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. 1996 Jun;16(6):3054–3065. doi: 10.1128/mcb.16.6.3054

Methylation of DNA repeats of decreasing sizes in Ascobolus immersus.

C Goyon 1, C Barry 1, A Grégoire 1, G Faugeron 1, J L Rossignol 1
PMCID: PMC231300  PMID: 8649417

Abstract

In Ascobolus immersus, DNA duplications are subject to the process of methylation induced premeiotically (MIP), which methylates the cytosine residues within the repeats and results in reversible gene silencing. The triggering of MIP requires pairing of the repeats, and its detection requires maintenance of the resulting methylation. MIP of kilobase-size duplications occurs frequently and leads to the methylation of all C residues in the repeats, including those belonging to non-CpG sequences. Using duplications of decreasing sizes, we observed that tandem repeats never escaped MIP when larger than 630 bp and showed a sudden and drastic drop in MIP frequencies when their sizes decreased from 630 to 317 bp. This contrasted with the progressive decrease of MIP frequencies observed with ectopic repeats, in which apparently the search for homology influences the MIP triggering efficiency. The minimal size actually required for a repeat to undergo detectable MIP was found to be close to 300 bp. Genomic sequencing and Southern hybridization analyses using restriction enzymes sensitive to C methylation showed a loss of methylation at non-CpG sites in short DNA segments, methylation being restricted to a limited number of CpG dinucleotides. Our data suggest the existence of two distinct mechanisms underlying methylation maintenance, one responsible for methylation at CpG sites and the other responsible for methylation at non-CpG sites.

