• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of geneticsGeneticsCurrent IssueInformation for AuthorsEditorial BoardSubscribeSubmit a Manuscript
Genetics. Jul 1998; 149(3): 1205–1219.
PMCID: PMC1460222

Distribution of a limited Sir2 protein pool regulates the strength of yeast rDNA silencing and is modulated by Sir4p.

Abstract

Transcriptional silencing in Saccharomyces cerevisiae occurs at the silent mating-type loci HML and HMR, at telomeres, and at the ribosomal DNA (rDNA) locus RDN1. Silencing in the rDNA occurs by a novel mechanism that depends on a single Silent Information Regulator (SIR) gene, SIR2. SIR4, essential for other silenced loci, paradoxically inhibits rDNA silencing. In this study, we elucidate a regulatory mechanism for rDNA silencing based on the finding that rDNA silencing strength directly correlates with cellular Sir2 protein levels. The endogenous level of Sir2p was shown to be limiting for rDNA silencing. Furthermore, small changes in Sir2p levels altered rDNA silencing strength. In rDNA silencing phenotypes, sir2 mutations were shown to be epistatic to sir4 mutations, indicating that SIR4 inhibition of rDNA silencing is mediated through SIR2. Furthermore, rDNA silencing is insensitive to SIR3 overexpression, but is severely reduced by overexpression of full-length Sir4p or a fragment of Sir4p that interacts with Sir2p. This negative effect of SIR4 overexpression was overridden by co-overexpression of SIR2, suggesting that SIR4 directly inhibits the rDNA silencing function of SIR2. Finally, genetic manipulations of SIR4 previously shown to promote extended life span also resulted in enhanced rDNA silencing. We propose a simple model in which telomeres act as regulators of rDNA silencing by competing for limiting amounts of Sir2 protein.

Full Text

The Full Text of this article is available as a PDF (546K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Adams AK, Holm C. Specific DNA replication mutations affect telomere length in Saccharomyces cerevisiae. Mol Cell Biol. 1996 Sep;16(9):4614–4620. [PMC free article] [PubMed]
  • Aparicio OM, Billington BL, Gottschling DE. Modifiers of position effect are shared between telomeric and silent mating-type loci in S. cerevisiae. Cell. 1991 Sep 20;66(6):1279–1287. [PubMed]
  • Baudin A, Ozier-Kalogeropoulos O, Denouel A, Lacroute F, Cullin C. A simple and efficient method for direct gene deletion in Saccharomyces cerevisiae. Nucleic Acids Res. 1993 Jul 11;21(14):3329–3330. [PMC free article] [PubMed]
  • Boeke JD, Garfinkel DJ, Styles CA, Fink GR. Ty elements transpose through an RNA intermediate. Cell. 1985 Mar;40(3):491–500. [PubMed]
  • Brachmann CB, Sherman JM, Devine SE, Cameron EE, Pillus L, Boeke JD. 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. [PubMed]
  • Brachmann CB, Davies A, Cost GJ, Caputo E, Li J, Hieter P, Boeke JD. Designer deletion strains derived from Saccharomyces cerevisiae S288C: a useful set of strains and plasmids for PCR-mediated gene disruption and other applications. Yeast. 1998 Jan 30;14(2):115–132. [PubMed]
  • Braunstein M, Rose AB, Holmes SG, Allis CD, Broach JR. Transcriptional silencing in yeast is associated with reduced nucleosome acetylation. Genes Dev. 1993 Apr;7(4):592–604. [PubMed]
