Mammalian development is associated with extensive changes in gene expression, chromatin accessibility, and nuclear structure. Here, we follow such changes associated with mouse embryonic stem cell differentiation and X inactivation by integrating, for the first time, allele-specific data from these three modalities obtained by high-throughput single-cell RNA-seq, ATAC-seq, and Hi-C. Allele-specific contact decay profiles obtained by single-cell Hi-C clearly show that the inactive X chromosome has a unique profile in differentiated cells that have undergone X inactivation. Loss of this inactive X-specific structure at mitosis is followed by its reappearance during the cell cycle, suggesting a “bookmark” mechanism. Differentiation of embryonic stem cells to follow the onset of X inactivation is associated with changes in contact decay profiles that occur in parallel on both the X chromosomes and autosomes. Single-cell RNA-seq and ATAC-seq show evidence of a delay in female versus male cells, due to the presence of two active X chromosomes at early stages of differentiation. The onset of the inactive X-specific structure in single cells occurs later than gene silencing, consistent with the idea that chromatin compaction is a late event of X inactivation. Single-cell Hi-C highlights evidence of discrete changes in nuclear structure characterized by the acquisition of very long-range contacts throughout the nucleus. Novel computational approaches allow for the effective alignment of single-cell gene expression, chromatin accessibility, and 3D chromosome structure. Based on trajectory analyses, three distinct nuclear structure states are detected reflecting discrete and profound simultaneous changes not only to the structure of the X chromosomes, but also to that of autosomes during differentiation. Our study reveals that long-range structural changes to chromosomes appear as discrete events, unlike progressive changes in gene expression and chromatin accessibility.

中文翻译：

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干细胞分化和 X 失活过程中核构型和基因表达的单细胞景观


哺乳动物的发育与基因表达、染色质可及性和核结构的广泛变化有关。在这里，我们通过首次整合通过高通量单细胞 RNA-seq、ATAC-seq 和 Hi 获得的这三种方式的等位基因特异性数据来跟踪与小鼠胚胎干细胞分化和 X 失活相关的变化。 -C。单细胞 Hi-C 获得的等位基因特异性接触衰减谱清楚地表明，失活的 X 染色体在经历 X 失活的分化细胞中具有独特的谱。这种非活性 X 特异性结构在有丝分裂时丢失，随后在细胞周期中重新出现，这表明存在“书签”机制。 X 失活发生后胚胎干细胞的分化与 X 染色体和常染色体上同时发生的接触衰变谱的变化有关。单细胞 RNA-seq 和 ATAC-seq 显示雌性细胞相对于雄性细胞存在延迟的证据，这是由于在分化早期阶段存在两条活性 X 染色体。单细胞中失活的 X 特异性结构的出现晚于基因沉默，这与染色质压缩是 X 失活的晚期事件的观点一致。单细胞 Hi-C 强调了核结构离散变化的证据，其特征是在整个细胞核中获得非常长距离的接触。新颖的计算方法可以有效对齐单细胞基因表达、染色质可及性和 3D 染色体结构。 基于轨迹分析，检测到三种不同的核结构状态，不仅反映了 X 染色体结构的离散且深刻的同时变化，而且还反映了分化过程中常染色体结构的变化。我们的研究表明，染色体的长期结构变化表现为离散事件，与基因表达和染色质可及性的渐进变化不同。