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Coincident sequence-specific RNA degradation of linked transgenes in the plant genome

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Abstract

The expression of transgenes in plant genomes can be inhibited by either transcriptional gene silencing or posttranscriptional gene silencing (PTGS). Overexpression of the chalcone synthase-A (CHS-A) transgene triggers PTGS of CHS-A and thus results in loss of flower pigmentation in petunia. We previously demonstrated that epigenetic inactivation of CHS-A transgene transcription leads to a reversion of the PTGS phenotype. Although neomycin phosphotransferase II (nptII), a marker gene co-introduced into the genome with the CHS-A transgene, is not normally silenced in petunia, even when CHS-A is silenced, here we found that nptII was silenced in a petunia line in which CHS-A PTGS was induced, but not in the revertant plants that had no PTGS of CHS-A. Transcriptional activity, accumulation of short interfering RNAs, and restoration of mRNA level after infection with viruses that had suppressor proteins of gene silencing indicated that the mechanism for nptII silencing was posttranscriptional. Read-through transcripts of the CHS-A gene toward the nptII gene were detected. Deep-sequencing analysis revealed a striking difference between the predominant size class of small RNAs produced from the read-through transcripts (22 nt) and that from the CHS-A RNAs (21 nt). These results implicate the involvement of read-through transcription and distinct phases of RNA degradation in the coincident PTGS of linked transgenes and provide new insights into the destabilization of transgene expression.

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Acknowledgments

We are grateful to Mike O’Dell for helpful discussions and maintenance of plant materials. We also thank Cathie Martin for the Antirrhinum ubiquitin DNA clone, Mineo Senda for small RNA analysis, Neal Gutterson and Richard Jorgensen for plant materials, and Michael Metzlaff, Richard B. Falavell, Kenji Nakahara, Tetsuya Yamada and Hanako Shimura for valuable comments during the course of this work. This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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  1. Research Faculty of Agriculture, Hokkaido University, Sapporo, 060-8589, Japan

    Megumi Kasai, Maiko Koseki, Kazunori Goto, Chikara Masuta, Shiho Ishii & Akira Kanazawa

  2. The New Zealand Institute for Plant and Food Research Ltd, Private Bag 92 169, Auckland, 1142, New Zealand

    Roger P. Hellens

  3. Graduate School of Science and Technology, Hirosaki University, Hirosaki, Aomori, 036-8561, Japan

    Akito Taneda

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  1. Megumi Kasai
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  2. Maiko Koseki
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Correspondence to Akira Kanazawa.

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Kasai, M., Koseki, M., Goto, K. et al. Coincident sequence-specific RNA degradation of linked transgenes in the plant genome. Plant Mol Biol 78, 259–273 (2012). https://doi.org/10.1007/s11103-011-9863-0

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