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Association between up-regulation of stress-responsive genes and hypomethylation of genomic DNA in tobacco plants

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Abstract

Transcripts that specifically accumulate in transgenic tobacco plants expressing an anti-sense construct for a tobacco type I DNA methyltransferase, NtMET1, were screened by the differential display method. Of the 31 genes identified, 16 encoded proteins with known functions; ten of these were related to biotic and abiotic stress responses, and the other six to cellular functions. In order to examine whether expression of these genes is correlated with DNA methylation status under natural stress conditions, a pathogen-responsive gene ( NtAlix1) was selected as representative, and assayed for transcript induction and genomic methylation in tobacco plants infected with tobacco mosaic virus (TMV). In inoculated leaves of wild-type plants, NtAlix1 transcripts began to accumulate 12 h after the onset of the hypersensitive response (HR), and levels remained high for up to 24 h. Changes in the methylation status at the locus became obvious 24 h later, as detected by digestion of genomic DNA with a methylation-sensitive restriction enzyme. The results suggest that the level of DNA methylation may change in response to external stresses, and that this is closely related to the activation of stress-responsive genes.

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References

  1. Bird AP (2002) DNA methylation patterns and epigenetic memory. Genes Dev 16:6-21

  2. Cao X, Jacobsen S (2002) Role of the Arabidopsis DRM methyltransferases in de novo DNA methylation and gene silencing. Curr BioL 12: 1138–1144

  3. Chatellard-Causse C, Blot B, Cristina N, Torch S, Missotten M, Sadoul R (2002) Alix (ALG-2-interacting protein X), a protein involved in apoptosis, binds to endophilins and induces cytoplasmic vacuolization. J Biol Chem 277:29108–29115

  4. Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156–159

  5. Ehrlich M (2000) DNA hypomethylation and cancer. In: Ehrlich M (ed) DNA alterations in cancer. Eaton Publishing, Natick, Mass., pp 273–291

  6. Fajkus J, Vyskot B, Bezdek M (1992) Changes in chromatin structure due to hypomethylation induced with 5-azacytidine or DL-ethionine. FEBS Lett 314:13–16

  7. Finnegan EJ, Kovac KA (2000) Plant DNA methyltransferases. Plant Mol Biol 43:189–201

  8. Finnegan EJ, Peacock WJ, Dennis ES (1996) Reduced DNA methylation in Arabidopsis thaliana results in abnormal plant development. Proc Natl Acad Sci USA 93:8449–8454

  9. Finnegan EJ, Genger RK, Peacock WJ, Dennis ES (1998) DNA methylation in plants. Annu Rev Plant Physiol Plant Mol Biol 49:223–247

  10. Ferguson-Smith AC, Surani MA (2001) Imprinting and the epigenetic asymmetry between parental genomes. Science 293:1086–1089

  11. Gong Z, Morales-Ruiz T, Ariza RR, Roldan-Arjona T, David L, Zhu JK (2002) ROS1, a repressor of transcriptional gene silencing in Arabidopsis, encodes a DNA glycosylase/lyase. Cell 111:803–814

  12. Hara K, Yagi M, Koizumi N, Kusano T, Sano H (2000) Screening of wound-responsive genes identifies an immediate-early expressed gene encoding a highly charged protein in tobacco plants. Plant Cell Physiol 41:684–691

  13. Jackson-Grusby L, Beard C, Possemato R, Tudor M, Fambrough D, Csankovszki G, Dausman J, Lee P, Wilson C, Lander E, Jaenisch R (2001) Loss of genomic methylation causes p53-dependent apoptosis and epigenetic deregulation. Nature Genet 27:31–39

  14. Kankel MW, Ramsey DE, Stokes TL, Flowers SK, Haag JR, Jeddeloh JA, Riddle NC, Verbsky ML, Richards EJ (2003) Arabidopsis MET1 cytosine methyltransferase mutants. Genetics 163:1109–1122

