Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA.
Harvard Stem Cell Institute, Cambridge, Massachusetts 02138, USA.
Nature. 2015 Feb 19;518(7539):355-359. doi: 10.1038/nature13990. Epub 2014 Dec 24.
Models derived from human pluripotent stem cells that accurately recapitulate neural development in vitro and allow for the generation of specific neuronal subtypes are of major interest to the stem cell and biomedical community. Notch signalling, particularly through the Notch effector HES5, is a major pathway critical for the onset and maintenance of neural progenitor cells in the embryonic and adult nervous system. Here we report the transcriptional and epigenomic analysis of six consecutive neural progenitor cell stages derived from a HES5::eGFP reporter human embryonic stem cell line. Using this system, we aimed to model cell-fate decisions including specification, expansion and patterning during the ontogeny of cortical neural stem and progenitor cells. In order to dissect regulatory mechanisms that orchestrate the stage-specific differentiation process, we developed a computational framework to infer key regulators of each cell-state transition based on the progressive remodelling of the epigenetic landscape and then validated these through a pooled short hairpin RNA screen. We were also able to refine our previous observations on epigenetic priming at transcription factor binding sites and suggest here that they are mediated by combinations of core and stage-specific factors. Taken together, we demonstrate the utility of our system and outline a general framework, not limited to the context of the neural lineage, to dissect regulatory circuits of differentiation.
从人类多能干细胞中衍生而来的能够在体外准确再现神经发育并允许生成特定神经元亚型的模型，是干细胞和生物医学领域的主要关注点。Notch 信号通路，特别是通过 Notch 效应物 HES5，是胚胎和成年神经系统中神经祖细胞起始和维持的关键途径。在这里，我们报告了从 HES5::eGFP 报告人人类胚胎干细胞系中衍生的六个连续神经祖细胞阶段的转录组和表观基因组分析。使用该系统，我们旨在模拟包括皮质神经干/祖细胞发生过程中的特化、扩增和模式形成在内的细胞命运决定。为了解剖协调特定于阶段的分化过程的调控机制，我们开发了一种计算框架，基于表观遗传景观的渐进式重塑，推断每个细胞状态转变的关键调节因子，然后通过汇集短发夹 RNA 筛选进行验证。我们还能够改进我们之前关于转录因子结合位点上表观遗传引发的观察结果，并在此建议它们是由核心和阶段特异性因子的组合介导的。总之，我们证明了我们的系统的实用性，并概述了一个通用框架，不仅限于神经谱系的背景，以剖析分化的调控回路。
