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. 2002 Sep;162(1):59–71. doi: 10.1093/genetics/162.1.59

A novel yeast silencer. the 2mu origin of Saccharomyces cerevisiae has HST3-, MIG1- and SIR-dependent silencing activity.

Arnold Grünweller 1, Ann E Ehrenhofer-Murray 1
PMCID: PMC1462261  PMID: 12242223

Abstract

Silencing in Saccharomyces cerevisiae is found at the mating-type loci HMR and HML, in subtelomeric regions, and at the rDNA locus. Repressed chromatin is built up by the recruitment of the Sir proteins via their interaction with DNA-binding proteins that bind to silencers. Here, we have performed a genetic screen for novel sequence elements within the yeast genome that display silencing activity. We isolated as a novel silencer element the origin of replication from the endogenous 2mu plasmid (2mu ARS). 2mu ARS-mediated silencing was dependent upon the Sir proteins, the origin recognition complex (ORC), and Hst3, a Sir2 histone deacetylase homolog, suggesting that it constituted a novel class of silencing in yeast. Moreover, 2mu ARS carried a binding site for Mig1, a transcriptional repressor of glucose-regulated genes. Both the Mig1-binding site and the MIG1 gene were necessary for full silencing activity of 2mu ARS. Furthermore, Hst3 was physically present at 2mu ARS in a silencing context as well as at the endogenous 2mu plasmid. Also, Hst3 regulated the repression of the flipase gene, although this was likely an indirect effect of HST3 on FLP1 expression.

