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. 1991 Feb;10(2):361–368. doi: 10.1002/j.1460-2075.1991.tb07957.x

A functional role for nucleosomes in the repression of a yeast promoter.

C Straka 1, W Hörz 1
PMCID: PMC452655  PMID: 1899374

Abstract

Induction of the PHO5 gene in S. cerevisiae was previously shown to be accompanied by the removal of four positioned nucleosomes from the promoter. In order to assess the role of nucleosomes in the cascade of gene activation, DNA corresponding to one of these nucleosomes was excised. In its place two foreign DNA segments of the same length were inserted: a fragment from the African green monkey alpha-satellite DNA which is known to associate with histones in a highly specific fashion to give a uniquely positioned nucleosome or, alternatively, a fragment derived from pBR322 DNA. The promoter constructs were fused to the lacZ gene on centromere plasmids and transformed into yeast cells. The satellite fragment formed a nucleosome which persisted under inducing conditions. At the same time the inducibility of the PHO5 promoter was virtually abolished. When various subfragments containing between 35 and 100 bp of the satellite segment were tested, they were all found to decrease the inducibility of the promoter, full repression required the full length molecule, however. In contrast, the pBR fragment made the promoter weakly constitutive, and induction proceeded to levels even higher than with a promoter lacking an insert. Analysis of the chromatin structure reveals a nucleosome on the pBR segment at noninducing conditions which is removed upon induction. It is concluded that the quality of the histone-DNA interactions at the promoter makes an intrinsic contribution to the regulation of the gene.

