A homozygous missense mutation in the gene encoding the estrogen receptor α (ERα) was previously identified in a female patient with estrogen insensitivity syndrome. We investigated the molecular features underlying the impaired transcriptional response of this mutant (ERα-Q375H) and four other missense mutations at this position designed to query alternative mechanisms. The identity of residue 375 greatly affected the sensitivity of the receptor to agonists without changing the ligand binding affinity. Instead, the mutations caused changes in the affinity of coactivator binding and alterations in the balance of coactivator and corepressor recruitment. Comparisons among the transcriptional regulatory responses of these six ERα genotypes to a set of ER agonists showed that both steric and electrostatic factors contributed to the functional deficits in gene regulatory activity of the mutant ERα proteins. ERα–coregulator peptide binding in vitro and RIME (rapid immunoprecipitation mass spectrometry of endogenous) analysis in cells showed that the degree of functional impairment paralleled changes in receptor-coregulator binding interactions. These findings uncover coupling between ligand binding and coregulator recruitment that affects the potency rather than the efficacy of the receptor response without substantially altering ligand binding affinity. This highlights a molecular mechanism for estrogen insensitivity syndrome involving mutations that perturb a bidirectional allosteric coupling between ligand binding and coregulator binding that determines receptor transcriptional output.

先前在一名患有雌激素不敏感综合征的女性患者中发现了编码雌激素受体α (ERα) 的基因中的纯合错义突变。我们研究了该突变体（ERα-Q375H）和该位置的其他四个错义突变的转录反应受损的分子特征，旨在查询替代机制。残基 375 的身份极大地影响了受体对激动剂的敏感性，而不会改变配体结合亲和力。相反，突变导致共激活因子结合亲和力的变化以及共激活因子和辅抑制因子募集平衡的改变。这六种 ERα 基因型对一组 ER 激动剂的转录调节反应之间的比较表明，空间和静电因素都导致突变 ERα 蛋白基因调节活性的功能缺陷。体外 ERα-调节剂肽结合和细胞中的 RIME（内源性快速免疫沉淀质谱）分析表明，功能障碍的程度与受体-调节剂结合相互作用的变化平行。这些发现揭示了配体结合和共调节因子募集之间的耦合，这会影响受体反应的效力而不是功效，而不会显着改变配体结合亲和力。