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. 1995 May 11;23(9):1454–1460. doi: 10.1093/nar/23.9.1454

Protein-DNA interactions in soluble telosomes from Saccharomyces cerevisiae.

J H Wright 1, V A Zakian 1
PMCID: PMC306882  PMID: 7784196

Abstract

Telomeric DNA in Saccharomyces is organized into a non-nucleosomal chromatin structure called the telosome that can be released from chromosome ends in soluble form by nuclease digestion (Wright, J. H., Gottschling, D. E. and Zakian, V. A. (1992) Genes Dev. 6, 197-210). The protein-DNA interactions of soluble telosomes were investigated by monitoring isolated telomeric DNA fragments for the retention of bound protein using both gel mobility shift and nitrocellulose filter-binding assays. Telosomal proteins remained associated with telomeric DNA at concentrations of ethidium bromide that dissociated nucleosomes. The protein-DNA interactions in the yeast telosome were also disrupted by much lower salt concentrations than those known to disrupt either the interactions of ciliate terminus-binding proteins with telomeric DNA or the interactions of histones with DNA in nucleosomes. Taken together, these data corroborate previously published nuclease mapping data indicating that telosomes are distinct in structure from conventional nucleosomes. These data also indicate that yeast do not possess telomere binding proteins similar to those detected in ciliates that remain tightly bound to telomeric DNA even in high salt. In addition, the characteristic gel mobility shift of soluble telosomes could be mimicked by complexes formed in vitro with yeast telomeric DNA and recombinant Rap1p suggesting that Rap1p, a known component of soluble yeast telosomes (Wright, J. H., Gottschling, D. E. and Zakian, V. A. (1992) Genes Dev. 6, 197-210; Conrad, M. N., Wright, J. H., Wolf, A. J. and Zakian, V. A. (1990) Cell 63, 739-750), is likely to be the major structural protein bound directly to yeast telomeric DNA.

