Nucleosome dynamics render heterochromatin accessible in living human cells

Introduction

Chromatin consists of repeating units called nucleosomes, which contain ~147 bp of DNA coiled around a central histone octamer core. The octamer is composed of two molecules each of H2A, H2B, H3 and H4¹. Nucleosomes are regularly spaced on the DNA, resembling beads on a string. This nucleosomal filament undergoes additional compaction to form euchromatin or heterochromatin. In mammalian cells, heterochromatin may be constitutive, involving the same genomic regions in all cells (typically gene-poor regions composed of short repeated sequences), such as pericentromeric and telomeric regions, or facultative, involving cell type-specific fully repressed genes^{2, 3, 4, 5, 6, 7, 8, 9, 10, 11–12}. Heterochromatin is associated with specific post-translational histone modifications: constitutive heterochromatin is marked by H3K9me2/3, whereas facultative heterochromatin is marked by H3K27me3, although there is some overlap¹³. Conversely, other histone marks, such as H3K4me1, H3K4me3, H3K36me3 and H3K27ac, are generally associated with euchromatin.

The highly condensed nature of heterochromatin suggests that access to the DNA may be limited or even prevented. However, large proteins and dextrans can penetrate heterochromatin domains to some extent when injected into living cells, suggesting that heterochromatin may be accessible¹⁴. Furthermore, heterochromatin protein 1 (HP1), which binds to H3K9me3 in constitutive heterochromatin, is mobile in living mammalian cells^15,16 and transcription of repeat sequences in constitutive heterochromatin occurs at low levels^7,17. These data indicate that constitutive heterochromatin is at least partially accessible some of the time. Liquid-liquid phase separation may also be important in constitutive heterochromatin, resulting in exclusion of specific proteins from the heterochromatin phase^{10,12,18, 19, 20–21}. These studies have led to a more nuanced view concerning the accessibility of constitutive heterochromatin.

Facultative heterochromatin contains inactive genes that are subject to Polycomb-mediated repression and are marked by H3K27me3 (reviewed by ref. ¹¹). Genome-wide MNase-seq and ATAC-seq studies on isolated nuclei from various organisms have shown that inactive genes lack nucleosome-depleted regions (NDRs) at their promoters, unlike active genes. This observation suggests that nucleosomes prevent transcription factor binding at regulatory elements, such as promoters and enhancers, resulting in repression^{22, 23, 24, 25–26}. However, inactive promoters are partially accessible in mouse liver cell nuclei²⁷. Although most transcription factors cannot access their cognate binding...