Fig. 2. Histone H3 and H4 post-translational modifications at the mRARβ2 promoter. (A) Functional organization of the RARβ2 promoter. cis-acting sequences are indicated along the primary sequence of the RARβ2 promoter, as well as the main trans-acting factors known to regulate its activity. (B) ChIP assays. Soluble, formaldehyde-crosslinked chromatin was immunoprecipitated with anti-acetylated H3 (Ac-H3), anti-acetylated H4 (Ac-H4, all forms) or anti-phospho H3 (P-H3) antibodies. Input DNA (1:10 of released chromatin) and immunoprecipitated DNAs were analyzed for the presence of promoter sequences using semi-quantitative PCR. One-tenth of immunoprecipitated DNA (50–200 ng DNA) was used as a template. Mock immunoprecipitations were carried out in the absence of primary antibody (no Ab). Amplified promoter regions were (relative to the transcription initiation site): –148/+31 for RARβ, –150/+79 for H4, –243/+28 for Oct3/4 and –63/+146 for γ-actin (relative to the first codon). Additional controls for this experiment appear in Supplementary figure 1.