AGO4 is specifically required for heterochromatic siRNA accumulation at Pol V-dependent loci in Arabidopsis thaliana

Plant J. 2017 Apr;90(1):37-47. doi: 10.1111/tpj.13463. Epub 2017 Feb 11.


In plants, 24 nucleotide long heterochromatic siRNAs (het-siRNAs) transcriptionally regulate gene expression by RNA-directed DNA methylation (RdDM). The biogenesis of most het-siRNAs depends on the plant-specific RNA polymerase IV (Pol IV), and ARGONAUTE4 (AGO4) is a major het-siRNA effector protein. Through genome-wide analysis of sRNA-seq data sets, we found that AGO4 is required for the accumulation of a small subset of het-siRNAs. The accumulation of AGO4-dependent het-siRNAs also requires several factors known to participate in the effector portion of the RdDM pathway, including RNA POLYMERASE V (POL V), DOMAINS REARRANGED METHYLTRANSFERASE 2 (DRM2) and SAWADEE HOMEODOMAIN HOMOLOGUE 1 (SHH1). Like many AGO proteins, AGO4 is an endonuclease that can 'slice' RNAs. We found that a slicing-defective AGO4 was unable to fully recover AGO4-dependent het-siRNA accumulation from ago4 mutant plants. Collectively, our data suggest that AGO4-dependent siRNAs are secondary siRNAs dependent on the prior activity of the RdDM pathway at certain loci.

Keywords: ARGONAUTE4; heterochromatic siRNA accumulation; sRNA-seq; secondary heterochromatic siRNAs; slicing ability.

MeSH terms

  • Arabidopsis / enzymology*
  • Arabidopsis / genetics
  • Arabidopsis / microbiology*
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism*
  • Argonaute Proteins / genetics
  • Argonaute Proteins / metabolism*
  • DNA-Directed RNA Polymerases / genetics
  • DNA-Directed RNA Polymerases / metabolism
  • Gene Expression Regulation, Plant / genetics
  • Gene Expression Regulation, Plant / physiology
  • Gene Silencing / physiology
  • Heterochromatin / genetics
  • Homeodomain Proteins / genetics
  • Homeodomain Proteins / metabolism
  • Methyltransferases / genetics
  • Methyltransferases / metabolism
  • RNA, Plant / genetics
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / physiology


  • AGO4 protein, Arabidopsis
  • Arabidopsis Proteins
  • Argonaute Proteins
  • Heterochromatin
  • Homeodomain Proteins
  • RNA, Plant
  • RNA, Small Interfering
  • SHH1 protein, Arabidopsis
  • Methyltransferases
  • DRM2 protein, Arabidopsis
  • RNA polymerase IV, Arabidopsis
  • RNA polymerase V, Arabidopsis
  • DNA-Directed RNA Polymerases

Associated data

  • GENBANK/GSE79119
  • GENBANK/GSE87333