Cooperative binding of Oct4, Sox2, and Klf4 with stage-specific transcription factors orchestrates reprogramming [RNA-seq]

Oct4, Sox2, Klf4, and cMyc (OSKM) reprogram somatic cells to pluripotency. To gain a mechanistic understanding of their function, we mapped OSKM-binding, stage-specific transcription-factors (TFs), and chromatin-states in discrete reprogramming stages and performed loss- and gain-of-function experiments. We found that early in reprogramming OSK extensively bind somatic-enhancers and initiate their decommissioning by recruiting Hdac1. Concurrently, OSK engage other sites, including specific pluripotency-enhancers, and induce the relocation of somatic TFs to these sites and away from somatic-enhancers, extending somatic-enhancer decommissioning genome-wide. Pluripotency-enhancer selection early in reprogramming occurs predominantly at sites with high OSK-motif densities and requires collaborative binding by OSK. Most pluripotency-enhancers are selected later and occupied by OS and stage-specific-TFs like Esrrb. Overexpression of stage-specific-TFs influences reprogramming efficiency by changing OSK-occupancy, somatic-enhancer decommissioning, and pluripotency-enhancer selection. We propose that collaborative interactions among OSK and with stage-specific-TFs direct both somatic-enhancer decommissioning and pluripotency-enhancer selection, which drives the enhancer reorganization underlying reprogramming

Oct4、Sox2、Klf4和cMyc（合称OSKM）可将体细胞重编程为多能性细胞。为阐明其功能的分子机制，本研究绘制了不同离散重编程阶段下OSKM结合位点、阶段特异性转录因子（transcription factor, TF）及染色质状态（chromatin state）的图谱，并开展了功能缺失（loss-of-function）与功能获得（gain-of-function）实验。研究发现，在重编程早期，OSK可广泛结合体细胞增强子（somatic enhancer），并通过招募组蛋白去乙酰化酶1（Hdac1）启动体细胞增强子的失活过程；与此同时，OSK还结合其他位点，包括特定的多能性增强子（pluripotency enhancer），并诱导体细胞转录因子向这些位点迁移、远离体细胞增强子，从而在全基因组范围内延长体细胞增强子的失活进程。重编程早期的多能性增强子筛选主要发生在OSK基序（OSK motif）密度较高的位点，且需要OSK的协同结合才能完成；多数多能性增强子则在后期完成筛选，并由OS及Esrrb等阶段特异性转录因子占据。阶段特异性转录因子的过表达可通过改变OSK结合占据情况、体细胞增强子失活进程以及多能性增强子筛选过程，影响重编程效率。我们提出，OSK之间以及OSK与阶段特异性转录因子之间的协同相互作用，同时调控体细胞增强子失活与多能性增强子筛选过程，进而驱动重编程背后的增强子重组事件。