Laboratory for Developmental Epigenetics, RIKEN Center for Biosystems Dynamics Research (BDR), Kobe, Japan.
RIKEN Center for Developmental Biology (CDB), Kobe, Japan.
Nat Genet. 2019 Mar;51(3):529-540. doi: 10.1038/s41588-019-0347-5. Epub 2019 Feb 25.
Here, we report a single-cell DNA replication sequencing method, scRepli-seq, a genome-wide methodology that measures copy number differences between replicated and unreplicated DNA. Using scRepli-seq, we demonstrate that replication-domain organization is conserved among individual mouse embryonic stem cells (mESCs). Differentiated mESCs exhibited distinct profiles, which were also conserved among cells. Haplotype-resolved scRepli-seq revealed similar replication profiles of homologous autosomes, while the inactive X chromosome was clearly replicated later than its active counterpart. However, a small degree of cell-to-cell replication-timing heterogeneity was present, which was smallest at the beginning and the end of S phase. In addition, developmentally regulated domains were found to deviate from others and showed a higher degree of heterogeneity, thus suggesting a link to developmental plasticity. Moreover, allelic expression imbalance was found to strongly associate with replication-timing asynchrony. Our results form a foundation for single-cell-level understanding of DNA replication regulation and provide insights into three-dimensional genome organization.
在这里，我们报告了一种单细胞 DNA 复制测序方法 scRepli-seq，这是一种测量复制和未复制 DNA 之间拷贝数差异的全基因组方法。使用 scRepli-seq，我们证明了复制域组织在单个小鼠胚胎干细胞（mESC）中是保守的。分化的 mESC 表现出不同的特征，这些特征在细胞之间也是保守的。单倍型解析 scRepli-seq 揭示了同源常染色体的相似复制特征，而失活的 X 染色体明显比其活性对应物复制得更晚。然而，存在微小的细胞间复制定时异质性，在 S 期的开始和结束时最小。此外，发育调控域与其他域分离，表现出更高程度的异质性，因此表明与发育可塑性有关。此外，等位基因表达失衡与复制定时的异步性强烈相关。我们的研究结果为单细胞水平的 DNA 复制调控提供了基础，并为三维基因组组织提供了新的见解。
