title: Detection of Pol IV/RDR2-dependent transcripts at the genomic scale in Arabidopsis reveals features and regulation of siRNA biogenesis.
abstract: Twenty-four-nucleotide small interfering (si)RNAs are central players in RNA-directed DNA methylation (RdDM), a process that establishes and maintains DNA methylation at transposable elements to ensure genome stability in plants. The plant-specific RNA polymerase IV (Pol IV) is required for siRNA biogenesis and is believed to transcribe RdDM loci to produce primary transcripts that are converted to double-stranded RNAs (dsRNAs) by RDR2 to serve as siRNA precursors. Yet, no such siRNA precursor transcripts have ever been reported. Here, through genome-wide profiling of RNAs in genotypes that compromise the processing of siRNA precursors, we were able to identify Pol IV/RDR2-dependent transcripts from tens of thousands of loci. We show that Pol IV/RDR2-dependent transcripts correspond to both DNA strands, whereas the RNA polymerase II (Pol II)-dependent transcripts produced upon derepression of the loci are derived primarily from one strand. We also show that Pol IV/RDR2-dependent transcripts have a 5' monophosphate, lack a poly(A) tail at the 3' end, and contain no introns; these features distinguish them from Pol II-dependent transcripts. Like Pol II-transcribed genic regions, Pol IV-transcribed regions are flanked by A/T-rich sequences depleted in nucleosomes, which highlights similarities in Pol II- and Pol IV-mediated transcription. Computational analysis of siRNA abundance from various mutants reveals differences in the regulation of siRNA biogenesis at two types of loci that undergo CHH methylation via two different DNA methyltransferases. These findings begin to reveal features of Pol IV/RDR2-mediated transcription at the heart of genome stability in plants.
authors.affiliation: Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, California 92521, USA;
authors.forename: Shaofang
authors.initials: S
authors.lastname: Li
authors.affiliation: Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA;
authors.forename: Lee E
authors.initials: LE
authors.lastname: Vandivier
authors.affiliation: Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, California 92521, USA;
authors.forename: Bin
authors.initials: B
authors.lastname: Tu
authors.affiliation: Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, California 92521, USA;
authors.forename: Lei
authors.initials: L
authors.lastname: Gao
authors.affiliation: Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, California 92521, USA;
authors.forename: So Youn
authors.initials: SY
authors.lastname: Won
authors.affiliation: Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, California 92521, USA;
authors.forename: Shengben
authors.initials: S
authors.lastname: Li
authors.affiliation: State Key Laboratory of Genetic Engineering and Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai 200433, China;
authors.forename: Binglian
authors.initials: B
authors.lastname: Zheng
authors.affiliation: Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA;
authors.forename: Brian D
authors.initials: BD
authors.lastname: Gregory
authors.affiliation: Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, California 92521, USA; Howard Hughes Medical Institute, University of California, Riverside, California 92521, USA xuemei.chen@ucr.edu.
authors.forename: Xuemei
authors.initials: X
authors.lastname: Chen
mesh_terms: D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000378:metabolism*; D019175:DNA Methylation; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D018745:Genome, Plant; D042822:Genomic Instability; D023281:Genomics; D008954:Models, Biological; D012322:RNA Precursors; D018749:RNA, Plant; D034741:RNA, Small Interfering / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D029681:Arabidopsis Proteins; D012322:RNA Precursors; D018749:RNA, Plant; D034741:RNA, Small Interfering; C499181:RNA polymerase IV, Arabidopsis; C519655:RDR2 protein, Arabidopsis; D012324:RNA-Dependent RNA Polymerase; D012321:DNA-Directed RNA Polymerases