Figure 5.

Assessing the interaction of Ser5P-PolII, RB1, ELF1, or MAZ with the core CXXC5 promoter elements in cellula. (a) ChIP. Fragmented chromatin from MCF7 cells processed for ChIP was immunoprecipitated with a species-specific IgG or an antibody specific to Ser5P-PolII, RB1, or ELF1. Fragmented chromatin of MCF7 cells transiently transfected with MAZ_ΔN was also processed for ChIP to ensure that the MAZ antibody directed to the carboxyl-terminus of MAZ to be used in ChIP is capable of precipitating the endogenous MAZ. ChIP precipitates were subjected to SDS-PAGE for Ser5P-PolII and RB1 or to SDS-PAGE for ELF1 and MAZ_ΔN followed by IB using antibodies indicated for ChIP. Asterisk (*) indicates the protein of interest. Input, IgG together with heavy (HC) and light (LC) chains of IgG are indicated. Molecular masses (MM) in kDa are denoted. (b) ChIP-qPCR. While identical primer sets for each antibody were used in assessing the interaction of a transcription factor (TF) to Segment A as the core CXXC5 promoter elements (CXXC5) or to the Exon2 of Myoglobin (MB) as a control, distinct primer sets were used for the promoter of a target gene of each TF. The mean ± SE of three independent experiments performed in triplicate is shown. Asterisk (*) indicates significant differences depicted as fold change compared to IgG.