Spatial and functional relationships among Pol V-associated loci, Pol IV-dependent siRNAs, and cytosine methylation in the Arabidopsis epigenome

Andrzej T. Wierzbicki, Ross Cocklin, Anoop Mayampurath, Ryan Lister, M. Jordan Rowley, Brian D. Gregory, Joseph R. Ecker, Haixu Tang, Craig S. Pikaard

Research output: Contribution to journalArticlepeer-review

95 Scopus citations

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.

Original languageEnglish (US)
Pages (from-to)1825-1836
Number of pages12
JournalGenes and Development
Volume26
Issue number16
DOIs
StatePublished - Aug 15 2012

Keywords

  • DNA methylation
  • DNA-dependent RNA polymerase
  • Epigenetics
  • Gene silencing
  • RNA-directed DNA methylation
  • Short interfering RNA

ASJC Scopus subject areas

  • Genetics
  • Developmental Biology

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