Project description:In Arabidopsis, CHG DNA methylation is controlled by the H3K9 methylation mark through a self-reinforcing loop between DNA methyltransferase CHROMOMETHYLASE3 (CMT3) and H3K9 histone methyltransferase KRYPTONITE/SUVH4 (KYP). We report on the structure of KYP in complex with methylated DNA, substrate H3 peptide and cofactor SAH, thereby defining the spatial positioning of the SRA domain relative to the SET domain. The methylated DNA is bound by the SRA domain with the 5mC flipped out of the DNA, while the H3(1-15) peptide substrate binds between the SET and post-SET domains, with the epsilon-ammonium of K9 positioned adjacent to bound SAH. These structural insights complemented by in vivo functional data on key mutants of residues lining the 5mC and H3K9-binding pockets within KYP, establish how methylated DNA recruits KYP to the histone substrate. Together, the structures of KYP and previously reported CMT3 complexes provide insights into molecular mechanisms linking DNA and histone methylation.
Project description:DNA methylation can be established by RNA-directed DNA methylation (RdDM) in plants. The association of RNA polymerase V (Pol V) with chromatin is a critical step for RdDM. While the SRA-domain-containing proteins SUVH2 and SUVH9 and the DDR complex are known to be required for the association of Pol V with chromatin, it is unknown whether the association of Pol V with chromatin requires other unidentified regulators. Here we found that SUVH9 is able to interact with a conserved histone-interaction protein, FVE, and a previously uncharacterized RRM domain-containing protein, which we named RRM1. We demonstrated that FVE facilitates the association of Pol V with chromatin and thus contributes to DNA methylation at a substantial subset of RdDM target loci, while RRM1 is only slightly involved in RdDM. FVE-dependent RdDM target loci are more abundant in gene-rich chromosome arms than FVE-independent RdDM target loci. FVE was previously shown to be a shared subunit of the RPD3-type histone deacetylase complex and the polycomb-type histone H3K27 trimethyltransferase complex, both of which are involved in transcriptional repression. This study reveals a previously uncharacterized role of FVE in RdDM and suggests that FVE may coordinate RdDM, histone deacetylation, and H3K27 trimethylation, thus ensuring transcriptional silencing of TEs in gene-rich chromosome arms to protect genes from harmful effects of potentially transcribed TEs
Project description:DNA methylation can be established by RNA-directed DNA methylation (RdDM) in plants. The association of RNA polymerase V (Pol V) with chromatin is a critical step for RdDM. While the SRA-domain-containing proteins SUVH2 and SUVH9 and the DDR complex are known to be required for the association of Pol V with chromatin, it is unknown whether the association of Pol V with chromatin requires other unidentified regulators. Here we found that SUVH9 is able to interact with a conserved histone-interaction protein, FVE, and a previously uncharacterized RRM domain-containing protein, which we named RRM1. We demonstrated that FVE facilitates the association of Pol V with chromatin and thus contributes to DNA methylation at a substantial subset of RdDM target loci, while RRM1 is only slightly involved in RdDM. FVE-dependent RdDM target loci are more abundant in gene-rich chromosome arms than FVE-independent RdDM target loci. FVE was previously shown to be a shared subunit of the RPD3-type histone deacetylase complex and the polycomb-type histone H3K27 trimethyltransferase complex, both of which are involved in transcriptional repression. This study reveals a previously uncharacterized role of FVE in RdDM and suggests that FVE may coordinate RdDM, histone deacetylation, and H3K27 trimethylation, thus ensuring transcriptional silencing of TEs in gene-rich chromosome arms to protect genes from harmful effects of potentially transcribed TEs.
Project description:DNA methylation can be established by RNA-directed DNA methylation (RdDM) in plants. The association of RNA polymerase V (Pol V) with chromatin is a critical step for RdDM. While the SRA-domain-containing proteins SUVH2 and SUVH9 and the DDR complex are known to be required for the association of Pol V with chromatin, it is unknown whether the association of Pol V with chromatin requires other unidentified regulators. Here we found that SUVH9 is able to interact with a conserved histone-interaction protein, FVE, and a previously uncharacterized RRM domain-containing protein, which we named RRM1. We demonstrated that FVE facilitates the association of Pol V with chromatin and thus contributes to DNA methylation at a substantial subset of RdDM target loci, while RRM1 is only slightly involved in RdDM. FVE-dependent RdDM target loci are more abundant in gene-rich chromosome arms than FVE-independent RdDM target loci. FVE was previously shown to be a shared subunit of the RPD3-type histone deacetylase complex and the polycomb-type histone H3K27 trimethyltransferase complex, both of which are involved in transcriptional repression. This study reveals a previously uncharacterized role of FVE in RdDM and suggests that FVE may coordinate RdDM, histone deacetylation, and H3K27 trimethylation, thus ensuring transcriptional silencing of TEs in gene-rich chromosome arms to protect genes from harmful effects of potentially transcribed TEs.
