Identification of Pol IV and RDR2-dependent precursors of 24 nt siRNAs guiding de novo DNA methylation in Arabidopsis

Genes contain instructions for processes in cells and therefore their activities must be carefully controlled. The addition of small chemical tags called methyl groups to DNA is one of the many ways by which cells can influence gene activity. These methyl groups can silence genes by altering the DNA so that is more tightly packed within the nucleus of the cell. Virus genes and mobile sections of DNA called transposable elements (sometimes known as jumping genes) are also silenced by DNA methylation to keep them from doing harm. In plants, methyl groups can be attached to DNA by proteins that are guided to the DNA by molecules called short interfering ribonucleic acids (or siRNAs for short). Each siRNA is made of a chain of 24 building blocks called nucleotides and is able to bind to matching RNA molecules that are attached to the target DNA. The siRNAs are made from longer RNA molecules in a process that involves trimming by an enzyme called DCL3. However, it is not clear how long these “precursor” molecules are before DCL3 cuts them down to size. Here, Blevins, Podicheti et al. studied how siRNAs are made in a plant called Arabidopsis thaliana. The experiments show that RNAs containing around 26-45 nucleotides accumulate in cells that lack DCL3 and these cells are unable to make 24 nucleotide long siRNAs. Furthermore, the purified DCL3 enzyme can cut these precursor RNAs to make the siRNAs. Because the precursors are relatively short, the experiments suggest that DCL3 only cuts each precursor RNA once when making siRNAs. Blevins, Podicheti et al. also show that the siRNA precursors are made by a partnership of two RNA synthesizing enzymes. Therefore, a challenge for the future will be to understand exactly how they work together.