Figure 3

SN-38 treatment induces binding of endogenous SXR to the native CYP3A4 promoter. A. ChIP assays were performed and DNA was further analyzed either by classical PCR or by semi-quantitative PCR using a primer set specific for the promoter or control region as indicated by the arrows on the schematic diagram of the CYP3A4 promoter. B. LS180 were treated with 10 μM of rifampicin (rif), 1 μg/ml CPT-11 or 10 ng/ml SN-38 for 4 h and subjected to formaldehyde cross-linking. Soluble chromatin was prepared by sonication. Precleared chromatin solution was immunoprecipitated by antibodies anti-SXR or Ig control, and precipitated. PCR was performed with the precipitated DNA (IP SXR) or the DNA present in 10% of total cell lysates used for each IP (input). C. Cells were treated with SN-38 for 4 h and subjected to ChIP as described above with anti-SXR, anti-RNA phospho-ARN polymerase II and anti-RNA polymerase II. Enrichment factors were determined by qPCR using GAPDH as threshold indicator (internal control for each IP).