Testing the effect of the ER expression level on the ER-dependent transcriptional activation. (A) Experimental testing of the model predictions about the non-monotonous dependence of ER -induced transcriptional activation on the level of ER expression. mRNA expression levels of TFF1, PGR, MYC and CTSD in four experimental cell lines were measured by real-time RT-PCR using specific primer pairs. RNA was collected at 48 h and was extracted from either untreated (control) cells or cells treated with 1 nM 17β- estradiol (E2), 1 μM tamoxifen (Tam), and a combination of 1 nM E2 and 1 μM tamoxifen (E2+Tam). mRNA levels are given in arbitrary units. (B) Relative expression levels of ERα in wild type MCF-7, LCC1, LCC2 and LCC9 cell lines; (C) Theoretical expression levels of a hypothetical estrogene responsive gene, calculated as p̅ (see equations 22–24 in Methods) for four different total concentrations of ER: ERt=10; 40; 70 and 100 nM. The parameters of transcriptional regulation were set equal to those presented in Tables 1 and 2; Hormone (Ht) and tamoxifen (It) concentrations were 1 nM and 100 nM respectively. The ratio of chosen theoretical ER levels roughly corresponded to the ratio of ER expression levels in four experimental cell lines.