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Plant and Cell Physiology Advance Access originally published online on February 21, 2007
Plant and Cell Physiology 2007 48(4):638-647; doi:10.1093/pcp/pcm028
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© The Author 2007. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oxfordjournals.org

Short Communication

Epigenetic Inactivation of Chalcone Synthase-A Transgene Transcription in Petunia Leads to a Reversion of the Post-Transcriptional Gene Silencing Phenotype

Akira Kanazawa1,*, Michael O'Dell2 and Roger P. Hellens3

1Graduate School of Agriculture, Hokkaido University, Sapporo, 060-8589 Japan
2Department of Genetics, John Innes Centre, Norwich Research Park, Colney, Norwich NR4 7UH, UK
3Gene Discovery and Function Department, HortResearch, Private Bag 92 169, Auckland, New Zealand

*Corresponding author: E-mail, kanazawa{at}res.agr.hokudai.ac.jp; Fax, +81-11-706-4933.


  Abstract

Petunia plants that exhibit a white-flowering phenotype as a consequence of chalcone synthase transgene-induced silencing occasionally give rise to revertant branches that produce flowers with wild-type pigmentation. Transcription run-on assays confirmed that the production of white flowers is caused by post-transcriptional gene silencing (PTGS), and indicated that transgene transcription is repressed in the revertant plants, providing evidence that induction of PTGS depends on the transcription rate. Transcriptional repression of the transgene was associated with cytosine methylation at CpG, CpNpG and CpNpN sites, and the expression was restored by treatment with either 5-azacytidine or trichostatin A. These results demonstrate that epigenetic changes occurred in the PTGS line, and these changes interfere with the initiation of transgene transcription, leading to a reversion of the PTGS phenotype.

Keywords: Bisulfite sequencing analysis - Cauliflower mosaic virus 35S promoter - Chalcone synthase - Cytosine methylation - Petunia hybrida - Post-transcriptional gene silencing

Abbreviations: CaMV, cauliflower mosaic virus; CHS, chalcone synthase; dsRNA, double-stranded RNA; Npt, neomycin phosphotransferase; NOS, nopaline synthase; OCS, octopine synthase; PTGS, post-transcriptional gene silencing; RdDM, RNA-directed DNA methylation; RT–PCR, reverse transcription–PCR; siRNA, short interfering RNA; TGS, transcriptional gene silencing

(Received October 30, 2006; Accepted February 17, 2007)
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