Chromatin structure of the inactive X chromosome revealed by in situ cryo-electron tomography

Abstract

The intricate organisation of chromatin in eukaryotic cells plays a central role in regulating gene transcription and other DNA-templated processes. Prior studies using light microscopy, biochemistry, and genomics have implicated histone and DNA modifications, histone variants, chromosomal proteins, non-coding RNA, and intrinsic biophysical properties of chromatin in determining chromatin organisation across multiple scales. In this study, we apply an innovative approach, direct visualisation of native chromatin structure and its modulation by different factors using cryo-electron tomography, exploiting the inactive X chromosome (Xi) in differentiating mouse embryonic stem cells as a model system of facultative heterochromatin. We show that Xi chromatin undergoes progressive compaction across scales, from individual nucleosomes to chromatin domains. We observe an enrichment of histone H1-bound chromatosomes that are distributed to form the core of Xi chromatin domains. Moreover, we demonstrate that histone deacetylation demarcates chromatin domains. Specifically, perturbations leading to histone acetylation in the Xi dissolve discrete chromatin-domain boundaries, resulting in a homogeneous chromatin distribution with a dense and uniform packing of nucleosomes. Together, these findings illuminate the multiscale organisation of native facultative heterochromatin and the contributions of histone H1 and histone acetylation to this organisation. This study opens a new avenue for investigating how chromatin modification and structure relate to gene activity in near-native cells.