Identification of DNA methylation changes at <i>cis</i>-regulatory elements during early steps of HSC differentiation using tagmentation-based whole genome bisulfite sequencing

Epigenetic alterations during cellular differentiation are a key molecular mechanism which both instructs and reinforces the process of lineage commitment. Within the haematopoietic system, progressive changes in the DNA methylome of haematopoietic stem cells (HSCs) are essential for the effective production of mature blood cells. Inhibition or loss of function of the cellular DNA methylation machinery has been shown to lead to a severe perturbation in blood production and is also an important driver of malignant transformation. HSCs constitute a very rare cell population in the bone marrow, capable of life-long self-renewal and multi-lineage differentiation. The low abundance of HSCs has been a major technological barrier to the global analysis of the CpG methylation status within both HSCs and their immediate progeny, the multipotent progenitors (MPPs). Within this Extra View article, we review the current understanding of how the DNA methylome regulates normal and malignant hematopoiesis. We also discuss the current methodologies that are available for interrogating the DNA methylation status of HSCs and MPPs and describe a new data set that was generated using tagmentation-based whole genome bisulfite sequencing (TWGBS) in order to comprehensively map methylated cytosines using the limited amount of genomic DNA that can be harvested from rare cell populations. Extended analysis of this data set clearly demonstrates the added value of genome-wide sequencing of methylated cytosines and identifies novel important <i>cis</i>-acting regulatory regions that are dynamically remodeled during the first steps of haematopoietic differentiation.细胞分化过程中的表观遗传改变，是指导并强化谱系定型进程的关键分子机制。在造血系统中，造血干细胞（hematopoietic stem cells, HSCs）的DNA甲基化组发生的渐进性改变，对于高效生成成熟血细胞至关重要。研究证实，细胞DNA甲基化系统的功能抑制或缺失，不仅会严重扰乱造血过程，同时也是恶性转化的重要驱动因素。 造血干细胞在骨髓中含量极为稀少，却具备终生自我更新与多谱系分化的能力。造血干细胞的低丰度，长期以来是对造血干细胞及其直接子代——多能祖细胞（multipotent progenitors, MPPs）的CpG甲基化状态进行全基因组分析的主要技术障碍。 在本篇特稿文章中，我们综述了当前学界对于DNA甲基化组调控正常造血与恶性造血的认知现状。此外，我们还探讨了当前可用于分析造血干细胞与多能祖细胞DNA甲基化状态的现有方法，并介绍了一项基于转座酶介导的全基因组亚硫酸氢盐测序（tagmentation-based whole genome bisulfite sequencing, TWGBS）构建的全新数据集：该技术可利用从稀有细胞群体中获取的微量基因组DNA，实现甲基化胞嘧啶的全基因组定位。 对该数据集的扩展分析，清晰证实了甲基化胞嘧啶全基因组测序的附加价值，并鉴定出在造血分化初始阶段发生动态重塑的新型关键顺式作用调控区域。