Distinct catalytic and non-catalytic roles of ARGONAUTE4 in RNA-directed DNA methylation.
DNA methylation has important functions in stable, transcriptional gene silencing, immobilization of transposable elements and genome organization. In Arabidopsis, DNA methylation can be induced by double-stranded RNA through the RNA interference (RNAi) pathway, a response known as RNA-directed DNA methylation. This requires a specialized set of RNAi components, including ARGONAUTE4 (AGO4). Here we show that AGO4 binds to small RNAs including small interfering RNAs (siRNAs) originating from transposable and repetitive elements, and cleaves target RNA transcripts. Single mutations in the Asp-Asp-His catalytic motif of AGO4 do not affect siRNA-binding activity but abolish its catalytic potential. siRNA accumulation and non-CpG DNA methylation at some loci require the catalytic activity of AGO4, whereas others are less dependent on this activity. Our results are consistent with a model in which AGO4 can function at target loci through two distinct and separable mechanisms. First, AGO4 can recruit components that signal DNA methylation in a manner independent of its catalytic activity. Second, AGO4 catalytic activity can be crucial for the generation of secondary siRNAs that reinforce its repressive effects.
Qi
Yijun
Y
Cold Spring Harbor Laboratory, Watson School of Biological Sciences and Howard Hughes Medical Institute, 1 Bungtown Road, Cold Spring Harbor, New York 11724, USA.
He
Xingyue
X
Wang
Xiu-Jie
XJ
Kohany
Oleksiy
O
Jurka
Jerzy
J
Hannon
Gregory J
GJ
D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D060565:Argonaute Proteins; D002384:Catalysis; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D018744:DNA, Plant / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D034622:RNA Interference; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
C471209:AGO4 protein, Arabidopsis; D029681:Arabidopsis Proteins; D060565:Argonaute Proteins; D018744:DNA, Plant; D018749:RNA, Plant; D034741:RNA, Small Interfering