  • Braunstein M, Sobel RE, Allis CD, Turner BM, Broach JR. Efficient transcriptional silencing in Saccharomyces cerevisiae requires a heterochromatin histone acetylation pattern. Mol Cell Biol. 1996 Aug;16(8):4349–4356. [PMC free article] [PubMed]
  • Bryk M, Banerjee M, Murphy M, Knudsen KE, Garfinkel DJ, Curcio MJ. Transcriptional silencing of Ty1 elements in the RDN1 locus of yeast. Genes Dev. 1997 Jan 15;11(2):255–269. [PubMed]
  • Buchman AR, Kimmerly WJ, Rine J, Kornberg RD. 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. [PMC free article] [PubMed]
  • Buck SW, Shore D. Action of a RAP1 carboxy-terminal silencing domain reveals an underlying competition between HMR and telomeres in yeast. Genes Dev. 1995 Feb 1;9(3):370–384. [PubMed]
  • Chapman KB, Boeke JD. Isolation and characterization of the gene encoding yeast debranching enzyme. Cell. 1991 May 3;65(3):483–492. [PubMed]
  • Chen XJ, Clark-Walker GD. sir2 mutants of Kluyveromyces lactis are hypersensitive to DNA-targeting drugs. Mol Cell Biol. 1994 Jul;14(7):4501–4508. [PMC free article] [PubMed]
  • Christianson TW, Sikorski RS, Dante M, Shero JH, Hieter P. Multifunctional yeast high-copy-number shuttle vectors. Gene. 1992 Jan 2;110(1):119–122. [PubMed]
  • Christman MF, Dietrich FS, Fink GR. Mitotic recombination in the rDNA of S. cerevisiae is suppressed by the combined action of DNA topoisomerases I and II. Cell. 1988 Nov 4;55(3):413–425. [PubMed]
  • Cockell M, Palladino F, Laroche T, Kyrion G, Liu C, Lustig AJ, Gasser SM. The carboxy termini of Sir4 and Rap1 affect Sir3 localization: evidence for a multicomponent complex required for yeast telomeric silencing. J Cell Biol. 1995 May;129(4):909–924. [PMC free article] [PubMed]
  • Cost GJ, Boeke JD. A useful colony colour phenotype associated with the yeast selectable/counter-selectable marker MET15. Yeast. 1996 Aug;12(10):939–941. [PubMed]
  • Curcio MJ, Garfinkel DJ. Single-step selection for Ty1 element retrotransposition. Proc Natl Acad Sci U S A. 1991 Feb 1;88(3):936–940. [PMC free article] [PubMed]
  • Derbyshire MK, Weinstock KG, Strathern JN. HST1, a new member of the SIR2 family of genes. Yeast. 1996 Jun 15;12(7):631–640. [PubMed]
  • Dobie K, Mehtali M, McClenaghan M, Lathe R. Variegated gene expression in mice. Trends Genet. 1997 Apr;13(4):127–130. [PubMed]
  • Fritze CE, Verschueren K, Strich R, Easton Esposito R. Direct evidence for SIR2 modulation of chromatin structure in yeast rDNA. EMBO J. 1997 Nov 3;16(21):6495–6509. [PMC free article] [PubMed]
  • Loo S, Rine J. Silencing and heritable domains of gene expression. Annu Rev Cell Dev Biol. 1995;11:519–548. [PubMed]
  • Lorenz MC, Muir RS, Lim E, McElver J, Weber SC, Heitman J. Gene disruption with PCR products in Saccharomyces cerevisiae. Gene. 1995 May 26;158(1):113–117. [PubMed]
  • Lustig AJ, Liu C, Zhang C, Hanish JP. Tethered Sir3p nucleates silencing at telomeres and internal loci in Saccharomyces cerevisiae. Mol Cell Biol. 1996 May;16(5):2483–2495. [PMC free article] [PubMed]
  • Gotta M, Laroche T, Formenton A, Maillet L, Scherthan H, Gasser SM. The clustering of telomeres and colocalization with Rap1, Sir3, and Sir4 proteins in wild-type Saccharomyces cerevisiae. J Cell Biol. 1996 Sep;134(6):1349–1363. [PMC free article] [PubMed]
