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. Author manuscript; available in PMC: 2014 Jul 24.
Published in final edited form as: Nat Commun. 2014;5:3127. doi: 10.1038/ncomms4127

Figure 3. Anti-EED (aa429-441) antibody disrupts PRC2 and attenuates its histone methyltransferase activity.

Figure 3

a, Anti-EED (aa429-441) antibody inhibits the EED: EZH2 interaction. Halo-tagged full-length EED protein was incubated with EZH2 in the presence of different amounts of control IgG, anti-EED (aa429-441) and anti-EED (aa186-304) antibodies. Halo-Link PD (pull down) followed by immunoblot analysis is shown. b, Anti-EED (aa429-441) antibody inhibits PRC2 mediated methylation of histone H3 in vitro. Indicated amounts of purified PRC2, control IgG, anti-EED (aa429-441) and anti-EED (aa186-304) antibodies were incubated with recombinant histone H3 with unmethylated H3K27 for two hours. Immunoblot analyses with anti-H3K27me3 and anti-H3K27me2 antibodies were then performed. c, Anti-EED (aa429-441) antibody inhibits PRC2 mediated methylation of nucleosomes in vitro. Indicated amounts of purified PRC2, control IgG, anti-EED (aa429-441) and anti-EED (aa186-304) antibodies were incubated with nucleosomes containing unmethylated H3K27. ELISA with anti-H3K27me3 antibody was then performed to measure PRC2 histone methyltransferase activity. Assays were repeated three times in triplicate. All error bars represent ±s.e.m.