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. 2003 Dec 1;31(23):6710–6721. doi: 10.1093/nar/gkg913

Figure 2.

Figure 2

Metal-dependent recruitment of c-fos to the MT-I promoter requires MTF-1, whereas the metal-dependent recruitment of MTF-1 does not require c-fos. (A–C) Mouse embryonic fibroblasts (c-fos knockout (c-fos–/–), MTF-KO, or wild-type MEFs) or Hepa cells remained untreated (–) or were treated for 3 h with 100 µM zinc (Zn) or 10 µM cadmium (Cd) prior to chromatin isolation. Chromatin immunoprecipitation was carried out using antibodies against the indicated transcription factors (c-fos, c-jun, USF-2) or beads alone (no ab). Products from triplicate PCRs were quantified relative to input DNA as described in the legend to Figure 1. Similar results were obtained using zinc or cadmium [shown only for cadmium in (A)]. (C) c-fos–/– mouse embryonic fibroblasts remained untreated or were treated with cadmium (10 µM) for 3 h and ChIP was carried out using the antibodies against MTF-1 or USF-2. (D) Western blot analysis of the cellular levels of c-fos. Hepa cells cultured in DMEM containing 10% or 2% FBS as indicated, were untreated or treated for 3 h with zinc or cadmium and whole cell extracts (10 µg) were prepared. After western blotting, the membrane was stained with Ponceau’s solution to ensure equal loading and transfer of protein. ECL was carried out using the same antibody against c-fos that was used in the ChIP experiments.