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Selected References

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

  1. Almer A., Hörz W. Nuclease hypersensitive regions with adjacent positioned nucleosomes mark the gene boundaries of the PHO5/PHO3 locus in yeast. EMBO J. 1986 Oct;5(10):2681–2687. doi: 10.1002/j.1460-2075.1986.tb04551.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Almer A., Rudolph H., Hinnen A., Hörz W. Removal of positioned nucleosomes from the yeast PHO5 promoter upon PHO5 induction releases additional upstream activating DNA elements. EMBO J. 1986 Oct;5(10):2689–2696. doi: 10.1002/j.1460-2075.1986.tb04552.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Andreadis A., Hsu Y. P., Hermodson M., Kohlhaw G., Schimmel P. Yeast LEU2. Repression of mRNA levels by leucine and primary structure of the gene product. J Biol Chem. 1984 Jul 10;259(13):8059–8062. [PubMed] [Google Scholar]
  4. Brent R., Ptashne M. A bacterial repressor protein or a yeast transcriptional terminator can block upstream activation of a yeast gene. Nature. 1984 Dec 13;312(5995):612–615. doi: 10.1038/312612a0. [DOI] [PubMed] [Google Scholar]
  5. Cordingley M. G., Riegel A. T., Hager G. L. Steroid-dependent interaction of transcription factors with the inducible promoter of mouse mammary tumor virus in vivo. Cell. 1987 Jan 30;48(2):261–270. doi: 10.1016/0092-8674(87)90429-6. [DOI] [PubMed] [Google Scholar]
  6. Eissenberg J. C., Cartwright I. L., Thomas G. H., Elgin S. C. Selected topics in chromatin structure. Annu Rev Genet. 1985;19:485–536. doi: 10.1146/annurev.ge.19.120185.002413. [DOI] [PubMed] [Google Scholar]
  7. Elgin S. C. The formation and function of DNase I hypersensitive sites in the process of gene activation. J Biol Chem. 1988 Dec 25;263(36):19259–19262. [PubMed] [Google Scholar]
  8. Fascher K. D., Schmitz J., Hörz W. Role of trans-activating proteins in the generation of active chromatin at the PHO5 promoter in S. cerevisiae. EMBO J. 1990 Aug;9(8):2523–2528. doi: 10.1002/j.1460-2075.1990.tb07432.x. [DOI] [PMC free article] [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. Gross D. S., Garrard W. T. Nuclease hypersensitive sites in chromatin. Annu Rev Biochem. 1988;57:159–197. doi: 10.1146/annurev.bi.57.070188.001111. [DOI] [PubMed] [Google Scholar]
  11. Guarente L. UASs and enhancers: common mechanism of transcriptional activation in yeast and mammals. Cell. 1988 Feb 12;52(3):303–305. doi: 10.1016/s0092-8674(88)80020-5. [DOI] [PubMed] [Google Scholar]
  12. Guarente L. Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. Methods Enzymol. 1983;101:181–191. doi: 10.1016/0076-6879(83)01013-7. [DOI] [PubMed] [Google Scholar]
  13. Han M., Grunstein M. Nucleosome loss activates yeast downstream promoters in vivo. Cell. 1988 Dec 23;55(6):1137–1145. doi: 10.1016/0092-8674(88)90258-9. [DOI] [PubMed] [Google Scholar]
  14. Han M., Kim U. J., Kayne P., Grunstein M. Depletion of histone H4 and nucleosomes activates the PHO5 gene in Saccharomyces cerevisiae. EMBO J. 1988 Jul;7(7):2221–2228. doi: 10.1002/j.1460-2075.1988.tb03061.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kayne P. S., Kim U. J., Han M., Mullen J. R., Yoshizaki F., Grunstein M. Extremely conserved histone H4 N terminus is dispensable for growth but essential for repressing the silent mating loci in yeast. Cell. 1988 Oct 7;55(1):27–39. doi: 10.1016/0092-8674(88)90006-2. [DOI] [PubMed] [Google Scholar]
  16. Knezetic J. A., Luse D. S. The presence of nucleosomes on a DNA template prevents initiation by RNA polymerase II in vitro. Cell. 1986 Apr 11;45(1):95–104. doi: 10.1016/0092-8674(86)90541-6. [DOI] [PubMed] [Google Scholar]
  17. Legrain M., De Wilde M., Hilger F. Isolation, physical characterization and expression analysis of the Saccharomyces cerevisiae positive regulatory gene PHO4. Nucleic Acids Res. 1986 Apr 11;14(7):3059–3073. doi: 10.1093/nar/14.7.3059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lorch Y., LaPointe J. W., Kornberg R. D. Nucleosomes inhibit the initiation of transcription but allow chain elongation with the displacement of histones. Cell. 1987 Apr 24;49(2):203–210. doi: 10.1016/0092-8674(87)90561-7. [DOI] [PubMed] [Google Scholar]
  19. Meyhack B., Bajwa W., Rudolph H., Hinnen A. Two yeast acid phosphatase structural genes are the result of a tandem duplication and show different degrees of homology in their promoter and coding sequences. EMBO J. 1982;1(6):675–680. doi: 10.1002/j.1460-2075.1982.tb01229.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Neubauer B., Linxweiler W., Hörz W. DNA engineering shows that nucleosome phasing on the African green monkey alpha-satellite is the result of multiple additive histone-DNA interactions. J Mol Biol. 1986 Aug 20;190(4):639–645. doi: 10.1016/0022-2836(86)90249-4. [DOI] [PubMed] [Google Scholar]