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Selected References

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

  1. Blackburn E. H., Chiou S. S. Non-nucleosomal packaging of a tandemly repeated DNA sequence at termini of extrachromosomal DNA coding for rRNA in Tetrahymena. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2263–2267. doi: 10.1073/pnas.78.4.2263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blackburn E. H. Telomerases. Annu Rev Biochem. 1992;61:113–129. doi: 10.1146/annurev.bi.61.070192.000553. [DOI] [PubMed] [Google Scholar]
  3. Buchman A. R., Kimmerly W. J., Rine J., Kornberg R. D. Two DNA-binding factors recognize specific sequences at silencers, upstream activating sequences, autonomously replicating sequences, and telomeres in Saccharomyces cerevisiae. Mol Cell Biol. 1988 Jan;8(1):210–225. doi: 10.1128/mcb.8.1.210. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Budarf M. L., Blackburn E. H. Chromatin structure of the telomeric region and 3'-nontranscribed spacer of Tetrahymena ribosomal RNA genes. J Biol Chem. 1986 Jan 5;261(1):363–369. [PubMed] [Google Scholar]
  5. Burton D. R., Butler M. J., Hyde J. E., Phillips D., Skidmore C. J., Walker I. O. The interaction of core histones with DNA: equilibrium binding studies. Nucleic Acids Res. 1978 Oct;5(10):3643–3663. doi: 10.1093/nar/5.10.3643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Conrad M. N., Wright J. H., Wolf A. J., Zakian V. A. RAP1 protein interacts with yeast telomeres in vivo: overproduction alters telomere structure and decreases chromosome stability. Cell. 1990 Nov 16;63(4):739–750. doi: 10.1016/0092-8674(90)90140-a. [DOI] [PubMed] [Google Scholar]
  7. Fang G. W., Cech T. R. Molecular cloning of telomere-binding protein genes from Stylonychia mytilis. Nucleic Acids Res. 1991 Oct 25;19(20):5515–5518. doi: 10.1093/nar/19.20.5515. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fang G., Gray J. T., Cech T. R. Oxytricha telomere-binding protein: separable DNA-binding and dimerization domains of the alpha-subunit. Genes Dev. 1993 May;7(5):870–882. doi: 10.1101/gad.7.5.870. [DOI] [PubMed] [Google Scholar]
  9. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  10. Funabiki H., Hagan I., Uzawa S., Yanagida M. Cell cycle-dependent specific positioning and clustering of centromeres and telomeres in fission yeast. J Cell Biol. 1993 Jun;121(5):961–976. doi: 10.1083/jcb.121.5.961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Giesman D., Best L., Tatchell K. The role of RAP1 in the regulation of the MAT alpha locus. Mol Cell Biol. 1991 Feb;11(2):1069–1079. doi: 10.1128/mcb.11.2.1069. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gilson E., Laroche T., Gasser S. M. Telomeres and the functional architecture of the nucleus. Trends Cell Biol. 1993 Apr;3(4):128–134. doi: 10.1016/0962-8924(93)90175-z. [DOI] [PubMed] [Google Scholar]
  13. Gilson E., Roberge M., Giraldo R., Rhodes D., Gasser S. M. Distortion of the DNA double helix by RAP1 at silencers and multiple telomeric binding sites. J Mol Biol. 1993 May 20;231(2):293–310. doi: 10.1006/jmbi.1993.1283. [DOI] [PubMed] [Google Scholar]
  14. Gottschling D. E., Zakian V. A. Telomere proteins: specific recognition and protection of the natural termini of Oxytricha macronuclear DNA. Cell. 1986 Oct 24;47(2):195–205. doi: 10.1016/0092-8674(86)90442-3. [DOI] [PubMed] [Google Scholar]
  15. Hardy C. F., Sussel L., Shore D. A RAP1-interacting protein involved in transcriptional silencing and telomere length regulation. Genes Dev. 1992 May;6(5):801–814. doi: 10.1101/gad.6.5.801. [DOI] [PubMed] [Google Scholar]
  16. Henson P., Walker I. O. The partial dissociation of nucleohistone by salts. Hydrodynamic and denaturation studies. Eur J Biochem. 1970 Jun;14(2):345–350. doi: 10.1111/j.1432-1033.1970.tb00295.x. [DOI] [PubMed] [Google Scholar]
  17. Hofmann J. F., Laroche T., Brand A. H., Gasser S. M. RAP-1 factor is necessary for DNA loop formation in vitro at the silent mating type locus HML. Cell. 1989 Jun 2;57(5):725–737. doi: 10.1016/0092-8674(89)90788-5. [DOI] [PubMed] [Google Scholar]
  18. Johnston R. F., Pickett S. C., Barker D. L. Autoradiography using storage phosphor technology. Electrophoresis. 1990 May;11(5):355–360. doi: 10.1002/elps.1150110503. [DOI] [PubMed] [Google Scholar]
  19. Klein F., Laroche T., Cardenas M. E., Hofmann J. F., Schweizer D., Gasser S. M. Localization of RAP1 and topoisomerase II in nuclei and meiotic chromosomes of yeast. J Cell Biol. 1992 Jun;117(5):935–948. doi: 10.1083/jcb.117.5.935. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Klobutcher L. A., Swanton M. T., Donini P., Prescott D. M. All gene-sized DNA molecules in four species of hypotrichs have the same terminal sequence and an unusual 3' terminus. Proc Natl Acad Sci U S A. 1981 May;78(5):3015–3019. doi: 10.1073/pnas.78.5.3015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kyrion G., Liu K., Liu C., Lustig A. J. RAP1 and telomere structure regulate telomere position effects in Saccharomyces cerevisiae. Genes Dev. 1993 Jul;7(7A):1146–1159. doi: 10.1101/gad.7.7a.1146. [DOI] [PubMed] [Google Scholar]
  22. Liu Z. P., Tye B. K. A yeast protein that binds to vertebrate telomeres and conserved yeast telomeric junctions. Genes Dev. 1991 Jan;5(1):49–59. doi: 10.1101/gad.5.1.49. [DOI] [PubMed] [Google Scholar]
  23. Longtine M. S., Wilson N. M., Petracek M. E., Berman J. A yeast telomere binding activity binds to two related telomere sequence motifs and is indistinguishable from RAP1. Curr Genet. 1989 Oct;16(4):225–239. doi: 10.1007/BF00422108. [DOI] [PubMed] [Google Scholar]