Project description:DNA methylation can be established by RNA-directed DNA methylation (RdDM) in plants. The association of RNA polymerase V (Pol V) with chromatin is a critical step for RdDM. While the SRA-domain-containing proteins SUVH2 and SUVH9 and the DDR complex are known to be required for the association of Pol V with chromatin, it is unknown whether the association of Pol V with chromatin requires other unidentified regulators. Here we found that SUVH9 is able to interact with a conserved histone-interaction protein, FVE, and a previously uncharacterized RRM domain-containing protein, which we named RRM1. We demonstrated that FVE facilitates the association of Pol V with chromatin and thus contributes to DNA methylation at a substantial subset of RdDM target loci, while RRM1 is only slightly involved in RdDM. FVE-dependent RdDM target loci are more abundant in gene-rich chromosome arms than FVE-independent RdDM target loci. FVE was previously shown to be a shared subunit of the RPD3-type histone deacetylase complex and the polycomb-type histone H3K27 trimethyltransferase complex, both of which are involved in transcriptional repression. This study reveals a previously uncharacterized role of FVE in RdDM and suggests that FVE may coordinate RdDM, histone deacetylation, and H3K27 trimethylation, thus ensuring transcriptional silencing of TEs in gene-rich chromosome arms to protect genes from harmful effects of potentially transcribed TEs.
Project description:Epigenetic information can be inherited through a process known as transgenerational epigenetic inheritance (TEI). TEI can be initiated and maintained by small RNAs and histone modifications. The latter includes histone methylation, deposited by proteins with methyltransferase activity, including SET domain-containing proteins. Other SET domain-containing proteins with no catalytic methyltransferase activity, also adopt roles in chromatin and gene expression regulation. Here, we describe SET-24, a SET domain-containing protein in Caenorhabditis elegans that belongs to a family of catalytically inactive SET proteins. SET-24 localises to germline nuclei and is required for germline immortality. Additionally, the inheritance of small RNA-driven epigenetic silencing is compromised in set-24 mutants. Using quantitative proteomics and yeast two-hybrid assays, we found that SET-24 interacts with Host Cell Factor 1 (HCF-1), a protein involved in epigenetic regulation and associated with known chromatin remodelling complexes, like COMPASS, which deposits H3K4me3. In set-24 mutants, hundreds of genes display increased H3K4me3 levels at their transcription start sites. While these changes are not matched at the transcriptional level, small RNA production is disrupted in approximately one fifth of those genes, which are normally targeted by small RNA pathways. We propose that SET-24 is a factor required to maintain epigenetic memory in the germline by maintaining a chromatin environment permissible to small RNA biogenesis over generations.
Project description:The RNA polymerase II (RNApII) C-terminal domain (CTD)-interacting domain (CID) proteins are involved in two distinct termination pathways and recognize different phosphorylated forms of CTD. To investigate the role of differential CTDM-^VCID interactions in the choice of termination pathway, we altered the CTD-binding specificity of Nrd1 by domain swapping. ChIP-chip was performed to examine the effect of Nrd1 CID swapping on genome-wide RNA polymerase II (Rpb3 antibody, Neoclone) occupancy. Nrd1 with the CID from Rtt103 (Nrd1[CID-Rtt103]; strain YSB2445) causes read-through transcription at many genes, but can trigger termination where multiple Nrd1/Nab3-binding sites and serine 2 phosphorylated CTD co-exist.
Project description:Retrotransposons are classified into long terminal repeat (LTR) or non-LTR retrotransoposons, and both types can mobilize in a “copy and paste” manner. Rice genome contains >40% of repetitive sequences or transposable elements (TEs) that scattered throughout the genome. TE activities are tightly regulated by distinct epigenetic mechanisms. Histone lysine methylation is catalyzed by SET domain group (SDG) protein and reversed by a family of Jumonji C (JmjC) domain-containing proteins. In rice, DNA methylation and histone methylation have been implicated in controlling copia-like LTR retrotransposon Tos17 activities. Whether any TEs are controlled by histone demethylase remains to be elusive. Here, we show that JMJ703 is a histone H3K4 specific demethylase in rice. Impaired JMJ703 disrupted genome-wide transcriptome and enhanced H3K4me3 resulting in pleiotropic defects. Two LINE elements, Karma and its N-terminal truncation were identified as direct targets of JMJ703 with increased frequency of transposition in jmj703, while Tos17 is unaffected. Our findings show that plants use H3K4me3 demethylase to constitutively remove active chromatin mark and maintain silencing status at a subset retrotransposons which localize in heterochromatin regions. Therefore, our work uncovers a novel mechanism to control retrotransposon activity by histone demethylation, which further strengthens the link between epigenetic silencing and genome stability. Examination of differences in H3K4 between jmj703 and wild type.