  22. Panzeri L., Philippsen P. Centromeric DNA from chromosome VI in Saccharomyces cerevisiae strains. EMBO J. 1982;1(12):1605–1611. doi: 10.1002/j.1460-2075.1982.tb01362.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Pavlović B., Hörz W. The chromatin structure at the promoter of a glyceraldehyde phosphate dehydrogenase gene from Saccharomyces cerevisiae reflects its functional state. Mol Cell Biol. 1988 Dec;8(12):5513–5520. doi: 10.1128/mcb.8.12.5513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Pederson D. S., Thoma F., Simpson R. T. Core particle, fiber, and transcriptionally active chromatin structure. Annu Rev Cell Biol. 1986;2:117–147. doi: 10.1146/annurev.cb.02.110186.001001. [DOI] [PubMed] [Google Scholar]
  25. Perlmann T., Wrange O. Specific glucocorticoid receptor binding to DNA reconstituted in a nucleosome. EMBO J. 1988 Oct;7(10):3073–3079. doi: 10.1002/j.1460-2075.1988.tb03172.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Piña B., Brüggemeier U., Beato M. Nucleosome positioning modulates accessibility of regulatory proteins to the mouse mammary tumor virus promoter. Cell. 1990 Mar 9;60(5):719–731. doi: 10.1016/0092-8674(90)90087-u. [DOI] [PubMed] [Google Scholar]
  27. Richard-Foy H., Hager G. L. Sequence-specific positioning of nucleosomes over the steroid-inducible MMTV promoter. EMBO J. 1987 Aug;6(8):2321–2328. doi: 10.1002/j.1460-2075.1987.tb02507.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Rosenberg H., Singer M., Rosenberg M. Highly reiterated sequences of SIMIANSIMIANSIMIANSIMIANSIMIAN. Science. 1978 Apr 28;200(4340):394–402. doi: 10.1126/science.205944. [DOI] [PubMed] [Google Scholar]
  29. Rudolph H., Hinnen A. The yeast PHO5 promoter: phosphate-control elements and sequences mediating mRNA start-site selection. Proc Natl Acad Sci U S A. 1987 Mar;84(5):1340–1344. doi: 10.1073/pnas.84.5.1340. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sengstag C., Hinnen A. A 28-bp segment of the Saccharomyces cerevisiae PHO5 upstream activator sequence confers phosphate control to the CYC1-lacZ gene fusion. Gene. 1988 Jul 30;67(2):223–228. doi: 10.1016/0378-1119(88)90399-x. [DOI] [PubMed] [Google Scholar]
  31. Simpson R. T. Nucleosome positioning can affect the function of a cis-acting DNA element in vivo. Nature. 1990 Jan 25;343(6256):387–389. doi: 10.1038/343387a0. [DOI] [PubMed] [Google Scholar]
  32. Solomon M. J., Larsen P. L., Varshavsky A. Mapping protein-DNA interactions in vivo with formaldehyde: evidence that histone H4 is retained on a highly transcribed gene. Cell. 1988 Jun 17;53(6):937–947. doi: 10.1016/s0092-8674(88)90469-2. [DOI] [PubMed] [Google Scholar]
  33. Sutcliffe J. G. Complete nucleotide sequence of the Escherichia coli plasmid pBR322. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):77–90. doi: 10.1101/sqb.1979.043.01.013. [DOI] [PubMed] [Google Scholar]
  34. Tschumper G., Carbon J. Sequence of a yeast DNA fragment containing a chromosomal replicator and the TRP1 gene. Gene. 1980 Jul;10(2):157–166. doi: 10.1016/0378-1119(80)90133-x. [DOI] [PubMed] [Google Scholar]
  35. Vogel K., Hörz W., Hinnen A. The two positively acting regulatory proteins PHO2 and PHO4 physically interact with PHO5 upstream activation regions. Mol Cell Biol. 1989 May;9(5):2050–2057. doi: 10.1128/mcb.9.5.2050. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wintersberger U., Smith P., Letnansky K. Yeast chromatin. Preparation from isolated nuclei, histone composition and transcription capacity. Eur J Biochem. 1973 Feb 15;33(1):123–130. doi: 10.1111/j.1432-1033.1973.tb02663.x. [DOI] [PubMed] [Google Scholar]
  37. Workman J. L., Roeder R. G. Binding of transcription factor TFIID to the major late promoter during in vitro nucleosome assembly potentiates subsequent initiation by RNA polymerase II. Cell. 1987 Nov 20;51(4):613–622. doi: 10.1016/0092-8674(87)90130-9. [DOI] [PubMed] [Google Scholar]
  38. Yaniv M., Cereghini S. Structure of transcriptionally active chromatin. CRC Crit Rev Biochem. 1986;21(1):1–26. doi: 10.3109/10409238609113607. [DOI] [PubMed] [Google Scholar]
  39. Zhang X. Y., Fittler F., Hörz W. Eight different highly specific nucleosome phases on alpha-satellite DNA in the African green monkey. Nucleic Acids Res. 1983 Jul 11;11(13):4287–4306. doi: 10.1093/nar/11.13.4287. [DOI] [PMC free article] [PubMed] [Google Scholar]

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