Abstract
In plants, transgenes containing Potato spindle tuber viroid (PSTVd) cDNA sequences were efficient targets of PSTVd infection-mediated RNA-directed DNA methylation. Here, we demonstrate that in PSTVd-infected tobacco plants, a 134 bp PSTVd fragment (PSTVd-134) did not become densely methylated when it was inserted into a chimeric Satellite tobacco mosaic virus (STMV) construct. Only about 4–5% of all cytosines (Cs) of the PSTVd-134 were methylated when flanked by satellite sequences. In the same plants, C methylation was approximately 92% when the PSTVd-134 was in a PSTVd full length sequence context and roughly 33% when flanked at its 3′ end by a 19 bp PSTVd and at its 5′ end by a short viroid-unrelated sequence. In addition, PSTVd small interfering RNAs (siRNAs) produced from the replicating viroid failed to target PSTVd-134-containing chimeric STMV RNA for degradation. Satellite RNAs appear to have adopted secondary structures that protect them against RNA interference (RNAi)—mediated degradation. Protection can be extended to short non-satellite sequences residing in satellite RNAs, rendering them poor targets for nuclear and cytoplasmic RNAi induced in trans.
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Acknowledgments
We are grateful to Michael Metzlaff (Bayer BioScience N.V., Belgium) for providing the pEV311 vector and TMGMV-infected tobacco material. We thank Michele Zwiebel and Milena Kitova for excellent technical assistance with tissue culture and Annerie Liebenberg for critical reading of the manuscript. This work was supported by the grants of the Sixth Research Framework Programs of the European Union, Project LSHG-CT-2006-037900 (SIROCCO) and LSHG-CT-2004-005120 (FOSRAK).
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Dalakouras, A., Moser, M., Krczal, G. et al. A chimeric satellite transgene sequence is inefficiently targeted by viroid-induced DNA methylation in tobacco. Plant Mol Biol 73, 439–447 (2010). https://doi.org/10.1007/s11103-010-9631-6
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DOI: https://doi.org/10.1007/s11103-010-9631-6