Characterising the role of DOUBLE-STRANDED RNA BINDING2 in the microRNA and RNA-directed DNA methylation pathways of Arabidopsis thaliana floral tissue

AL Eamens

Centre for Plant Science, School of Environmental and Life Science, The University of Newcastle, Callaghan NSW 2308, Australia

In plants, microRNAs (miRNAs) are a class of non-protein-coding small RNA (sRNA), 21 to 22 nucleotides in length, and are key regulators of developmental gene expression. We have previously demonstrated that the DOUBLE-STRANDED RNA BINDING2 (DRB2) protein is directly involved in the production (liberation of the miRNA sRNA from the double-stranded RNA precursor transcript) of a subset of miRNAs in specific and developmentally important vegetative tissues of the genetic model species, Arabidopsis thaliana. Here, I will report that in Arabidopsis thaliana floral tissue, DRB2 plays an indirect role in the miRNA pathway to regulate the abundance of individual miRNA sRNAs. More specifically, the global population of another class of non-protein-coding sRNA, the repeat-associated small-interfering RNAs (rasiRNAs), is dramatically altered in the floral tissue of the drb2 knockout mutant. rasiRNA sRNAs, 24 nucleotides in length, are distinct to the miRNA class of sRNA and regulate the expression of both protein-coding and non-protein-coding genes at the transcriptional level via the RNA-directed DNA methylation (RdDM) mechanism of RNA silencing. I will go on to show that for many of the miRNAs with altered abundance in the floral tissue of the drb2 knockout mutant, the observed alterations result from changes to the methylation status of the promoter sequences of the miRNA genes from which the miRNA precursor transcripts are transcribed, and are not caused by defects in miRNA production in the absence of DRB2 activity as previously demonstrated in Arabidopsis thaliana vegetative tissues.