Molecular basis of neuronal subtype bias introduced by proneural factors Ascl1 and Neurog2 (ATAC-seq). Molecular basis of neuronal subtype bias introduced by proneural factors Ascl1 and Neurog2 (ATAC-seq)

Basic helix-loop-helix (bHLH) proneural transcription factors (TFs) Ascl1 and Neurog2 are integral to the development of the nervous system. Here, we investigated the molecular mechanisms by which Ascl1 and Neurog2 control the acquisition of generic neuronal fate and impose neuronal subtype identity. Using direct neuronal programming of embryonic stem cells, we found that Ascl1 and Neurog2 regulate distinct targets by binding to largely different sets of sites. Their divergent binding pattern is not determined by the previous chromatin state but distinguished by specific E-box enrichments which reflect the DNA sequence preference of the bHLH domain. The divergent Ascl1 and Neurog2 binding patterns result in distinct chromatin accessibility and enhancer activity landscapes that shape the binding and activity of downstream TFs during neuronal specification. Our findings suggest that proneural factors contribute to neuronal diversity by differentially altering the chromatin landscapes that shape the binding of neuronally expressed TFs. Overall design: ATAC-seq was used to profile chromatin accessibility upon overexpression of Ascl1 and Neurog2 in mouse embryonic stem cells. t=0 (uninduced) chromatin was used as a control.

碱性螺旋-环-螺旋（basic helix-loop-helix, bHLH）型成神经转录因子（transcription factors, TFs）Ascl1与Neurog2在神经系统发育过程中发挥关键作用。本研究探究了Ascl1与Neurog2调控通用神经元命运获得并赋予神经元亚型身份的分子机制。借助胚胎干细胞直接神经元重编程技术，我们发现Ascl1与Neurog2可通过结合截然不同的位点集合来调控不同的靶基因。二者的差异化结合模式并非由预存染色质状态决定，而是由特异性富集的E盒（E-box）所区分，这一特征反映了bHLH结构域的DNA序列偏好性。Ascl1与Neurog2的差异化结合模式会造就截然不同的染色质可及性与增强子活性图谱，进而在神经元特化过程中调控下游转录因子（TFs）的结合与活性。本研究结果表明，成神经转录因子可通过差异化重塑调控神经元表达转录因子结合的染色质图谱，从而促进神经元多样性的形成。整体实验设计：采用转座酶可及性染色质测序（ATAC-seq）技术，对小鼠胚胎干细胞中过表达Ascl1与Neurog2后的染色质可及性进行分析，以未诱导（t=0）的染色质样本作为对照。