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Selected References

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  1. Barry C., Faugeron G., Rossignol J. L. Methylation induced premeiotically in Ascobolus: coextension with DNA repeat lengths and effect on transcript elongation. Proc Natl Acad Sci U S A. 1993 May 15;90(10):4557–4561. doi: 10.1073/pnas.90.10.4557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bates G., Lehrach H. Trinucleotide repeat expansions and human genetic disease. Bioessays. 1994 Apr;16(4):277–284. doi: 10.1002/bies.950160411. [DOI] [PubMed] [Google Scholar]
  3. Browne M. J., Burdon R. H. The sequence specificity of vertebrate DNA methylation. Nucleic Acids Res. 1977 Apr;4(4):1025–1037. doi: 10.1093/nar/4.4.1025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clark S. J., Harrison J., Frommer M. CpNpG methylation in mammalian cells. Nat Genet. 1995 May;10(1):20–27. doi: 10.1038/ng0595-20. [DOI] [PubMed] [Google Scholar]
  5. Colot V., Rossignol J. L. Isolation of the Ascobolus immersus spore color gene b2 and study in single cells of gene silencing by methylation induced premeiotically. Genetics. 1995 Dec;141(4):1299–1314. doi: 10.1093/genetics/141.4.1299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. DOSKOCIL J., SORM F. Distribution of 5-methylcytosine in pyrimidine sequences of deoxyribonucleic acids. Biochim Biophys Acta. 1962 Jun 11;55:953–959. doi: 10.1016/0006-3002(62)90909-5. [DOI] [PubMed] [Google Scholar]
  7. Faugeron G., Goyon C., Grégoire A. Stable allele replacement and unstable non-homologous integration events during transformation of Ascobolus immersus. Gene. 1989 Mar 15;76(1):109–119. doi: 10.1016/0378-1119(89)90013-9. [DOI] [PubMed] [Google Scholar]
  8. Faugeron G., Rhounim L., Rossignol J. L. How does the cell count the number of ectopic copies of a gene in the premeiotic inactivation process acting in Ascobolus immersus? Genetics. 1990 Mar;124(3):585–591. doi: 10.1093/genetics/124.3.585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Frommer M., McDonald L. E., Millar D. S., Collis C. M., Watt F., Grigg G. W., Molloy P. L., Paul C. L. A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1827–1831. doi: 10.1073/pnas.89.5.1827. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Goyon C., Faugeron G., Rossignol J. L. Molecular cloning and characterization of the met2 gene from Ascobolus immersus. Gene. 1988 Mar 31;63(2):297–308. doi: 10.1016/0378-1119(88)90533-1. [DOI] [PubMed] [Google Scholar]
  11. Goyon C., Faugeron G. Targeted transformation of Ascobolus immersus and de novo methylation of the resulting duplicated DNA sequences. Mol Cell Biol. 1989 Jul;9(7):2818–2827. doi: 10.1128/mcb.9.7.2818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Goyon C., Nogueira T. I., Faugeron G. Perpetuation of cytosine methylation in Ascobolus immersus implies a novel type of maintenance methylase. J Mol Biol. 1994 Jul 1;240(1):42–51. doi: 10.1006/jmbi.1994.1416. [DOI] [PubMed] [Google Scholar]
  13. Gruenbaum Y., Naveh-Many T., Cedar H., Razin A. Sequence specificity of methylation in higher plant DNA. Nature. 1981 Aug 27;292(5826):860–862. doi: 10.1038/292860a0. [DOI] [PubMed] [Google Scholar]
  14. Holliday R., Pugh J. E. DNA modification mechanisms and gene activity during development. Science. 1975 Jan 24;187(4173):226–232. [PubMed] [Google Scholar]
  15. Hynes M. J., Corrick C. M., King J. A. Isolation of genomic clones containing the amdS gene of Aspergillus nidulans and their use in the analysis of structural and regulatory mutations. Mol Cell Biol. 1983 Aug;3(8):1430–1439. doi: 10.1128/mcb.3.8.1430. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Irelan J. T., Hagemann A. T., Selker E. U. High frequency repeat-induced point mutation (RIP) is not associated with efficient recombination in Neurospora. Genetics. 1994 Dec;138(4):1093–1103. doi: 10.1093/genetics/138.4.1093. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jinks-Robertson S., Michelitch M., Ramcharan S. Substrate length requirements for efficient mitotic recombination in Saccharomyces cerevisiae. Mol Cell Biol. 1993 Jul;13(7):3937–3950. doi: 10.1128/mcb.13.7.3937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Leonhardt H., Bestor T. H. Structure, function and regulation of mammalian DNA methyltransferase. EXS. 1993;64:109–119. doi: 10.1007/978-3-0348-9118-9_5. [DOI] [PubMed] [Google Scholar]
  19. Liskay R. M., Letsou A., Stachelek J. L. Homology requirement for efficient gene conversion between duplicated chromosomal sequences in mammalian cells. Genetics. 1987 Jan;115(1):161–167. doi: 10.1093/genetics/115.1.161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Meyer P., Niedenhof I., ten Lohuis M. Evidence for cytosine methylation of non-symmetrical sequences in transgenic Petunia hybrida. EMBO J. 1994 May 1;13(9):2084–2088. doi: 10.1002/j.1460-2075.1994.tb06483.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Nelson M., McClelland M. Site-specific methylation: effect on DNA modification methyltransferases and restriction endonucleases. Nucleic Acids Res. 1991 Apr 25;19 (Suppl):2045–2071. doi: 10.1093/nar/19.suppl.2045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Nicolas A., Petes T. D. Polarity of meiotic gene conversion in fungi: contrasting views. Experientia. 1994 Mar 15;50(3):242–252. doi: 10.1007/BF01924007. [DOI] [PubMed] [Google Scholar]
  23. RIZET G., ENGELMANN N., LEFORT C., LISSOUBA P., MOUSSEAU J. [On an Ascomycete of interest for the study of certain aspects of the problem of gene structure]. C R Hebd Seances Acad Sci. 1960 Mar 14;250:2050–2052. [PubMed] [Google Scholar]
  24. Rhounim L., Grégoire A., Salama S., Faugeron G. Clustering of multiple transgene integrations in highly-unstable Ascobolus immersus transformants. Curr Genet. 1994 Oct;26(4):344–351. doi: 10.1007/BF00310499. [DOI] [PubMed] [Google Scholar]
  25. Rhounim L., Rossignol J. L., Faugeron G. Epimutation of repeated genes in Ascobolus immersus. EMBO J. 1992 Dec;11(12):4451–4457. doi: 10.1002/j.1460-2075.1992.tb05546.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Riggs A. D. X inactivation, differentiation, and DNA methylation. Cytogenet Cell Genet. 1975;14(1):9–25. doi: 10.1159/000130315. [DOI] [PubMed] [Google Scholar]
  27. Rossignol J. L., Faugeron G. Gene inactivation triggered by recognition between DNA repeats. Experientia. 1994 Mar 15;50(3):307–317. doi: 10.1007/BF01924014. [DOI] [PubMed] [Google Scholar]
  28. Selker E. U. DNA methylation and chromatin structure: a view from below. Trends Biochem Sci. 1990 Mar;15(3):103–107. doi: 10.1016/0968-0004(90)90193-f. [DOI] [PubMed] [Google Scholar]
  29. Selker E. U., Fritz D. Y., Singer M. J. Dense nonsymmetrical DNA methylation resulting from repeat-induced point mutation in Neurospora. Science. 1993 Dec 10;262(5140):1724–1728. doi: 10.1126/science.8259516. [DOI] [PubMed] [Google Scholar]
  30. Weiner B. M., Kleckner N. Chromosome pairing via multiple interstitial interactions before and during meiosis in yeast. Cell. 1994 Jul 1;77(7):977–991. doi: 10.1016/0092-8674(94)90438-3. [DOI] [PubMed] [Google Scholar]

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