Pioneer factor Pax7 initiates two-step cell-cycle dependent chromatin opening (ATAC-seq). Pioneer factor Pax7 initiates two-step cell-cycle dependent chromatin opening (ATAC-seq)

Pioneer transcription factors direct cell differentiation by deploying new enhancer repertoires through their unique ability to target and initiate remodelling of closed chromatin. The initial steps of their action remain undefined although pioneers were shown to interact with nucleosomal target DNA and with some chromatin remodelling complexes. We now define the sequence of events that enable the pioneer Pax7 with its unique abilities. Chromatin condensation exerted by linker histone H1 is the first constraint on Pax7 recruitment, and this establishes the initial speed of chromatin remodelling. The first step of pioneer action involves recruitment of the KDM1A (Lsd1) H3K9me2 demethylase for removal of this repressive mark, as well as recruitment of the MLL complex for deposition of the activating H3K4me1 mark. Further progression of pioneer action requires passage through cell division, and this involves dissociation of pioneer targets from perinuclear lamin B. Only then, the SWI/SNF remodeling complex and the coactivator p300 are recruited, leading to nucleosome displacement and enhancer activation. Thus, the unique features of pioneer actions are those occurring in the lamin-associated compartment of the nucleus. This model is consistent with prior work that showed a dependence on cell division for establishment of new cell fates. Overall design: ATAC-seq in mouse corticotrope AtT-20 cells with Pax7 or ER-Pax7 ectopic expression. Treated with tamoxifen (tam, ER-Pax7 activation) or mimosine (mim, cell cycle arrest).

先锋转录因子（Pioneer transcription factors）凭借其靶向并启动封闭染色质重塑的独特能力，调控全新增强子组的部署，从而指导细胞分化。尽管已有研究证实先锋因子可与核小体靶DNA及部分染色质重塑复合物结合，但其作用的初始步骤仍未明确。本研究明确了具备独特功能的先锋因子Pax7发挥作用的事件时序：连接组蛋白H1介导的染色质浓缩是Pax7招募的首个限制因素，并决定了染色质重塑的初始速率。先锋因子作用的第一步，是招募KDM1A（Lsd1）H3K9me2去甲基化酶以清除该抑制性修饰，同时招募MLL复合物以沉积激活性H3K4me1修饰。先锋因子作用的后续推进则依赖细胞周期分裂，此过程涉及先锋因子靶位点从核周核纤层蛋白B上解离。完成上述步骤后，方可招募SWI/SNF重塑复合物与共激活因子p300，进而引发核小体移位与增强子激活。由此可见，先锋因子作用的独特特征均发生于细胞核的核纤层相关区域。该模型与此前揭示的“新细胞命运建立依赖细胞分裂”的研究结果一致。
实验设计方案：对异位表达Pax7或ER-Pax7的小鼠促肾上腺皮质细胞AtT-20进行转座酶可及性测序（ATAC-seq）。实验分组分别经他莫昔芬（tam，用于激活ER-Pax7）或含羞草氨酸（mim，用于阻断细胞周期）处理。