Robust regulatory interplay of enhancers, facilitators, and promoters in a native chromatin context [ChIP-Seq]

Enhancers are gene-distal cis-regulatory elements that drive cell type-specific gene expression. While significant progress has been made in identifying enhancers and characterizing their epigenomic features, much less effort has been devoted to elucidating mechanistic interactions among clusters of functionally linked regulatory elements within their endogenous chromatin contexts. Here, we developed a novel recombinase-mediated genome rewriting platform and applied our divergent transcription architectural model to understand how a long-range human enhancer confers a remarkable 10,000-fold activation to its target gene, NMU, at its native locus. Our systematic dissection reveals transcription factor synergy at this enhancer and highlights the interplay between a divergently transcribed core enhancer unit and emerging new types of cis-regulatory elements—notably, intrinsically inactive facilitators that augment and buffer core enhancer activity, and an adjacent retroviral long terminal repeat promoter that represses enhancer activity. We discuss the broader implications of our focused study on enhancer mechanisms and regulation genome-wide. Chromatin immunoprecipitation sequencing (ChIP-seq) targeting p300 for recombinant eNMU motif mutants; two biological replicates per genotype

增强子（enhancers）是一类位于基因远端的顺式调控元件（cis-regulatory elements），可介导细胞类型特异性的基因表达。尽管目前在增强子的鉴定及其表观基因组特征解析领域已取得长足进展，但针对内源性染色质环境中功能关联的调控元件簇之间的机制性相互作用的解析，相关研究仍相对不足。本研究开发了一种新型重组酶介导的基因组重写平台，并应用双向转录结构模型，旨在解析人类远端增强子如何在其内源位点对靶基因NMU实现高达10000倍的转录激活。我们的系统性解析揭示了该增强子区域的转录因子协同作用，并凸显了双向转录核心增强子单元与新型顺式调控元件之间的相互调控关系：具体包括可增强并缓冲核心增强子活性的本征无活性辅助因子，以及毗邻的、可抑制增强子活性的逆转录病毒长末端重复序列启动子。本研究还探讨了这项聚焦性研究对全基因组范围内增强子机制与调控研究的广泛启示。本研究的实验数据集包括：针对重组eNMU基序突变体的p300靶向染色质免疫共沉淀测序（ChIP-seq），每种基因型设置两次生物学重复。