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. 2009 Jun 5;10(7):776–782. doi: 10.1038/embor.2009.88

Figure 3.

Figure 3

MC1568 blocks MEF2D transcriptional activity in C2C12 cells and shows inhibitory activities in mice. (A) Histone deacetylase (HDAC) assay on myocyte enhancer factor 2D (MEF2D) immunoprecipitation (IP) from C2C12 cells in the presence of MC1568 (5 μM) with or without valproic acid (VPA; 1 mM); the inset shows the quantities of co-immunoprecipitated MEF2D, HDAC4 and HDAC3. (B) Chromatin immunoprecipitation (ChIP) assay of acetyl-H3 (AcH3) levels on the myogenin promoter in C2C12 cells. (C) ChIP assay of MEF2D on the myogenin (Myo) and muscle creatine kinase (MCK) promoters in C2C12 cells. (D) MC1568 increases acetylation of tubulin (AcTub) in selected organs of mice in a dose-dependent manner. (E) HDAC4 and HDAC5 expression levels in skeletal muscle and heart after MC1568 treatment (50 mg/kg); note that although differentially expressed both HDACs are present. Bottom: IPs using HDAC4 and HDAC5 antibodies to reveal MEF2D and HDAC3 complexes in skeletal muscle and heart. (F) Heart-extract-HDAC5 immunoprecipitation assay on treatment with 1, 10 and 50 mg/kg MC1568. (G) HDAC assay from heart extracts on MEF2D IP in the presence of MC1568 with or without 1 mM VPA. DM, differentiation medium; DPM, disintegrations per minute; ERK, extracellular signal regulated kinase.