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. Author manuscript; available in PMC: 2011 Aug 17.
Published in final edited form as: Dev Cell. 2010 Aug 17;19(2):220–231. doi: 10.1016/j.devcel.2010.07.007

Figure 3. β-catenin associates with a Histone H3(R8) methyltransferase before the MBT.

Figure 3

(A) β-catenin was immunoprecipitated from 16-cell embryos and HMT activity was visualized by fluorography (1 day exposure) for incorporation of [3H] methyl groups into calf thymus histones (top panel). Equal loading of histones is shown by coomassie staining (lower panel). “Input” represents HMT activity in embryo lysates, with activity toward both H3 and H4. (B) β-catenin IP/HMT assays were performed on either wild type (WT) recombinant H3.3 or H3.3 with the indicated point mutations. (C) β-catenin IP/HMT assays were performed on peptides corresponding to unmodified H3 (aa 1-15, lanes 1&2), asymmetrically dimethylated R8 (aa 1-15, lane 3), unmodified H3 (aa 1-21, lanes 4&5), acetylated K9 (aa 1-20, lane 6) and trimethylated K9 (aa 1-24, lane 7). (D) MBT-stage control and lithium chloride treated (LiCl, 300mM for 10 minutes, 1 hour prior to harvest) embryos were subjected to ChIP for either H3K9me1 or H3K9me3. (E) ChIP was performed as described in panel D, using instead antibodies to either H3R8me2a or H3R8me2s (Pal et al., 2004). (F) ChIP was performed on 1000-cell stage control and β-catenin knockdown (β-MO) embryos with the Active Motif H3R8me2a antiserum. See also Figure S2.