Up to seven members of the histone H1 family may contribute to chromatin compaction and its regulation in human somatic cells. In breast cancer cells, knock-down of multiple H1 variants deregulates many genes, promotes the appearance of genome-wide accessibility sites and triggers an interferon response via activation of heterochromatic repeats. However, how these changes in the expression profile relate to the re-distribution of H1 variants as well as to genome conformational changes have not been yet studied. Here, we combined ChIP-seq of five endogenous H1 variants with Chromosome Conformation Capture analysis in wild-type and H1.2/H1.4 knock-down T47D cells. The results indicate that H1 variants coexist in the genome in two large groups depending on the local GC content and that their distribution is robust with respect to H1 depletion. Despite the small changes in H1 variants distribution, knock-down of H1 translated into more isolated but de-compacted chromatin structures at the scale of topologically associating domains (TADs). Such changes in TAD structure correlated with a coordinated gene expression response of their resident genes. This is the first report describing simultaneous profiling of five endogenous H1 variants and giving functional evidence of genome topology alterations upon H1 depletion in human cancer cells.

An epigenetic characterization of Giemsa bands (G bands) revealed their utility as epigenetic units to investigate the differential distribution of linker histones. GC content is a strong driver in histone H1 distribution: H1X is enriched at high GC bands and H1.2 is abundant at low GC, compacted bands. Hi‐C analysis showed that TADs with a high H1.2/H1X ratio strongly overlap with B compartment, inaccessible chromatin and low‐GC bands.

Human somatic cells may contain up to seven members of the histone H1 family contributing to chromatin compaction and its regulation. In breast cancer cells, knock-down (KD) of each H1 variant results in specific gene expression changes. We have previously shown that combined KD of H1.2 and H1.4 (H1 KD) deregulates many genes, promotes the appearance of accessibility sites genome wide and triggers an interferon response via activation of heterochromatic repeats. Here we describe for the first time ChIP-seq profiling of five endogenous H1 variants at the same time, obtaining that H1 variants are differentially distributed at low (H1.2, H1.5, H1.0) and high (H1X, H1.4) GC content regions. Further, we report that H1 KD promotes redistribution of some of the variants and changes on genome architecture. H1 KD decreased topological associating domain (TAD) border definition and interactions, both between TADs and intra-TAD. In addition, many TADs presented a coordinated gene expression response to H1 KD. Up-regulated genes accumulate within TADs with low gene density and high H1.2 content. In conclusion, our data suggests that the equilibrium between distinct histone H1 variants helps maintaining the topological organization of the genome and the proper expression of particular gene programs.