Single-cell landscape of nuclear configuration and gene expression during stem cell differentiation and X inactivation

Genome Biol. 2021 Sep 27;22(1):279. doi: 10.1186/s13059-021-02432-w.


Background: 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.

Results: 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.

Conclusions: 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.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Alleles
  • Animals
  • Cell Cycle
  • Cell Differentiation / genetics*
  • Cell Line
  • Cell Nucleus / genetics
  • Female
  • Gene Expression*
  • Genome
  • Male
  • Mice
  • Mouse Embryonic Stem Cells / metabolism*
  • RNA-Seq
  • Single-Cell Analysis
  • X Chromosome / chemistry
  • X Chromosome Inactivation*