  • Maillet L, Boscheron C, Gotta M, Marcand S, Gilson E, Gasser SM. Evidence for silencing compartments within the yeast nucleus: a role for telomere proximity and Sir protein concentration in silencer-mediated repression. Genes Dev. 1996 Jul 15;10(14):1796–1811. [PubMed]
  • Gotta M, Strahl-Bolsinger S, Renauld H, Laroche T, Kennedy BK, Grunstein M, Gasser SM. Localization of Sir2p: the nucleolus as a compartment for silent information regulators. EMBO J. 1997 Jun 2;16(11):3243–3255. [PMC free article] [PubMed]
  • Marcand S, Buck SW, Moretti P, Gilson E, Shore D. Silencing of genes at nontelomeric sites in yeast is controlled by sequestration of silencing factors at telomeres by Rap 1 protein. Genes Dev. 1996 Jun 1;10(11):1297–1309. [PubMed]
  • Gottlieb S, Esposito RE. A new role for a yeast transcriptional silencer gene, SIR2, in regulation of recombination in ribosomal DNA. Cell. 1989 Mar 10;56(5):771–776. [PubMed]
  • Marshall M, Mahoney D, Rose A, Hicks JB, Broach JR. Functional domains of SIR4, a gene required for position effect regulation in Saccharomyces cerevisiae. Mol Cell Biol. 1987 Dec;7(12):4441–4452. [PMC free article] [PubMed]
  • Moazed D, Johnson D. A deubiquitinating enzyme interacts with SIR4 and regulates silencing in S. cerevisiae. Cell. 1996 Aug 23;86(4):667–677. [PubMed]
  • Moazed D, Kistler A, Axelrod A, Rine J, Johnson AD. Silent information regulator protein complexes in Saccharomyces cerevisiae: a SIR2/SIR4 complex and evidence for a regulatory domain in SIR4 that inhibits its interaction with SIR3. Proc Natl Acad Sci U S A. 1997 Mar 18;94(6):2186–2191. [PMC free article] [PubMed]
  • Haber JE, George JP. A mutation that permits the expression of normally silent copies of mating-type information in Saccharomyces cerevisiae. Genetics. 1979 Sep;93(1):13–35. [PMC free article] [PubMed]
  • Moretti P, Freeman K, Coodly L, Shore D. Evidence that a complex of SIR proteins interacts with the silencer and telomere-binding protein RAP1. Genes Dev. 1994 Oct 1;8(19):2257–2269. [PubMed]
  • MORTIMER RK, JOHNSTON JR. Life span of individual yeast cells. Nature. 1959 Jun 20;183(4677):1751–1752. [PubMed]
  • Hecht A, Laroche T, Strahl-Bolsinger S, Gasser SM, Grunstein M. Histone H3 and H4 N-termini interact with SIR3 and SIR4 proteins: a molecular model for the formation of heterochromatin in yeast. Cell. 1995 Feb 24;80(4):583–592. [PubMed]
  • Palladino F, Laroche T, Gilson E, Axelrod A, Pillus L, Gasser SM. SIR3 and SIR4 proteins are required for the positioning and integrity of yeast telomeres. Cell. 1993 Nov 5;75(3):543–555. [PubMed]
  • Petes TD, Botstein D. Simple Mendelian inheritance of the reiterated ribosomal DNA of yeast. Proc Natl Acad Sci U S A. 1977 Nov;74(11):5091–5095. [PMC free article] [PubMed]
  • Hill JE, Myers AM, Koerner TJ, Tzagoloff A. Yeast/E. coli shuttle vectors with multiple unique restriction sites. Yeast. 1986 Sep;2(3):163–167. [PubMed]
  • Philippsen P, Thomas M, Kramer RA, Davis RW. Unique arrangement of coding sequences for 5 S, 5.8 S, 18 S and 25 S ribosomal RNA in Saccharomyces cerevisiae as determined by R-loop and hybridization analysis. J Mol Biol. 1978 Aug 15;123(3):387–404. [PubMed]
