Supplementary Material for: Impact of Thyroid Hormones on Estrogen Receptor α-Dependent Transcriptional Mechanisms in Ventromedial Hypothalamus and Preoptic Area

Elevated levels of thyroid hormones (TH) reduce estradiol (E2)-dependent female sexual behavior. E2 stimulates progesterone receptor <i>(Pgr)</i> and oxytocin receptor <i>(Oxtr)</i> within the ventromedial hypothalamus and preoptic area, critical hypothalamic nuclei for sexual and maternal behavior, respectively. Here, we investigated the impact of TH on E2-dependent transcriptional mechanisms in female mice. First, we observed that triiodothyronine (T₃) inhibited the E2 induction of <i>Pgr</i> and <i>Oxtr.</i> We hypothesized that differences in histone modifications and receptor recruitment could explain the influence of TH on E2-responsive <i>Pgr</i> and <i>Oxtr</i> expression. We observed that histone H3 acetylation (H3Ac) and methylation (H3K4me3) was gene and brain-region specific. We then analyzed the recruitment of estrogen receptor α (ERα) and TH receptor α (TRα) on the putative regulatory sequences of <i>Pgr</i> and <i>Oxtr</i>. Interestingly, T₃ inhibited E2-induced ERα binding to a specific <i>Pgr</i> enhancer site, whereas TRα binding was not affected, corroborating our theory that the competitive binding of TRα to an ERα binding site can inhibit ERα transactivation and the subsequent E2-responsive gene expression. On the <i>Oxtr</i> promoter, E2 and T₃ worked together to modulate ERα and TRα binding. Finally, the E2-dependent induction of cofactors was reduced by hypothyroidism and T₃. Thus, we determined that the <i>Pgr</i> and <i>Oxtr</i> promoter regions are responsive to E2 and that T₃ interferes with the E2 regulation of <i>Pgr</i> and <i>Oxtr</i> expression by altering the recruitment of receptors to DNA and changing the availability of cofactors. Collectively, our findings provide insights into molecular mechanisms of response to E2 and TH interactions controlling sex behavior in the hypothalamus.