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Selected References

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  1. Andrulis E. D., Neiman A. M., Zappulla D. C., Sternglanz R. Perinuclear localization of chromatin facilitates transcriptional silencing. Nature. 1998 Aug 6;394(6693):592–595. doi: 10.1038/29100. [DOI] [PubMed] [Google Scholar]
  2. Aparicio O. M., Billington B. L., Gottschling D. E. Modifiers of position effect are shared between telomeric and silent mating-type loci in S. cerevisiae. Cell. 1991 Sep 20;66(6):1279–1287. doi: 10.1016/0092-8674(91)90049-5. [DOI] [PubMed] [Google Scholar]
  3. Aparicio O. M., Gottschling D. E. Overcoming telomeric silencing: a trans-activator competes to establish gene expression in a cell cycle-dependent way. Genes Dev. 1994 May 15;8(10):1133–1146. doi: 10.1101/gad.8.10.1133. [DOI] [PubMed] [Google Scholar]
  4. Boeke J. D., LaCroute F., Fink G. R. A positive selection for mutants lacking orotidine-5'-phosphate decarboxylase activity in yeast: 5-fluoro-orotic acid resistance. Mol Gen Genet. 1984;197(2):345–346. doi: 10.1007/BF00330984. [DOI] [PubMed] [Google Scholar]
  5. Bone J. R., Roth S. Y. Recruitment of the yeast Tup1p-Ssn6p repressor is associated with localized decreases in histone acetylation. J Biol Chem. 2000 Oct 30;276(3):1808–1813. doi: 10.1074/jbc.M008668200. [DOI] [PubMed] [Google Scholar]
  6. Brachmann C. B., Sherman J. M., Devine S. E., Cameron E. E., Pillus L., Boeke J. D. The SIR2 gene family, conserved from bacteria to humans, functions in silencing, cell cycle progression, and chromosome stability. Genes Dev. 1995 Dec 1;9(23):2888–2902. doi: 10.1101/gad.9.23.2888. [DOI] [PubMed] [Google Scholar]
  7. Braunstein M., Rose A. B., Holmes S. G., Allis C. D., Broach J. R. Transcriptional silencing in yeast is associated with reduced nucleosome acetylation. Genes Dev. 1993 Apr;7(4):592–604. doi: 10.1101/gad.7.4.592. [DOI] [PubMed] [Google Scholar]
  8. Dubey D. D., Davis L. R., Greenfeder S. A., Ong L. Y., Zhu J. G., Broach J. R., Newlon C. S., Huberman J. A. Evidence suggesting that the ARS elements associated with silencers of the yeast mating-type locus HML do not function as chromosomal DNA replication origins. Mol Cell Biol. 1991 Oct;11(10):5346–5355. doi: 10.1128/mcb.11.10.5346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ehrenhofer-Murray A. E., Gossen M., Pak D. T., Botchan M. R., Rine J. Separation of origin recognition complex functions by cross-species complementation. Science. 1995 Dec 8;270(5242):1671–1674. doi: 10.1126/science.270.5242.1671. [DOI] [PubMed] [Google Scholar]
  10. Fangman W. L., Brewer B. J. Activation of replication origins within yeast chromosomes. Annu Rev Cell Biol. 1991;7:375–402. doi: 10.1146/annurev.cb.07.110191.002111. [DOI] [PubMed] [Google Scholar]
  11. Fourel G., Revardel E., Koering C. E., Gilson E. Cohabitation of insulators and silencing elements in yeast subtelomeric regions. EMBO J. 1999 May 4;18(9):2522–2537. doi: 10.1093/emboj/18.9.2522. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fox C. A., Ehrenhofer-Murray A. E., Loo S., Rine J. The origin recognition complex, SIR1, and the S phase requirement for silencing. Science. 1997 Jun 6;276(5318):1547–1551. doi: 10.1126/science.276.5318.1547. [DOI] [PubMed] [Google Scholar]
  13. Fox C. A., Loo S., Dillin A., Rine J. The origin recognition complex has essential functions in transcriptional silencing and chromosomal replication. Genes Dev. 1995 Apr 15;9(8):911–924. doi: 10.1101/gad.9.8.911. [DOI] [PubMed] [Google Scholar]
  14. Friedman K. L., Diller J. D., Ferguson B. M., Nyland S. V., Brewer B. J., Fangman W. L. Multiple determinants controlling activation of yeast replication origins late in S phase. Genes Dev. 1996 Jul 1;10(13):1595–1607. doi: 10.1101/gad.10.13.1595. [DOI] [PubMed] [Google Scholar]
  15. Friedman K. L., Raghuraman M. K., Fangman W. L., Brewer B. J. Analysis of the temporal program of replication initiation in yeast chromosomes. J Cell Sci Suppl. 1995;19:51–58. doi: 10.1242/jcs.1995.supplement_19.7. [DOI] [PubMed] [Google Scholar]
  16. Gardner K. A., Rine J., Fox C. A. A region of the Sir1 protein dedicated to recognition of a silencer and required for interaction with the Orc1 protein in saccharomyces cerevisiae. Genetics. 1999 Jan;151(1):31–44. doi: 10.1093/genetics/151.1.31. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hecht A., Strahl-Bolsinger S., Grunstein M. Spreading of transcriptional repressor SIR3 from telomeric heterochromatin. Nature. 1996 Sep 5;383(6595):92–96. doi: 10.1038/383092a0. [DOI] [PubMed] [Google Scholar]
