Target Gene Repression Based on Dismissal of Polymerase II from Estrogen Receptor Trans-bound Enhancers Is Associated With Clinical Outcome in Human Breast Cancer [GRO-seq]

We find that 17-ß-estradiol (E2)-bound estrogen receptor a (ERa) is bound in trans to a cohort of FOXA1-dependent, constitutively activate enhancers, inactivating these enhancers by decommissioning/removing enhancer Polymerase II (Pol II), despite recruitment of coactivators. This is based on the surprising recruitment by the ERa DNA binding domain of the histone demethylase, KDM2A, which, functioning independently of its demethylase function. KDM2A mediates recruitment of NEDD4 complexes that ubiquitinate and dismisses Pol II from these "repressive" enhancers, resulting in the E2 down-regulated transcriptional program. Overall design: Trans-bound estrogen receptor a (ERa) to the E2 repressed enhancers are proved by ERa pbox mutated ChIP-seq. ERa DNA binding domain interacts with histone demethylase, KDM2A, which mediates recruitment of NEDD4 complexes that dismisses Pol II from these "repressive" enhancers, resulting in the E2 down-regulated transcriptional program shown by GRO-seq and Pol II ChIP-seq. Overall, our data suggest that KDM2A plays central roles in a nuclear receptor-induced transcriptional repression program.

我们发现，结合17-β-雌二醇（17-β-estradiol, E2）的雌激素受体α（estrogen receptor α, ERα）可反式结合于一组依赖于叉头框蛋白A1（FOXA1）的组成型活化增强子；尽管该复合物招募了辅激活因子（coactivators），却通过停用或移除增强子处的RNA聚合酶II（Polymerase II, Pol II）使这些增强子失活。这一现象的机制基础是ERα的DNA结合域（DNA binding domain）非预期地招募了组蛋白去甲基化酶（histone demethylase）KDM2A，且KDM2A的功能并不依赖于其自身的去甲基化酶活性。KDM2A可介导招募NEDD4复合物（NEDD4 complexes），该复合物可对这些"抑制性"增强子（enhancers）处的Pol II进行泛素化（ubiquitinate）并将其解离，最终引发E2下调的转录程序。
整体实验设计：通过ERα pbox突变的染色质免疫共沉淀测序（Chromatin Immunoprecipitation Sequencing, ChIP-seq）验证了可反式结合于E2抑制性增强子的ERα。ERα的DNA结合域与组蛋白去甲基化酶KDM2A相互作用，KDM2A介导招募NEDD4复合物以将这些"抑制性"增强子处的Pol II解离，这一结论可通过全局运行转录测序（Global Run-On Sequencing, GRO-seq）及Pol II ChIP-seq得到证实。综上，我们的数据表明KDM2A在核受体（nuclear receptor）诱导的转录抑制程序中发挥核心作用。