SRA- and SET-domain-containing proteins link RNA polymerase V occupancy to DNA methylation.
RNA-directed DNA methylation in Arabidopsis thaliana depends on the upstream synthesis of 24-nucleotide small interfering RNAs (siRNAs) by RNA POLYMERASE IV (Pol IV) and downstream synthesis of non-coding transcripts by Pol V. Pol V transcripts are thought to interact with siRNAs which then recruit DOMAINS REARRANGED METHYLTRANSFERASE 2 (DRM2) to methylate DNA. The SU(VAR)3-9 homologues SUVH2 and SUVH9 act in this downstream step but the mechanism of their action is unknown. Here we show that genome-wide Pol V association with chromatin redundantly requires SUVH2 and SUVH9. Although SUVH2 and SUVH9 resemble histone methyltransferases, a crystal structure reveals that SUVH9 lacks a peptide-substrate binding cleft and lacks a properly formed S-adenosyl methionine (SAM)-binding pocket necessary for normal catalysis, consistent with a lack of methyltransferase activity for these proteins. SUVH2 and SUVH9 both contain SRA (SET- and RING-ASSOCIATED) domains capable of binding methylated DNA, suggesting that they function to recruit Pol V through DNA methylation. Consistent with this model, mutation of DNA METHYLTRANSFERASE 1 (MET1) causes loss of DNA methylation, a nearly complete loss of Pol V at its normal locations, and redistribution of Pol V to sites that become hypermethylated. Furthermore, tethering SUVH9 [corrected] with a zinc finger to an unmethylated site is sufficient to recruit Pol V and establish DNA methylation and gene silencing. These results indicate that Pol V is recruited to DNA methylation through the methyl-DNA binding SUVH2 and SUVH9 proteins, and our mechanistic findings suggest a means for selectively targeting regions of plant genomes for epigenetic silencing.
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Lianna M
LM
Johnson
Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA.
Jiamu
J
Du
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Christopher J
CJ
Hale
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Sylvain
S
Bischof
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.|Howard Hughes Medical Institute, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Suhua
S
Feng
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Ramakrishna K
RK
Chodavarapu
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Xuehua
X
Zhong
Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA.
Giuseppe
G
Marson
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Matteo
M
Pellegrini
Genome Center and Department of Biochemistry and Molecular Medicine, University of California at Davis, Davis, CA 95616, USA.
David J
DJ
Segal
Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA.
Dinshaw J
DJ
Patel
Department of Molecular, Cell and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA.|Howard Hughes Medical Institute, University of California at Los Angeles, Los Angeles, CA 90095, USA.
Steven E
SE
Jacobsen
D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D055162:Biocatalysis; D002843:Chromatin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D035264:Flowers / Q000254:growth & development; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D018745:Genome, Plant / Q000235:genetics; D011495:Histone-Lysine N-Methyltransferase / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D018749:RNA, Plant / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D016335:Zinc Fingers
D029681:Arabidopsis Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D018749:RNA, Plant; D034741:RNA, Small Interfering; C499608:MET1 protein, Arabidopsis; D004248:DNA (Cytosine-5-)-Methyltransferases; D011495:Histone-Lysine N-Methyltransferase; C501526:SUVH2 protein, Arabidopsis; C587137:SUVH9 protein, Arabidopsis; C538841:RNA polymerase V, Arabidopsis; D012321:DNA-Directed RNA Polymerases; C504888:DRD1 protein, Arabidopsis