  18. Huberman J. A., Spotila L. D., Nawotka K. A., el-Assouli S. M., Davis L. R. The in vivo replication origin of the yeast 2 microns plasmid. Cell. 1987 Nov 6;51(3):473–481. doi: 10.1016/0092-8674(87)90643-x. [DOI] [PubMed] [Google Scholar]
  19. Imai S., Armstrong C. M., Kaeberlein M., Guarente L. Transcriptional silencing and longevity protein Sir2 is an NAD-dependent histone deacetylase. Nature. 2000 Feb 17;403(6771):795–800. doi: 10.1038/35001622. [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. Landry J., Sutton A., Tafrov S. T., Heller R. C., Stebbins J., Pillus L., Sternglanz R. The silencing protein SIR2 and its homologs are NAD-dependent protein deacetylases. Proc Natl Acad Sci U S A. 2000 May 23;97(11):5807–5811. doi: 10.1073/pnas.110148297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Loo S., Rine J. Silencers and domains of generalized repression. Science. 1994 Jun 17;264(5166):1768–1771. doi: 10.1126/science.8209257. [DOI] [PubMed] [Google Scholar]
  23. Marahrens Y., Stillman B. A yeast chromosomal origin of DNA replication defined by multiple functional elements. Science. 1992 Feb 14;255(5046):817–823. doi: 10.1126/science.1536007. [DOI] [PubMed] [Google Scholar]
  24. McNally F. J., Rine J. A synthetic silencer mediates SIR-dependent functions in Saccharomyces cerevisiae. Mol Cell Biol. 1991 Nov;11(11):5648–5659. doi: 10.1128/mcb.11.11.5648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Nehlin J. O., Carlberg M., Ronne H. Control of yeast GAL genes by MIG1 repressor: a transcriptional cascade in the glucose response. EMBO J. 1991 Nov;10(11):3373–3377. doi: 10.1002/j.1460-2075.1991.tb04901.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nielsen A. L., Oulad-Abdelghani M., Ortiz J. A., Remboutsika E., Chambon P., Losson R. Heterochromatin formation in mammalian cells: interaction between histones and HP1 proteins. Mol Cell. 2001 Apr;7(4):729–739. doi: 10.1016/s1097-2765(01)00218-0. [DOI] [PubMed] [Google Scholar]
  27. Ostling J., Ronne H. Negative control of the Mig1p repressor by Snf1p-dependent phosphorylation in the absence of glucose. Eur J Biochem. 1998 Feb 15;252(1):162–168. doi: 10.1046/j.1432-1327.1998.2520162.x. [DOI] [PubMed] [Google Scholar]
  28. Pak D. T., Pflumm M., Chesnokov I., Huang D. W., Kellum R., Marr J., Romanowski P., Botchan M. R. Association of the origin recognition complex with heterochromatin and HP1 in higher eukaryotes. Cell. 1997 Oct 31;91(3):311–323. doi: 10.1016/s0092-8674(00)80415-8. [DOI] [PubMed] [Google Scholar]
  29. Palacios DeBeer M. A., Fox C. A. A role for a replicator dominance mechanism in silencing. EMBO J. 1999 Jul 1;18(13):3808–3819. doi: 10.1093/emboj/18.13.3808. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Pryde F. E., Louis E. J. Limitations of silencing at native yeast telomeres. EMBO J. 1999 May 4;18(9):2538–2550. doi: 10.1093/emboj/18.9.2538. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Quandt K., Frech K., Karas H., Wingender E., Werner T. MatInd and MatInspector: new fast and versatile tools for detection of consensus matches in nucleotide sequence data. Nucleic Acids Res. 1995 Dec 11;23(23):4878–4884. doi: 10.1093/nar/23.23.4878. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Rao H., Marahrens Y., Stillman B. Functional conservation of multiple elements in yeast chromosomal replicators. Mol Cell Biol. 1994 Nov;14(11):7643–7651. doi: 10.1128/mcb.14.11.7643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Reynolds A. E., McCarroll R. M., Newlon C. S., Fangman W. L. Time of replication of ARS elements along yeast chromosome III. Mol Cell Biol. 1989 Oct;9(10):4488–4494. doi: 10.1128/mcb.9.10.4488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rivier D. H., Ekena J. L., Rine J. HMR-I is an origin of replication and a silencer in Saccharomyces cerevisiae. Genetics. 1999 Feb;151(2):521–529. doi: 10.1093/genetics/151.2.521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Rivier D. H., Rine J. An origin of DNA replication and a transcription silencer require a common element. Science. 1992 May 1;256(5057):659–663. doi: 10.1126/science.1585179. [DOI] [PubMed] [Google Scholar]
