title: Spatial and functional relationships among Pol V-associated loci, Pol IV-dependent siRNAs, and cytosine methylation in the Arabidopsis epigenome.
abstract: Multisubunit RNA polymerases IV and V (Pols IV and V) mediate RNA-directed DNA methylation and transcriptional silencing of retrotransposons and heterochromatic repeats in plants. We identified genomic sites of Pol V occupancy in parallel with siRNA deep sequencing and methylcytosine mapping, comparing wild-type plants with mutants defective for Pol IV, Pol V, or both Pols IV and V. Approximately 60% of Pol V-associated regions encompass regions of 24-nucleotide (nt) siRNA complementarity and cytosine methylation, consistent with cytosine methylation being guided by base-pairing of Pol IV-dependent siRNAs with Pol V transcripts. However, 27% of Pol V peaks do not overlap sites of 24-nt siRNA biogenesis or cytosine methylation, indicating that Pol V alone does not specify sites of cytosine methylation. Surprisingly, the number of methylated CHH motifs, a hallmark of RNA-directed de novo methylation, is similar in wild-type plants and Pol IV or Pol V mutants. In the mutants, methylation is lost at 50%-60% of the CHH sites that are methylated in the wild type but is gained at new CHH positions, primarily in pericentromeric regions. These results indicate that Pol IV and Pol V are not required for cytosine methyltransferase activity but shape the epigenome by guiding CHH methylation to specific genomic sites.
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Andrzej T
authors.initials: AT
authors.lastname: Wierzbicki
authors.forename: Ross
authors.initials: R
authors.lastname: Cocklin
authors.forename: Anoop
authors.initials: A
authors.lastname: Mayampurath
authors.forename: Ryan
authors.initials: R
authors.lastname: Lister
authors.forename: M Jordan
authors.initials: MJ
authors.lastname: Rowley
authors.forename: Brian D
authors.initials: BD
authors.lastname: Gregory
authors.forename: Joseph R
authors.initials: JR
authors.lastname: Ecker
authors.forename: Haixu
authors.initials: H
authors.lastname: Tang
authors.forename: Craig S
authors.initials: CS
authors.lastname: Pikaard
mesh_terms: D020816:Amino Acid Motifs; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D003596:Cytosine / Q000378:metabolism*; D019175:DNA Methylation; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D018745:Genome, Plant; D009154:Mutation; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D034741:RNA, Small Interfering; D003596:Cytosine; C499181:RNA polymerase IV, Arabidopsis; C538841:RNA polymerase V, Arabidopsis; D012321:DNA-Directed RNA Polymerases