  15. Kakutani T, Munakata K, Richards EJ, Hirochika H (1999) Meiotically and mitotically stable inheritance of DNA hypomethylation induced by ddm1 mutation of Arabidopsis thaliana. Genetics 151:831–838

  16. Kovarik A, Koukalova B, Holy A, Bezdek M (1994) Sequence-specific hypomethylation of the tobacco genome induced with dihydroxypropyladenine, ethinine and 5-azacytidine. FEBS Lett 353:309–311

  17. Lindroth AM, Cao X, Jackson JP, Zilberman D, McCallum CM, Henikoff S, Jacobsen SE (2001) Requirement of CHROMOMETHYLASE3 for maintenance of CpXpG methylation. Science 292:2077–2080

  18. Miura A, Yonebayashi S, Watanabe K, Toyama T, Shimada H, Kakutani T (2000) Mobilization of transposons by a mutation abolishing DNA methylation in Arabidopsis. Nature 411:212–214

  19. Murray MG, Thompson WF (1980) Rapid isolation of high molecular weight plant DNA. Nucleic Acids Res 8:4321–4325

  20. Nakano Y, Steward N, Kusano T, Sano H (2000) A tobacco NtMET1 encoding a DNA methyltransferase: molecular characterization and abnormal phenotypes of antisense transgenic tobacco plants. Plant Cell Physiol 41:448–457

  21. Ronemus MJ, Galbiati M, Ticker C, Chen J, Dellaporta SL (1996) Demethylation-induced developmental pleiotropy in Arabidopsis. Science 273:654–657

  22. Saze H, Mittelsten Scheid O, Paszkowski J (2003) Maintenance of CpG methylation is essential for epigenetic inheritance during plant gametogenesis. Nature Genet 34:65–69

  23. Steward N, Ito M, Yamaguchi Y, Koizumi N, Sano H (2002) Periodic DNA methylation in maize nucleosomes and demethylation by environmental stress. J Biol Chem 277:37741–37746

  24. Thomashow MF (1998) Role of cold-responsive genes in plant freezing tolerance. Plant Physiol 118:1–8

  25. Wada Y, Ohya H, Yamaguchi Y, Koizumi N, Sano H (2003) Preferential de novo methylation of cytosine residues in non-CpG sequences by a domains rearranged DNA methyltransferase from tobacco plants. J Biol Chem 278:42386–42393

  26. Yoda H, Ogawa M, Yamaguchi Y, Koizumi N, Sano H (2002) Identification of early responsive genes associated with the hypersensitive response and properties of a WRKY-type transcription factor in tobacco plants upon tobacco mosaic virus infection. Mol Genet Genomics 267:154–161

  27. Yoshida KT, Naito S, Takeda G (1994) cDNA cloning of regeneration-specific genes in rice by differential screening of randomly amplified cDNA using RAPD primers. Plant Cell Physiol 35:1003–1009

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Acknowledgements

The authors are grateful to Dr. M. Bezdek (Academy of Sciences of the Czech Republic) for providing the plasmid HRS60.1, Drs N. Koizumi and Y. Yamaguchi (Nara Institute of Science and Technology) for valuable discussion and suggestions, to Ms M. Kobayashi (Nara Institute of Science and Technology) for technical assistance and to Dr M. Moore (Intermal, Nagoya) for critical reading of the manuscript. This work was supported by a grant from the Research for the Future Program (JSPS-RTFT001604) of the Japan Society for the Promotion of Science

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Correspondence to H. Sano.

Additional information

Communicated by G. Jürgens

The first two authors contributed equally to this work

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Wada, Y., Miyamoto, K., Kusano, T. et al. Association between up-regulation of stress-responsive genes and hypomethylation of genomic DNA in tobacco plants. Mol Genet Genomics 271, 658–666 (2004). https://doi.org/10.1007/s00438-004-1018-4

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Keywords

  • 5-Methylcytosine
  • DNA methyltransferase
  • Tobacco mosaic virus
  • Hypersensitive response
  • Nicotiana tabacum.