  36. Roy A., Exinger F., Losson R. cis- and trans-acting regulatory elements of the yeast URA3 promoter. Mol Cell Biol. 1990 Oct;10(10):5257–5270. doi: 10.1128/mcb.10.10.5257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Rusché L. N., Rine J. Conversion of a gene-specific repressor to a regional silencer. Genes Dev. 2001 Apr 15;15(8):955–967. doi: 10.1101/gad.873601. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Sherman F. Getting started with yeast. Methods Enzymol. 1991;194:3–21. doi: 10.1016/0076-6879(91)94004-v. [DOI] [PubMed] [Google Scholar]
  39. Sherman J. M., Stone E. M., Freeman-Cook L. L., Brachmann C. B., Boeke J. D., Pillus L. The conserved core of a human SIR2 homologue functions in yeast silencing. Mol Biol Cell. 1999 Sep;10(9):3045–3059. doi: 10.1091/mbc.10.9.3045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. 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]
  41. Singh J., Klar A. J. Active genes in budding yeast display enhanced in vivo accessibility to foreign DNA methylases: a novel in vivo probe for chromatin structure of yeast. Genes Dev. 1992 Feb;6(2):186–196. doi: 10.1101/gad.6.2.186. [DOI] [PubMed] [Google Scholar]
  42. Smith J. S., Boeke J. D. An unusual form of transcriptional silencing in yeast ribosomal DNA. Genes Dev. 1997 Jan 15;11(2):241–254. doi: 10.1101/gad.11.2.241. [DOI] [PubMed] [Google Scholar]
  43. Smith J. S., Brachmann C. B., Celic I., Kenna M. A., Muhammad S., Starai V. J., Avalos J. L., Escalante-Semerena J. C., Grubmeyer C., Wolberger C. A phylogenetically conserved NAD+-dependent protein deacetylase activity in the Sir2 protein family. Proc Natl Acad Sci U S A. 2000 Jun 6;97(12):6658–6663. doi: 10.1073/pnas.97.12.6658. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Som T., Armstrong K. A., Volkert F. C., Broach J. R. Autoregulation of 2 micron circle gene expression provides a model for maintenance of stable plasmid copy levels. Cell. 1988 Jan 15;52(1):27–37. doi: 10.1016/0092-8674(88)90528-4. [DOI] [PubMed] [Google Scholar]
  45. Stevenson J. B., Gottschling D. E. Telomeric chromatin modulates replication timing near chromosome ends. Genes Dev. 1999 Jan 15;13(2):146–151. doi: 10.1101/gad.13.2.146. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Sutton A., Heller R. C., Landry J., Choy J. S., Sirko A., Sternglanz R. A novel form of transcriptional silencing by Sum1-1 requires Hst1 and the origin recognition complex. Mol Cell Biol. 2001 May;21(10):3514–3522. doi: 10.1128/MCB.21.10.3514-3522.2001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Tanny J. C., Moazed D. Coupling of histone deacetylation to NAD breakdown by the yeast silencing protein Sir2: Evidence for acetyl transfer from substrate to an NAD breakdown product. Proc Natl Acad Sci U S A. 2000 Dec 26;98(2):415–420. doi: 10.1073/pnas.031563798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Triolo T., Sternglanz R. Role of interactions between the origin recognition complex and SIR1 in transcriptional silencing. Nature. 1996 May 16;381(6579):251–253. doi: 10.1038/381251a0. [DOI] [PubMed] [Google Scholar]
  49. Uetz P., Giot L., Cagney G., Mansfield T. A., Judson R. S., Knight J. R., Lockshon D., Narayan V., Srinivasan M., Pochart P. A comprehensive analysis of protein-protein interactions in Saccharomyces cerevisiae. Nature. 2000 Feb 10;403(6770):623–627. doi: 10.1038/35001009. [DOI] [PubMed] [Google Scholar]
  50. Wach A., Brachat A., Pöhlmann R., Philippsen P. New heterologous modules for classical or PCR-based gene disruptions in Saccharomyces cerevisiae. Yeast. 1994 Dec;10(13):1793–1808. doi: 10.1002/yea.320101310. [DOI] [PubMed] [Google Scholar]
  51. Weiler K. S., Wakimoto B. T. Heterochromatin and gene expression in Drosophila. Annu Rev Genet. 1995;29:577–605. doi: 10.1146/annurev.ge.29.120195.003045. [DOI] [PubMed] [Google Scholar]
  52. Wu J., Suka N., Carlson M., Grunstein M. TUP1 utilizes histone H3/H2B-specific HDA1 deacetylase to repress gene activity in yeast. Mol Cell. 2001 Jan;7(1):117–126. doi: 10.1016/s1097-2765(01)00160-5. [DOI] [PubMed] [Google Scholar]
  53. Wyrick J. J., Holstege F. C., Jennings E. G., Causton H. C., Shore D., Grunstein M., Lander E. S., Young R. A. Chromosomal landscape of nucleosome-dependent gene expression and silencing in yeast. Nature. 1999 Nov 25;402(6760):418–421. doi: 10.1038/46567. [DOI] [PubMed] [Google Scholar]

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