  • Holmes SG, Rose AB, Steuerle K, Saez E, Sayegh S, Lee YM, Broach JR. Hyperactivation of the silencing proteins, Sir2p and Sir3p, causes chromosome loss. Genetics. 1997 Mar;145(3):605–614. [PMC free article] [PubMed]
  • Huang H, Kahana A, Gottschling DE, Prakash L, Liebman SW. The ubiquitin-conjugating enzyme Rad6 (Ubc2) is required for silencing in Saccharomyces cerevisiae. Mol Cell Biol. 1997 Nov;17(11):6693–6699. [PMC free article] [PubMed]
  • Pillus L, Rine J. Epigenetic inheritance of transcriptional states in S. cerevisiae. Cell. 1989 Nov 17;59(4):637–647. [PubMed]
  • Huang H, Kahana A, Gottschling DE, Prakash L, Liebman SW. The ubiquitin-conjugating enzyme Rad6 (Ubc2) is required for silencing in Saccharomyces cerevisiae. Mol Cell Biol. 1997 Nov;17(11):6693–6699. [PMC free article] [PubMed]
  • Renauld H, Aparicio OM, Zierath PD, Billington BL, Chhablani SK, Gottschling DE. Silent domains are assembled continuously from the telomere and are defined by promoter distance and strength, and by SIR3 dosage. Genes Dev. 1993 Jul;7(7A):1133–1145. [PubMed]
  • Ivy JM, Klar AJ, Hicks JB. Cloning and characterization of four SIR genes of Saccharomyces cerevisiae. Mol Cell Biol. 1986 Feb;6(2):688–702. [PMC free article] [PubMed]
  • Rine J, Herskowitz I. Four genes responsible for a position effect on expression from HML and HMR in Saccharomyces cerevisiae. Genetics. 1987 May;116(1):9–22. [PMC free article] [PubMed]
  • Kennedy BK, Austriaco NR, Jr, Zhang J, Guarente L. Mutation in the silencing gene SIR4 can delay aging in S. cerevisiae. Cell. 1995 Feb 10;80(3):485–496. [PubMed]
  • Rine J, Strathern JN, Hicks JB, Herskowitz I. A suppressor of mating-type locus mutations in Saccharomyces cerevisiae: evidence for and identification of cryptic mating-type loci. Genetics. 1979 Dec;93(4):877–901. [PMC free article] [PubMed]
  • Kennedy BK, Gotta M, Sinclair DA, Mills K, McNabb DS, Murthy M, Pak SM, Laroche T, Gasser SM, Guarente L. Redistribution of silencing proteins from telomeres to the nucleolus is associated with extension of life span in S. cerevisiae. Cell. 1997 May 2;89(3):381–391. [PubMed]
  • Schnell R, Rine J. A position effect on the expression of a tRNA gene mediated by the SIR genes in Saccharomyces cerevisiae. Mol Cell Biol. 1986 Feb;6(2):494–501. [PMC free article] [PubMed]
  • Klar AJ, Fogel S, Macleod K. MAR1-a Regulator of the HMa and HMalpha Loci in SACCHAROMYCES CEREVISIAE. Genetics. 1979 Sep;93(1):37–50. [PMC free article] [PubMed]
  • Shore D. RAP1: a protean regulator in yeast. Trends Genet. 1994 Nov;10(11):408–412. [PubMed]
  • Kostriken R, Strathern JN, Klar AJ, Hicks JB, Heffron F. A site-specific endonuclease essential for mating-type switching in Saccharomyces cerevisiae. Cell. 1983 Nov;35(1):167–174. [PubMed]
  • Sikorski RS, 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. [PMC free article] [PubMed]
  • Liu H, Krizek J, Bretscher A. Construction of a GAL1-regulated yeast cDNA expression library and its application to the identification of genes whose overexpression causes lethality in yeast. Genetics. 1992 Nov;132(3):665–673. [PMC free article] [PubMed]
