Fig. 8

DNA fragment bound by the ternary complex including Tax-A is more abundant than that including Tax-B in the CXCL10 promoter. To investigate whether HTLV-1 subgroups affects the binding of Tax to the endogenous CXCL10 promoter, we performed chromatin immunoprecipitation (ChIP) assays. Chromatin fragments were prepared from Jurkat T-cells transfected with plasmid expressing Tax and immunoprecipitated with specific anti-Tax monoclonal antibody (Lt-4). The immunoprecipitated DNA was then amplified by PCR. a Schematic representation of the CXCL10 promoter sequence. “Target region”, which was amplified by PCR, includes two NF-κB sites and an AP-1 site. b, c Chromatin immunoprecipitation (ChIP) assays. Representative examples of PCR products obtained from chromatin material that was directly immunoprecipitated with anti-Tax monoclonal antibody (Lt-4) (b). The ternary protein complex including Tax was found to be associated with the CXCL10 proximal promoter sequence including the two NF-κB sites and an AP-1 site. The abundance of a specific DNA fragment bound by the ternary complex, including Tax, is higher for Tax-A than for Tax-B. Cell lysates were subjected to ChIP assays using Lt-4 antibody followed by qRT-PCR using the specific primer sets (c). Three independent experiments were performed. Data shown as mean ± SD