Heterochromatin plays important roles in the structure, maintenance, and function of the eukaryotic genome. It is associated with special histone modifications and specialized non-histone proteins and assumes a more compact structure than euchromatin. Genes embedded in heterochromatin are generally transcriptionally silent. It was found previously that several mutations of proliferating cell nuclear antigen (PCNA), a DNA replication processivity factor, reduce transcriptional silencing at heterochromatin loci in Saccharomyces cerevisiae. However, the notion that PCNA plays a role in transcriptional silencing was recently questioned because of a potential problem concerning the silencing assays used in prior studies. To determine if PCNA is a bona fide contributor to heterochromatin-mediated transcriptional silencing, we examined the effects of PCNA mutations on heterochromatin structure. We found evidence implicating PCNA in the maintenance of the high-order structure and stability of heterochromatin, which indicates a role of DNA replication in heterochromatin maintenance.
Proliferating cell nuclear antigen (PCNA) heterochromatin high order structure heterochromatin stability DNA supercoiling
Proliferating cell nuclear antigen
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We thank Dr. Peter Burgers for providing us with pBL230 and pBL230-x plasmids encoding wild-type and mutant PCNA alleles and Dr. Zhiguo Zhang for yeast strains ZGY005, ZGY005-6, ZGY005-8, ZGY005-9, and ZGY005-79. We also thank Hiram Lyon for assistance in performing sucrose gradient assay. This work was supported by NSF grant MCB-1158008 to X.B.
Alani E, Cao L, Kleckner N (1987) A method for gene disruption that allows repeated use of URA3 selection in the construction of multiply disrupted yeast strains. Genetics 116:541–545CrossRefPubMedPubMedCentralGoogle Scholar
Ayyagari R, Impellizzeri KJ, Yoder BL, Gary SL, Burgers PM (1995) A mutational analysis of the yeast proliferating cell nuclear antigen indicates distinct roles in DNA replication and DNA repair. Mol Cell Biol 15:4420–4429CrossRefPubMedPubMedCentralGoogle Scholar
Bi X, Broach JR (1997) DNA in transcriptionally silent chromatin assumes a distinct topology that is sensitive to cell cycle progression. Mol Cell Biol 17:7077–7087CrossRefPubMedPubMedCentralGoogle Scholar
Bi X, Yu Q, Sandmeier JJ, Elizondo S (2004) Regulation of transcriptional silencing in yeast by growth temperature. J Mol Biol 344:893–905CrossRefPubMedGoogle Scholar
Cheng TH, Li YC, Gartenberg MR (1998) Persistence of an alternate chromatin structure at silenced loci in the absence of silencers. Proc Natl Acad Sci U S A 95:5521–5526CrossRefPubMedPubMedCentralGoogle Scholar
Ehrenhofer-Murray AE, Kamakaka RT, Rine J (1999) A role for the replication proteins PCNA, RF-C, polymerase epsilon and Cdc45 in transcriptional silencing in Saccharomyces cerevisiae. Genetics 153:1171–1182PubMedPubMedCentralGoogle Scholar
Eissenberg JC, Ayyagari R, Gomes XV, Burgers PM (1997) Mutations in yeast proliferating cell nuclear antigen define distinct sites for interaction with DNA polymerase delta and DNA polymerase epsilon. Mol Cell Biol 17:6367–6378CrossRefPubMedPubMedCentralGoogle Scholar
Hoege C, Pfander B, Moldovan GL, Pyrowolakis G, Jentsch S (2002) RAD6-dependent DNA repair is linked to modification of PCNA by ubiquitin and SUMO. Nature 419:135–141CrossRefPubMedGoogle Scholar
Krishna TS, Kong XP, Gary S, Burgers PM, Kuriyan J (1994) Crystal structure of the eukaryotic DNA polymerase processivity factor PCNA. Cell 79:1233–1243CrossRefPubMedGoogle Scholar
Kueng S, Oppikofer M, Gasser SM (2013) SIR proteins and the assembly of silent chromatin in budding yeast. Annu Rev Genet 47:275–306CrossRefPubMedGoogle Scholar
Miller A, Yang B, Foster T, Kirchmaier AL (2008) Proliferating cell nuclear antigen and ASF1 modulate silent chromatin in Saccharomyces cerevisiae via lysine 56 on histone H3. Genetics 179:793–809CrossRefPubMedPubMedCentralGoogle Scholar
Miller A, Chen J, Takasuka TE, Jacobi JL, Kaufman PD, Irudayaraj JM, Kirchmaier AL (2010) Proliferating cell nuclear antigen (PCNA) is required for cell cycle-regulated silent chromatin on replicated and nonreplicated genes. J Biol Chem 285:35142–35154CrossRefPubMedPubMedCentralGoogle Scholar
Moggs JG, Grandi P, Quivy JP, Jónsson ZO, Hübscher U, Becker PB, Almouzni G (2000) ACAF-1-PCNA-mediated chromatin assembly pathway triggered by sensing DNA damage. Mol Cell Biol 20:1206–1218CrossRefPubMedPubMedCentralGoogle Scholar
Rusche LN, Kirchmaier AL, Rine J (2003) The establishment, inheritance, and function of silenced chromatin in Saccharomyces cerevisiae. Annu Rev Biochem 72:481–516CrossRefPubMedGoogle Scholar
Sharp JA, Fouts ET, Krawitz DC, Kaufman PD (2001) Yeast histone deposition protein Asf1p requires Hir proteins and PCNA for heterochromatic silencing. Curr Biol 11:463–473CrossRefPubMedGoogle Scholar
Shibahara K, Stillman B (1999) Replication-dependent marking of DNA by PCNA facilitates CAF-1-coupled inheritance of chromatin. Cell 96:575–585CrossRefPubMedGoogle Scholar
Simpson RT, Thoma F, Brubaker JM (1985) Chromatin reconstituted from tandemly repeated cloned DNA fragments and core histones: a model system for study of higher order structure. Cell 42:799–808CrossRefPubMedGoogle Scholar