BPS2026 – Chaperone functions of H1 linker histone driving canonical and centromeric chromatin assembly

Linker histone H1 has been traditionally recognized for stabilizing nucleosomes and promoting compaction of nucleosomes and higher order chromatin assembly. Our recent AFM studies of canonical and centromere nucleosomes revealed two modes of H1 binding to the nucleosomes forming centrosomes. Additionally, we found that the free DNA in the nucleosome solution decreased upon incubation with H1 histone, suggesting its capability to reform nucleosomes. Here, we tested this hypothesis by performing nucleosome reconstruction experiments with H1 histone. Atomic force microscopy (AFM) was employed to characterize the structural outcomes at single-particle resolution. In this study, H1 linker histone was added to the DNA-octameric histone mixture, the aliquots from the reaction mixture were taken and imaged with AFM. The nucleosome-like particles (NLP) were identified on the AFM images. Approximately 75% of particles assembled in the presence of H1 exhibited nucleosome-like morphology, whereas only 12% of such particles were detected in the absence of H1. The contour length analysis revealed that canonical H3 nucleosome-like particles wrapped ∼133 base pairs of DNA, with an average height of 2.3 nm. Comparable results were observed for centromeric CENP-A nucleosome-like particles, which also displayed ∼133 base pairs of wrapping and a mean height of 2.2 nm. These findings establish that H1 has chaperone functions, driving the assembly of nucleosome-like particles and maintaining consistent structural features across distinct nucleosome types. These findings were validated across independent replicates, highlighting that H1 functions not only as a passive stabilizer of chromatin but also as an active structural driver of nucleosome and chromatosome assembly. By initiating and reinforcing the formation of nucleosomal units, H1 contributes directly to chromatin architecture at the earliest stages of organization. This dual function broadens the mechanistic roles of linker histones in regulating genome packaging and accessibility.