  • Sinclair DA, Guarente L. Extrachromosomal rDNA circles--a cause of aging in yeast. Cell. 1997 Dec 26;91(7):1033–1042. [PubMed]
  • Sinclair DA, Mills K, Guarente L. Accelerated aging and nucleolar fragmentation in yeast sgs1 mutants. Science. 1997 Aug 29;277(5330):1313–1316. [PubMed]
  • Loo S, Rine J. Silencers and domains of generalized repression. Science. 1994 Jun 17;264(5166):1768–1771. [PubMed]
  • Strahl-Bolsinger S, Hecht A, Luo K, Grunstein M. SIR2 and SIR4 interactions differ in core and extended telomeric heterochromatin in yeast. Genes Dev. 1997 Jan 1;11(1):83–93. [PubMed]
  • Strathern JN, Klar AJ, Hicks JB, Abraham JA, Ivy JM, Nasmyth KA, McGill C. Homothallic switching of yeast mating type cassettes is initiated by a double-stranded cut in the MAT locus. Cell. 1982 Nov;31(1):183–192. [PubMed]
  • Skryabin KG, Eldarov MA, Larionov VL, Bayev AA, Klootwijk J, de Regt VC, Veldman GM, Planta RJ, Georgiev OI, Hadjiolov AA. Structure and function of the nontranscribed spacer regions of yeast rDNA. Nucleic Acids Res. 1984 Mar 26;12(6):2955–2968. [PMC free article] [PubMed]
  • Sussel L, Vannier D, Shore D. Epigenetic switching of transcriptional states: cis- and trans-acting factors affecting establishment of silencing at the HMR locus in Saccharomyces cerevisiae. Mol Cell Biol. 1993 Jul;13(7):3919–3928. [PMC free article] [PubMed]
  • Smeal T, Claus J, Kennedy B, Cole F, Guarente L. Loss of transcriptional silencing causes sterility in old mother cells of S. cerevisiae. Cell. 1996 Feb 23;84(4):633–642. [PubMed]
  • Szostak JW, Wu R. Insertion of a genetic marker into the ribosomal DNA of yeast. Plasmid. 1979 Oct;2(4):536–554. [PubMed]
  • 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. [PMC free article] [PubMed]
  • Thompson JS, Hecht A, Grunstein M. Histones and the regulation of heterochromatin in yeast. Cold Spring Harb Symp Quant Biol. 1993;58:247–256. [PubMed]
  • Stavenhagen JB, Zakian VA. Internal tracts of telomeric DNA act as silencers in Saccharomyces cerevisiae. Genes Dev. 1994 Jun 15;8(12):1411–1422. [PubMed]
  • 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. [PubMed]
  • Sternglanz R. Histone acetylation: a gateway to transcriptional activation. Trends Biochem Sci. 1996 Oct;21(10):357–358. [PubMed]
  • Weiler KS, Wakimoto BT. Heterochromatin and gene expression in Drosophila. Annu Rev Genet. 1995;29:577–605. [PubMed]
  • Stone EM, Pillus L. Activation of an MAP kinase cascade leads to Sir3p hyperphosphorylation and strengthens transcriptional silencing. J Cell Biol. 1996 Nov;135(3):571–583. [PMC free article] [PubMed]
  • Yu CE, Oshima J, Fu YH, Wijsman EM, Hisama F, Alisch R, Matthews S, Nakura J, Miki T, Ouais S, et al. Positional cloning of the Werner's syndrome gene. Science. 1996 Apr 12;272(5259):258–262. [PubMed]
  • Stone EM, Swanson MJ, Romeo AM, Hicks JB, Sternglanz R. The SIR1 gene of Saccharomyces cerevisiae and its role as an extragenic suppressor of several mating-defective mutants. Mol Cell Biol. 1991 Apr;11(4):2253–2262. [PMC free article] [PubMed]

Articles from Genetics are provided here courtesy of Genetics Society of America

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...