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SRX20209035: GSM7290819: Input for DDM1 ChIP-seq in ddm1 biol Rep1; Arabidopsis thaliana; ChIP-Seq
1 ILLUMINA (NextSeq 2000) run: 40M spots, 12.1G bases, 3.5Gb downloads

External Id: GSM7290819_r1
Submitted by: CSHL
Study: Chromatin remodeling of histone H3 variants by DDM1 underlies epigenetic inheritance of DNA methylation [ChIP-seq]
show Abstracthide Abstract
Epigenetic inheritance refers to the faithful replication of DNA and histone modification independent of DNA sequence. Nucleosomes block access to DNA methyltransferase during S phase, unless they are remodeled by Decrease in DNA methylation 1 (DDM1[Lsh/HELLS]), a Snf2-like master regulator of epigenetic inheritance. We show that DDM1 activity results in replacement of the transcriptional histone variant H3.3 for the replicative variant H3.1. Inddm1mutants, DNA methylation can be restored by loss of the H3.3 chaperone HIRA, while the H3.1 chaperone CAF-1 becomes essential. The single-particle cryo-EM structure at 3.2 Å of DDM1 with a variant nucleosome reveals direct engagement at SHL2 with histone H3.3 at or near variant residues required for assembly, as well as with the deacetylated H4 tail. An N-terminal autoinhibitory domain binds H2A variants to allow remodeling, while a di-sulphide bond in the helicase domain is essential for activity in vivo and in vitro. Differential remodeling of H3 and H2A variants in vitro reflects preferential deposition in vivo. DDM1 co-localizes with H3.1 and H3.3 during the cell cycle, and with the DNA methyltransferase MET1[Dnmt1]. DDM1 localization to the chromosome is blocked by H4K16 acetylation, which accumulates at DDM1 targets in ddm1 mutants, as does the sperm cell specific H3.3 variant MGH3 in pollen, which acts as a placeholder nucleosome and contributes to epigenetic inheritance. Overall design: Chromatin Immunoprecipitation sequencing was performed on wild-type and ddm1 mutants for DDM1 (vs Input), H3.3 (vs H3), H3K27me1 (vs H3), H4K16ac (vs H4) and MGH3 (vs Input) Please note that processed data generated from both ChIP and Input/control sample is linked to the corresponding ChIP sample records.
Sample: Input for DDM1 ChIP-seq in ddm1 biol Rep1
SAMN34567984 • SRS17533143 • All experiments • All runs
Library:
Name: GSM7290819
Instrument: NextSeq 2000
Strategy: ChIP-Seq
Source: GENOMIC
Selection: ChIP
Layout: PAIRED
Construction protocol: ChIP was performed as previously described (Parent et al., 2021), starting with 1g of seedlings. In brief, after crosslinking in 1% formaldehyde for 10 min, the tissue was ground to a fine powder in liquid nitrogen, chromatin was extracted with 1% SDS Tris-based lysis buffer and sonicated to ~200bp fragments. Chromatin was cleared with protein A magnetic beads and incubated overnight with the antibody listed below. Immune complexes were eluted with low, and then high salt buffers, before reversing crosslinks and purifying DNA fragments with ChIP DNA clean and concentrator kit (Zymo Research). Low input ChIP for MGH3-GFP was performed with ~10e8 pollen grains. Fixation was for 15 min and subsequent grinding with acid-washed glass beads. DNA fragments underwent a preliminary phenol-chloroform and ethanol precipitation step before clean up. NEBNext Ultra II DNA Library Prep Kit
Runs: 1 run, 40M spots, 12.1G bases, 3.5Gb
Run# of Spots# of BasesSizePublished
SRR2442124339,982,48312.1G3.5Gb2023-08-28

ID:
27629166

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