Identification of trans-acting siRNAs and their regulatory cascades in grapevine.

Abstract

Trans-acting small interfering RNAs (ta-siRNAs) play an essential role in the regulation of plant gene expression, but relevant reports are still limited. Bioinformatic analyses indicate that many ta-siRNA-producing loci (TASs) are present in plants, implying the existence of as yet undiscovered ta-siRNAs and related regulatory pathways. To expand our knowledge of these plant gene regulators, we applied high-throughput computational and experimental methods to grapevine (Vitis vinifera L.). Based on bioinformatic predictions, we identified 49 TASs from 49 055 small RNA clusters. Using RNA degradome analysis, we experimentally validated 5 TASs, 22 ta-siRNAs and 37 ta-siRNA targets. The cis-activities of ta-siRNAs were also confirmed, which suggested an inactive mechanism of TAS transcription, and a produced mechanism of multiple forms of small RNA from same TAS. We examined the conservation of newly identified ta-siRNA regulatory cascades and found that while the cascade related to vviTAS3 was conserved, cascades related to vviTAS7, vviTAS8, vviTAS9 and vviTAS10 were grape-specific. These results broaden the known scope of ta-siRNA regulation.