Ablation of Linker Histone H1.4 Reveals Focal Regulation of AP-1 Directed Enhancers and Chromatin Accessibility [4sU]
Linker histone H1 and the four nucleosomal histone proteins form the greater chromatosome superstructure of chromatin. While a critical component of the epigenetic landscape, the specific role of H1 has remained elusive due in part to compensation by other H1 family members, a lack of genome-wide analyses, and suitable model systems. Ablation of H1.4 in a human osteosarcoma line with high H1-4 expression resulted in broad changes in DNA accessibility as well as gene expression. Genes in critical signaling pathways are upregulated with H1.4 loss (RAR, acetylcholine/glutaminergic receptors) while extracellular matrix and collagen biosynthesis genes are repressed. The gene loci of the same critical pathways were enriched in differentially accessible chromatin after loss of H1.4, and many of this differential chromatin overlaps regions of known enhancer regions. The AP-1 binding motif was highly enriched at sites with reduced accessibility while sites with gained chromatin accessibility were enriched in more basal factor motifs (TBP, SRF). At sites of altered chromatin accessibility following H1.4-loss, concomitant changes to core histone proteins were observed (H3K27ac and H3K4me1), and the changes in PTMs was concordant with the change in accessibility. Alterations to chromatin accessibility and epigenetic activity were enriched at regions with known enhancer-like chromatin, and the size of the altered chromatin footprints were consistent with enhancer regions. Sites which gain chromatin accessibility localize to polycomb, heterochromatin and quiescent/low chromatin states, but were found to gain RNA transcription in cells lacking H1.4. Broad changes in chromatin accessibility and activity, as well as robust changes in gene transcription when H1.4 is abolished, place linker H1.4 as a hitherto underappreciated regulator of transcription response patterns as well as a dynamic member of regulatory chromatin. Overall design: Establishment of two Linker histone H1.4-deficient human osteosarcoma cell lines and interrogation of changes to chromatin accessibilty (ATAC-seq) and chromatin activity (H3K27ac and H3K4me1 ChIP-seq). Further analysis of changes to both gene expression (RNA-seq) and nascent transcription (4sU-seq) were also performed.