Extended and dynamic linker histone-DNA interactions control chromatosome compaction

Abstract

Chromatosomes, composed of nucleosomes and linker histones, play a fundamental role in chromatin regulation. However, a detailed understanding of their structure is lacking, partially due to their complex dynamics. Using single-molecule DNA unzipping with optical tweezers to map histone-DNA interactions in the chromatosome, we reveal a symmetrical compaction of the nucleosome core, governed by the linker histone globular domain contacts at the dyad. Dynamic mapping revealed that the C-terminal domain binds both DNA linkers, at ~ ±115 and ~ ±140 bp from the dyad, to compact the nucleosome entry and exit. Although H1 interacts more transiently with one linker, these dynamic contacts are crucial for stabilizing the binding to the second one. Extensive unzipping of the linker DNA, which mimics its invasion by motor proteins, shifts H1 off the dyad, triggering nucleosome decompaction. Our findings reveal molecular details of chromatosome compaction and its potential modulation by the transcription machinery.

Linker histone H1.8 inhibits chromatin binding of condensins and DNA topoisomerase II to tune chromosome length and individualization

“…Since the condensin complexes were intact at the assay conditions ( Figure 2—figure supplement 1 ), we suspect that non-SMC subunits of condensins were less stable on the nucleosome array than SMC subunits, while H1.8 reduced binding of all condensin subunits to the nucleosome array. The reduced binding of condensins in the presence of H1.8 can be explained by either direct competition between H1.8 and condensins for the linker DNA or by H1.8-mediated formation of a higher-order structure of the nucleosome array ( Song et al, 2014 ; Rudnizky et al, 2021 ). The direct competition model can be tested by using mononucleosomes with linker DNAs instead of the nucleosome array since H1.8 binding to mononucleosomes does not promote higher-order structure or aggregation ( White et al, 2016 ).…”

“…Now we showed that the linker histone H1.8 limits binding of condensin I and II to chromatin both in vitro and in Xenopus egg extracts. H1.8 suppressed condensin binding on both mononucleosomes and nucleosome arrays, suggesting that H1.8 is able to compete out condensins for the same linker DNA targets ( Rudnizky et al, 2021 ), though the capacity of linker histones to promote higher-order structures or phase separation may also limit the access of condensin ( Song et al, 2014 ; Gibson et al, 2019 ). Since condensin I subunits are most abundant chromatin proteins whose levels were enhanced by H1.8 depletion ( Figure 1—figure supplement 1E , Figure 1—source data 2 ), and chromosome lengths can be dictated by the amount of condensins in a manner independently of H1.8 ( Figure 3F ), we propose that regulating linker histone stoichiometry could serve as a rheostat to control chromosome length through tuning the condensin level on chromatin ( Figure 8 ).…”

Single molecule characterization of the binding kinetics of a transcription factor and its modulation by DNA sequence and methylation

“…Our method is based on monitoring the thermal fluctuations of DNA, which are suppressed upon protein binding. With the ability to obtain the full probability distribution for the binding and residence times of a TF to a known site, while at the same time offering the possibility to gradually increase the complexity of the system and follow binding in the vicinity of nucleosomes 54 and chromatosomes 55 , we expect this method to offer important mechanistic insights to complement the information obtained from live cell measurements. We demonstrate our approach by studying binding of Egr1 to the Lhb proximal promoter.…”