The same strand of B1-X35S is transcribed by Pol III and Pol II. (A) IVT of B1-X35S analyzed using direction-specific sense and antisense with extracts of Hepa-1 cells treated with or without 3 μg/mL α-AMA. A negative qPCR control was performed in the absence of RT template (−RT). (B) Binding of Pol III and Pol II on B1-X35S was determined by re-ChIP. First ChIP used anti-Pol III antibody, and the resulting DNA was immunoprecipitated again with anti-Pol II or anti-TFIIIC (positive control) antibodies. Re-ChIP for GAPDH was used as negative control. Data are presented as percentage of input from the first ChIP. (C) Pol III binding in vitro to B1-X35S A+B mutant was determined by DNA affinity assays with anti-Pol III RPC32 antibody. (D) EBA performed as for Figure 1A using wild-type B1-X35S and B1-X35S A+B mutant with or without AHR transfection. (B) Average data from five B1-X35S containing genes (Dad1, Lpp, Cabin1, Tbc1d1, and Rtl1). Data are shown as mean ±SD.
