OsChz1 acts as a histone chaperone in modulating chromatin organization and genome function in rice

While the yeast Chz1 acts as a specific histone-chaperone for H2A.Z, functions of CHZ-domain proteins in multicellular eukaryotes remain obscure. Here, we report on the functional characterization of OsChz1, a sole CHZ-domain protein identified in rice. OsChz1 interacts with both the canonical H2A-H2B dimer and the variant H2A.Z-H2B dimer. Within crystal structure the C-terminal region of OsChz1 binds H2A-H2B via an acidic region, pointing to a previously unknown recognition mechanism. Knockout of OsChz1 leads to multiple plant developmental defects. At genome-wide level, loss of OsChz1 causes mis-regulations of thousands of genes and broad alterations of nucleosome occupancy as well as reductions of H2A.Z-enrichment. While OsChz1 associates with chromatin regions enriched of repressive histone marks (H3K27me3 and H3K4me2), its loss does not affect the genome landscape of DNA methylation. Taken together, it is emerging that OsChz1 functions as an important H2A/H2A.Z-H2B chaperone in dynamic regulation of chromatin for higher eukaryote development.

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Wen-Hui Shen
Oct 20, 2020 For RT-qPCR and ChIP-qPCR, data were collected with CFX ConnectTM Real-Time System (BIO-RAD); RNA-seq, ChIP-seq and MNase-seq data were collected using IIllumina HiSeq3000 instrument via the custom service of GENERGY BIO (Shanghai, China). Images of gels or blots were collected by Clinx Science Instruments (Shanghai, China). Confocal images were acquired with ZEN 2010 software (version 1.0.3846.26637). ITC data were collected by MicroCal iTC200 (GE Healthcare). X-ray diffraction data were collected on beamline BL17U1 and BL19U1 at Shanghai Synchrotron Radiation Facility.
Neighbor-Joining phylogeny based on full-length amino acid sequence alignment was calculated using MEGA (v10.1.8) and illustrated using FigTree v1.4.3; Publicly available computational tools for analyzing RNA-seq, ChIP-seq and BS-seq data were used as described in the Methods section and they include: FastQC (v0.11.7), CUTADAPT (v1.10), HISAT2 (v2.1.0), Samtools (v1.9), FeatureCounts (v1.6. Data Policy information about availability of data All manuscripts must include a data availability statement. This statement should provide the following information, where applicable: -Accession codes, unique identifiers, or web links for publicly available datasets -A list of figures that have associated raw data -A description of any restrictions on data availability Field-specific reporting Please select the one below that is the best fit for your research. If you are not sure, read the appropriate sections before making your selection.

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All studies must disclose on these points even when the disclosure is negative. The authors declare that all data supporting the findings of this work are available within the paper and its Supplementary Information files, and that they are available from the corresponding author upon reasonable request. The RNA-seq, ChIP-seq, MNase-seq and BS-seq data that support the findings of this study have been deposited to NCBI GEO with the accession number GSE155269 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE155269). Structural factors and coordinates have been deposited in the Protein Data Bank (PDB) with accession code 6M2M (https://www.rcsb.org/structure/6M2M) for the OsChz1-H2A-H2B complex. The source data underlying Figures 1b, 2a ,3d,5c,d,6a,d,f,g,7b,c,d,f,8a,as well as Supplementary Figures 4,7,8b,10a,c,e,12  No data was excluded from analysis in this study.
For H2A.Z, H3 and OsChz1-Myc ChIP-seq experiments, 2 replicates were performed along with DNA input as ChIP-seq control. For RNA-seq, RT-PCR and ChIP-PCR, three replicates were performed as indicated in the figure legends. For MNase-seq and BS-seq, 2 replicates were performed. Flowering time experiments, GST pull-down assay, Co-IP and ITC experiments were replicated independently at least twice with similar results. All statistical experiments were performed in at least three biological replicates to allow for calculation of statistical significance.
All normal growing samples were selected randomly.
Blinding is not relevant to this study. All experiments were assigned into groups including relevant controls and analysis was done objectively and without bias.