TGF-β1 blocks DAP10 transcription. ChIP assays. Primary human NK cells (A) or NKL cells (B) were left untreated (), treated with IL-2 (■), or treated with IL-2 plus TGF-β1 () for 3 days (primary cells) or 36 hours (NKL cells). Cells were fixed, sonicated, and immunoprecipitated with an anti-RNA pol II antibody or IgG isotype control. The amount of DAP10 DNA immunoprecipitated in each experiment was assessed by quantitative PCR using 2 different promoter-specific primer sets and calculated relative to a standard curve generated with serial dilutions of human genomic DNA. All reactions were done in triplicate and the average value of triplicate was used for calculating the relative levels of each mRNA species. Relative quantification of the target genes was made with the 2nd derivative maximum method using the Roche Lightcycler software and calculating the fold change over the 18S rRNA or actin level. Results are expressed as the mean of RNA pol II fold enrichment over isotype control antibody, normalized to values from untreated cells. For primary NK cells, a representative result from 3 separate experiments with different donors is shown. For NKL cells, n = 2, error bar represents SEM.