Pol II-directed short RNAs suppress the nuclear export of mRNA
The synthesis and subsequent nuclear export of non-coding RNA (ncRNA) directed by RNA polymerase (Pol) II is very sensitive to abiotic and biotic external stimuli including pathogen challenges. To assess whether stress-induced ncRNAs may suppress the nuclear export of mRNA, we exploited the ability of Agrobacterium tumefaciens to co-deliver Pol I, II and III promoter-based vectors for the transcription of short (s) ncRNAs, GFP mRNA or genomic RNA of plant viruses (Tobacco mosaic virus, TMV; or Potato virus X, PVX) into the nucleus of Nicotiana benthamiana cells. We showed that, in contrast to Pol I- and Pol III-derived sncRNAs, all tested Pol II-derived sncRNAs (U6 RNA, tRNA or artificial RNAs) resulted in decreased expression of GFP and host mRNA. The level of this inhibitory effect depended on the non-coding transcript length and promoter strength. Short coding RNA (scRNA) can also compete with mRNA for nuclear export. We showed that scRNA, an artificial 117-nt short sequence encoding Elastin-Like peptide element tandems with FLAG sequence (ELF) and the 318-nt N. benthamiana antimicrobial peptide thionin (defensin) gene efficiently decreased GFP expression. The stress-induced export of Pol II-derived sncRNA and scRNA into the cytoplasm via the mRNA export pathway may block nucleocytoplasmic traffic including the export of mRNA responsible for antivirus protection. Consistent with this model, we observed that Pol II-derived sncRNAs as well as scRNA, thionin and ELF strongly enhanced the cytoplasmic reproduction of TMV and PVX RNA.
KeywordsNon-coding RNAs Nuclear export mRNA Plant virus RNA polymerase II Short coding RNA
We thank Drs. C. S. Pikaard (at Washington University St. Louis), T. Okuno (Kyoto University), A. E. Simon (University of Maryland, College Park) and H. Beier (University of Würzburg) for generously providing the Pol I promoter containing plasmid pBor2, pBICmiR171prec, GNC encoding plasmid, and plasmid pNtY1 encoding Nicotiana rustica pre-tRNATyr, respectively. We also thank members of the MSU Department of Virology for helpful discussions and technical assistance. This work was partly supported by the Russian Foundation for Basic Research (grants 08-04-00106 and 08-04-12073) and Icon Genetics GmbH. The work in the VC laboratory is supported by grants from NIH, NSF, NRI USDA CSREE, BARD, and BSF.
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