Transcribing the RNAs of RNA-Directed DNA Methylation

The plant nuclear multisubunit RNA polymerases, Pol IV and Pol V evolved as specialized forms of DNA-dependent RNA polymerase II that transcribe noncoding RNAs for gene silencing, primarily of transposons. Pol IV physically associates with the single-subunit RNA-dependent RNA polymerases, RDR2 and together they generate double-stranded RNAs. These double-stranded RNAs are diced, by DCL3, into 24 nt short interfering RNAs (siRNAs) that associate with an Argonaute protein, primarily AGO4 or AGO6, and direct the cytosine methylation of matching DNA sequences by DRM2 by binding to Pol V transcripts, and Pol V itself, somehow recruiting proteins that carry out the chemical modification of the adjacent DNA and associated histones. How these biochemical reactions take place, and how the siRNAs and scaffold RNAs contribute to the reactions, is unclear. Using biochemistry, genetics and genomics, my lab is working to understand the enzymes and RNAs of the RNA-directed DNA methylation pathway. Enzymatic properties of Pol IV, Pol V and RDR2, determined in vitro, and chracteristics of the RNAs they synthesize will be the focus of my talk.