title: Arabidopsis HDA6 regulates locus-directed heterochromatin silencing in cooperation with MET1.
abstract: Heterochromatin silencing is pivotal for genome stability in eukaryotes. In Arabidopsis, a plant-specific mechanism called RNA-directed DNA methylation (RdDM) is involved in heterochromatin silencing. Histone deacetylase HDA6 has been identified as a component of such machineries; however, its endogenous targets and the silencing mechanisms have not been analyzed globally. In this study, we investigated the silencing mechanism mediated by HDA6. Genome-wide transcript profiling revealed that the loci silenced by HDA6 carried sequences corresponding to the RDR2-dependent 24-nt siRNAs, however their transcript levels were mostly unaffected in the rdr2 mutant. Strikingly, we observed significant overlap of genes silenced by HDA6 to those by the CG DNA methyltransferase MET1. Furthermore, regardless of dependence on RdDM pathway, HDA6 deficiency resulted in loss of heterochromatic epigenetic marks and aberrant enrichment for euchromatic marks at HDA6 direct targets, along with ectopic expression of these loci. Acetylation levels increased significantly in the hda6 mutant at all of the lysine residues in the H3 and H4 N-tails, except H4K16. Interestingly, we observed two different CG methylation statuses in the hda6 mutant. CG methylation was sustained in the hda6 mutant at some HDA6 target loci that were surrounded by flanking DNA-methylated regions. In contrast, complete loss of CG methylation occurred in the hda6 mutant at the HDA6 target loci that were isolated from flanking DNA methylation. Regardless of CG methylation status, CHG and CHH methylation were lost and transcriptional derepression occurred in the hda6 mutant. Furthermore, we show that HDA6 binds only to its target loci, not the flanking methylated DNA, indicating the profound target specificity of HDA6. We propose that HDA6 regulates locus-directed heterochromatin silencing in cooperation with MET1, possibly recruiting MET1 to specific loci, thus forming the foundation of silent chromatin structure for subsequent non-CG methylation.
authors.affiliation: Plant Genomic Network Research Team, RIKEN Plant Science Center, Yokohama, Kanagawa, Japan.
authors.forename: Taiko Kim
authors.initials: TK
authors.lastname: To
authors.forename: Jong-Myong
authors.initials: JM
authors.lastname: Kim
authors.forename: Akihiro
authors.initials: A
authors.lastname: Matsui
authors.forename: Yukio
authors.initials: Y
authors.lastname: Kurihara
authors.forename: Taeko
authors.initials: T
authors.lastname: Morosawa
authors.forename: Junko
authors.initials: J
authors.lastname: Ishida
authors.forename: Maho
authors.initials: M
authors.lastname: Tanaka
authors.forename: Takaho
authors.initials: T
authors.lastname: Endo
authors.forename: Tetsuji
authors.initials: T
authors.lastname: Kakutani
authors.forename: Tetsuro
authors.initials: T
authors.lastname: Toyoda
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kimura
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
authors.forename: Kazuo
authors.initials: K
authors.lastname: Shinozaki
authors.forename: Motoaki
authors.initials: M
authors.lastname: Seki
mesh_terms: D000107:Acetylation; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D056426:Genetic Loci; D006570:Heterochromatin / Q000378:metabolism*; D006655:Histone Deacetylases / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding
chemical_list: D029681:Arabidopsis Proteins; D006570:Heterochromatin; C499608:MET1 protein, Arabidopsis; D004248:DNA (Cytosine-5-)-Methyltransferases; C432452:AT5G63110 protein, Arabidopsis; D006655:Histone Deacetylases