  24. Louis E. J., Haber J. E. Evolutionarily recent transfer of a group I mitochondrial intron to telomere regions in Saccharomyces cerevisiae. Curr Genet. 1991 Nov;20(5):411–415. doi: 10.1007/BF00317070. [DOI] [PubMed] [Google Scholar]
  25. Lundblad V., Szostak J. W. A mutant with a defect in telomere elongation leads to senescence in yeast. Cell. 1989 May 19;57(4):633–643. doi: 10.1016/0092-8674(89)90132-3. [DOI] [PubMed] [Google Scholar]
  26. Lustig A. J., Kurtz S., Shore D. Involvement of the silencer and UAS binding protein RAP1 in regulation of telomere length. Science. 1990 Oct 26;250(4980):549–553. doi: 10.1126/science.2237406. [DOI] [PubMed] [Google Scholar]
  27. Lustig A. J., Petes T. D. Identification of yeast mutants with altered telomere structure. Proc Natl Acad Sci U S A. 1986 Mar;83(5):1398–1402. doi: 10.1073/pnas.83.5.1398. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Makarov V. L., Lejnine S., Bedoyan J., Langmore J. P. Nucleosomal organization of telomere-specific chromatin in rat. Cell. 1993 May 21;73(4):775–787. doi: 10.1016/0092-8674(93)90256-p. [DOI] [PubMed] [Google Scholar]
  29. McMurray C. T., van Holde K. E. Binding of ethidium bromide causes dissociation of the nucleosome core particle. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8472–8476. doi: 10.1073/pnas.83.22.8472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Norris D., Dunn B., Osley M. A. The effect of histone gene deletions on chromatin structure in Saccharomyces cerevisiae. Science. 1988 Nov 4;242(4879):759–761. doi: 10.1126/science.2847314. [DOI] [PubMed] [Google Scholar]
  31. Price C. M. Telomere structure in Euplotes crassus: characterization of DNA-protein interactions and isolation of a telomere-binding protein. Mol Cell Biol. 1990 Jul;10(7):3421–3431. doi: 10.1128/mcb.10.7.3421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Runge K. W., Zakian V. A. Introduction of extra telomeric DNA sequences into Saccharomyces cerevisiae results in telomere elongation. Mol Cell Biol. 1989 Apr;9(4):1488–1497. doi: 10.1128/mcb.9.4.1488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Runge K. W., Zakian V. A. Properties of the transcriptional enhancer in Saccharomyces cerevisiae telomeres. Nucleic Acids Res. 1990 Apr 11;18(7):1783–1787. doi: 10.1093/nar/18.7.1783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sandell L. L., Zakian V. A. Loss of a yeast telomere: arrest, recovery, and chromosome loss. Cell. 1993 Nov 19;75(4):729–739. doi: 10.1016/0092-8674(93)90493-a. [DOI] [PubMed] [Google Scholar]
  35. Saunders M. J., Yeh E., Grunstein M., Bloom K. Nucleosome depletion alters the chromatin structure of Saccharomyces cerevisiae centromeres. Mol Cell Biol. 1990 Nov;10(11):5721–5727. doi: 10.1128/mcb.10.11.5721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Schulz V. P., Zakian V. A. The saccharomyces PIF1 DNA helicase inhibits telomere elongation and de novo telomere formation. Cell. 1994 Jan 14;76(1):145–155. doi: 10.1016/0092-8674(94)90179-1. [DOI] [PubMed] [Google Scholar]
  37. Shampay J., Blackburn E. H. Generation of telomere-length heterogeneity in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1988 Jan;85(2):534–538. doi: 10.1073/pnas.85.2.534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Shampay J., Szostak J. W., Blackburn E. H. DNA sequences of telomeres maintained in yeast. Nature. 1984 Jul 12;310(5973):154–157. doi: 10.1038/310154a0. [DOI] [PubMed] [Google Scholar]
  39. Shore D., Nasmyth K. Purification and cloning of a DNA binding protein from yeast that binds to both silencer and activator elements. Cell. 1987 Dec 4;51(5):721–732. doi: 10.1016/0092-8674(87)90095-x. [DOI] [PubMed] [Google Scholar]
  40. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  41. Sussel L., Shore D. Separation of transcriptional activation and silencing functions of the RAP1-encoded repressor/activator protein 1: isolation of viable mutants affecting both silencing and telomere length. Proc Natl Acad Sci U S A. 1991 Sep 1;88(17):7749–7753. doi: 10.1073/pnas.88.17.7749. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Tommerup H., Dousmanis A., de Lange T. Unusual chromatin in human telomeres. Mol Cell Biol. 1994 Sep;14(9):5777–5785. doi: 10.1128/mcb.14.9.5777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wang S. S., Zakian V. A. Sequencing of Saccharomyces telomeres cloned using T4 DNA polymerase reveals two domains. Mol Cell Biol. 1990 Aug;10(8):4415–4419. doi: 10.1128/mcb.10.8.4415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Wright J. H., Gottschling D. E., Zakian V. A. Saccharomyces telomeres assume a non-nucleosomal chromatin structure. Genes Dev. 1992 Feb;6(2):197–210. doi: 10.1101/gad.6.2.197. [DOI] [PubMed] [Google Scholar]
  45. Zakian V. A., Runge K., Wang S. S. How does the end begin? Formation and maintenance of telomeres in ciliates and yeast. Trends Genet. 1990 Jan;6(1):12–16. doi: 10.1016/0168-9525(90)90043-6. [DOI] [PubMed] [Google Scholar]
  46. Zakian V. A. Structure and function of telomeres. Annu Rev Genet. 1989;23:579–604. doi: 10.1146/annurev.ge.23.120189.003051. [DOI] [PubMed] [Google Scholar]
  47. Zakian V. A., Wagner D. W., Fangman W. L. Yeast L double-stranded ribonucleic acid is synthesized during the G1 phase but not the S phase of the cell cycle. Mol Cell Biol. 1981 Aug;1(8):673–679. doi: 10.1128/mcb.1.8.673. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. de Lange T. Human telomeres are attached to the nuclear matrix. EMBO J. 1992 Feb;11(2):717–724. doi: 10.1002/j.1460-2075.1992.tb05104.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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