Fig. 4
From: EZHIP constrains Polycomb Repressive Complex 2 activity in germ cells
EZHIP mitigates PRC2-cofactors interactions. a Genome-browser representation of SUZ12 enrichment in U2OS WT, EED−/−, and EZHIP−/−. Duplicates are merged, same scale for all tracks. b Immunofluorescence staining of EED and EZHIP (top) or H3K27me2 and EZHIP (bottom) on U2OS, nuclei are stained with DAPI. Representative results. Scale bar, 2 μm. c Left, purification scheme for PRC2. Right, histone methyltransferase (HMT) assay to monitor the enzymatic activity of PRC2 purified from WT (left panel) or EZHIP−/− (middle panel) U2OS cells (titration: 1, 2, 5 × ) on native nucleosomes. Right panel, same assay as previously, but this time titrating recombinant hEZHIP on PRC2-purified from EZHIP−/− U2OS cells (PRC2 quantity 5 × ). The upper panels are autoradiography, and the lower panels are the corresponding SDS-PAGE coomassie staining. Representative image. d Quantification of EZH2-Flag IP through mass spectrometry (iBAQ values). Heatmap representing the Log2-transformed median centered values. Horizontal axis: U2OS WT and EZHIP−/−, n = 3. Vertical axes: PRC2 components. Values are normalized on iBAQ values from untagged U2OS WT and EZHIP−/−. e EZH2-Flag Co-IP from nuclear extracts either WT or EZHIP−/−, and probed with antibodies against EZH2, EED, AEBP2, or JARID2
