Mammalian mediator structure and conformation regulation by the Tail module

The Mediator complex plays an essential and multi-faceted role in regulation of RNA polymerase II transcription in all eukaryotes. Structural analysis of yeast Mediator provided an understanding of the conserved core of the complex and its interaction with RNA polymerase II, but failed to reveal how Mediator might enable a response to interaction with transcription factors (TFs). Here we present an atomic model of mammalian (Mus musculus) Mediator, derived from a 4.0 Å resolution cryo-EM map of the complex. The mammalian Mediator structure reveals how the previously unresolved Tail module, which includes a number of metazoan specific subunits, modulates Mediator conformation and interactions. The structure also suggests how a bi-partite organization of TF-interacting subunits can be exploited to enable a Mediator mechanism that combines conformational control of molecular interactions and enhancement of transcription through phase separation and enhancer-specific recruitment Overall design: ChIP-seq in mouse CH12 cell line

中介体复合物（Mediator complex）在所有真核生物的RNA聚合酶II（RNA polymerase II）转录调控过程中，发挥着不可或缺且多维度的核心作用。此前针对酵母中介体的结构解析，仅阐明了该复合物保守核心区域的构成及其与RNA聚合酶II的相互作用模式，却未能揭示中介体如何响应转录因子（Transcription Factors, TFs）的结合并介导相关信号响应。本研究报道了源自该复合物4.0埃分辨率冷冻电镜（cryo-electron microscopy, cryo-EM）图谱的哺乳动物（小家鼠，Mus musculus）中介体原子分辨率模型。该哺乳动物中介体结构揭示了此前尚未解析的尾部模块（Tail module）如何调控中介体的构象与相互作用——该模块包含多个后生动物特异性亚基。该结构同时阐明了TF结合亚基的二分体组织模式如何被利用，从而形成兼具分子相互作用构象调控、通过相分离与增强子特异性招募实现转录增强功能的中介体作用机制。实验整体设计：在小鼠CH12细胞系中开展染色质免疫共沉淀测序（Chromatin Immunoprecipitation sequencing, ChIP-seq）实验。