34. McGrew J., Diehl B., Fitzgerald-Hayes M. Single base-pair mutations in centromere element III cause aberrant chromosome segregation in Saccharomyces cerevisiae. Mol Cell Biol. 1986 Feb;6(2):530–538. doi: 10.1128/mcb.6.2.530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mellor J., Jiang W., Funk M., Rathjen J., Barnes C. A., Hinz T., Hegemann J. H., Philippsen P. CPF1, a yeast protein which functions in centromeres and promoters. EMBO J. 1990 Dec;9(12):4017–4026. doi: 10.1002/j.1460-2075.1990.tb07623.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Ng R., Carbon J. Mutational and in vitro protein-binding studies on centromere DNA from Saccharomyces cerevisiae. Mol Cell Biol. 1987 Dec;7(12):4522–4534. doi: 10.1128/mcb.7.12.4522. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Niedenthal R., Stoll R., Hegemann J. H. In vivo characterization of the Saccharomyces cerevisiae centromere DNA element I, a binding site for the helix-loop-helix protein CPF1. Mol Cell Biol. 1991 Jul;11(7):3545–3553. doi: 10.1128/mcb.11.7.3545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pfarr C. M., Coue M., Grissom P. M., Hays T. S., Porter M. E., McIntosh J. R. Cytoplasmic dynein is localized to kinetochores during mitosis. Nature. 1990 May 17;345(6272):263–265. doi: 10.1038/345263a0. [DOI] [PubMed] [Google Scholar]
  39. Philippsen P., Stotz A., Scherf C. DNA of Saccharomyces cerevisiae. Methods Enzymol. 1991;194:169–182. doi: 10.1016/0076-6879(91)94014-4. [DOI] [PubMed] [Google Scholar]
  40. Pluta A. F., Cooke C. A., Earnshaw W. C. Structure of the human centromere at metaphase. Trends Biochem Sci. 1990 May;15(5):181–185. doi: 10.1016/0968-0004(90)90158-8. [DOI] [PubMed] [Google Scholar]
  41. Ptashne M., Gann A. A. Activators and targets. Nature. 1990 Jul 26;346(6282):329–331. doi: 10.1038/346329a0. [DOI] [PubMed] [Google Scholar]
  42. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Schulman I., Bloom K. S. Centromeres: an integrated protein/DNA complex required for chromosome movement. Annu Rev Cell Biol. 1991;7:311–336. doi: 10.1146/annurev.cb.07.110191.001523. [DOI] [PubMed] [Google Scholar]
  44. Sharp P. M., Tuohy T. M., Mosurski K. R. Codon usage in yeast: cluster analysis clearly differentiates highly and lowly expressed genes. Nucleic Acids Res. 1986 Jul 11;14(13):5125–5143. doi: 10.1093/nar/14.13.5125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Sherman F. Getting started with yeast. Methods Enzymol. 1991;194:3–21. doi: 10.1016/0076-6879(91)94004-v. [DOI] [PubMed] [Google Scholar]
  46. Sikorski R. S., Boeke J. D. In vitro mutagenesis and plasmid shuffling: from cloned gene to mutant yeast. Methods Enzymol. 1991;194:302–318. doi: 10.1016/0076-6879(91)94023-6. [DOI] [PubMed] [Google Scholar]
  47. Sikorski R. S., Hieter P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics. 1989 May;122(1):19–27. doi: 10.1093/genetics/122.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Spencer F., Hieter P. Centromere DNA mutations induce a mitotic delay in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):8908–8912. doi: 10.1073/pnas.89.19.8908. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Steuer E. R., Wordeman L., Schroer T. A., Sheetz M. P. Localization of cytoplasmic dynein to mitotic spindles and kinetochores. Nature. 1990 May 17;345(6272):266–268. doi: 10.1038/345266a0. [DOI] [PubMed] [Google Scholar]
  50. Xiao Z., McGrew J. T., Schroeder A. J., Fitzgerald-Hayes M. CSE1 and CSE2, two new genes required for accurate mitotic chromosome segregation in Saccharomyces cerevisiae. Mol Cell Biol. 1993 Aug;13(8):4691–4702. doi: 10.1128/mcb.13.8.4691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Yen T. J., Li G., Schaar B. T., Szilak I., Cleveland D. W. CENP-E is a putative kinetochore motor that accumulates just before mitosis. Nature. 1992 Oct 8;359(6395):536–539. doi: 10.1038/359536a0. [DOI] [PubMed] [Google Scholar]

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