Figure 6.

c-Jun is recruited to target gene promoters. Panel A, MCF-7 cells were hormone deprived and treated with 10⁻⁶m CdCl₂ (Cd) or mock treated [control (C)]. ChIP analysis was performed with α-c-Jun antibodies, and c-Jun recruitment to target gene promoters was determined by promoter-specific primers using semiquantitative RT-PCR. Panel B, Band intensities of PCR product were quantitated using Quantity One (Bio-Rad). Panels C and D, ChIP re-ChIP was used to determine whether ERα and c-Jun are part of the same promoter complex. ChIP assay was performed as in A with antibodies against ERα or c-Jun. Protein-DNA complexes of the first ChIP were extracted and re-ChIPed with the second antibody. Normal rabbit serum was used as a negative control. The occupancy of ERα/c-Jun complexes on target promoters was analyzed using promoter-specific primers and semiquantitative RT-PCR. Band intensities of PCR products were quantitated using Quantity One (Bio-Rad).