FIG. 3.

Estrogen agonists inhibit glucocorticoid-induced chromatin remodeling. (A) Estrogen agonists inhibit glucocorticoid-induced SstI hypersensitivity. Nuclei were harvested from cells pretreated with 10 nM E2 and DES or 100 nM genistein (lanes 3 to 8) followed by treatment with 1 nM DEX (lane 2) or ER ligands (Horm) and DEX (lanes 4, 6, and 8) for 1 h. Nuclei were digested with Sst-I in vivo and with HaeIII in vitro to provide an internal control. DNA was amplified by reiterative primer extension using Taq DNA polymerase. PCR products were analyzed using 8% polyacrylamide denaturing gels and exposed to phosphorimager screens. (B) Estrogen antagonists do not inhibit glucocorticoid-induced SstI hypersensitivity. Cells were pretreated as described in the legend for panel A, but with 100 nM tamoxifen (lanes 5 and 6) or raloxifene (lanes 7 to 8). Cells treated with E2 (lanes 3 to 4) were included as a control. (C) BRG1 association with the MMTV promoter is inhibited by E2. Cells were untreated (lane 1) or pretreated with 10 nM E2 (lanes 3 and 4) or 100 nM raloxifene (lanes 5 and 6) followed by treatment with 1 nM DEX (lane 2) or ER ligands and DEX (lanes 4 and 6) for 1 h. Cells were harvested, and chromatin was prepared by sonication. The fragmented chromatin was immunoprecipitated with antibodies against BRG1 or nonspecific antibody control (normal serum IgG [NS]). DNA from the input and immunoprecipitates was analyzed by PCR with primers specific for the MMTV promoter. PCR products were analyzed with 6% nondenaturing gels and exposed to phosphorimager screens for further analysis. E, E2; D, DES; G, genistein; R, raloxifene; T, tamoxifen.