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Fig. 2 | BMC Biology

Fig. 2

From: Histone variants H2A.Z and H3.3 coordinately regulate PRC2-dependent H3K27me3 deposition and gene expression regulation in mES cells

Fig. 2

Promotion of PRC2 enzymatic activity by H2A.Z through facilitating chromatin compaction. a PRC2 histone methyltransferase activity in vitro (HMT assay) on the nucleosomal substrates containing a different combination of histone variants (H2A/H3.1, H2A.Z/H3.1, H2A/H3.3, and H2A.Z/H3.3). Purified PRC2 complex and isotope-labeled S-adenosylmethionine (3H-SAM) were applied in HMT assay. PRC2 activity was measured by auto-chromatography and a liquid scintillation counter. The relative activities of PRC2 on histone variants-containing nucleosomes were normalized with the PRC2 activity on the canonical nucleosome (H2A/H3.1-nucleosome). b Sequence alignment between amino acid sequences of H2A and H2A.Z, divergent residues in extended acid patch domain are highlighted with a black box. c Analytical ultracentrifuge assay (AUC) to monitor the effect of mutations at the extended acid patch of H2A.Z (S99K, D98N, and D98N/S99K) on the H2A.Z-enhanced chromatin compaction. d HMT assays to monitor the effect of mutations at the extended acid patch (S99K, D98N, and D98N/S99K) on the H2A.Z-enhanced PRC2 activities. e HMT assay to monitor the effect of deletion of histone H4 N-terminal domain (NTD) on H2A.Z-enhanced PRC2 enzymatic activity. f HMT assay to monitor the effect of mutations on H3.1 (V89M/M90G and A31S/S87A) on H2A.Z-enhanced PRC2 enzymatic activity

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