The role of histone modifications in regulating nuclear rigidity and fate plasticity

Abstract

Histone modifications annotate genome function and respond to the environment by modulating gene regulation. They can locally tune the chromatin structure to control expression. In aggregate, they globally influence nuclear rigidity and subsequent cellular behaviors. However, the contribution of specific histone modifications to chromatin structure and function has not been fully investigated both in vitro and in vivo using multiple experimental modalities. Here, we probed the functional interplay of histone modifications and the mechanical environment through four separate experiments.

First, we developed an in vitro phase separation panel to study the association of 77 (modified and unmodified) histones with a chromatin phase. We found that acetylation of the histone H4 tail forms a multiphase structure. Given these results, we hypothesized that histone modifications can directly alter the structure and function of chromatin and proceeded to evaluate this in vivo.

Second, it has been shown in vivo that histone acetylation can anchor onto dense regions of chromatin called chromocenters, analogous to the in vitro recombinant chromatin phase. Given our in vitro results, we speculated that in vivo disruption of chromocenters may reduce H4 acetylation abundance and alter cell fate.We chemically decondensed chromatin using a histone deacetylase (HDAC) inhibitor and found that global chromatin decondensation can influence the accessibility of fate-defining transcription factors.

Third, we physically dissolved chromocenters and used mass spectrometry to characterize histone modification changes following mechanical stretching. We saw that dissolving chromocenters with mechanical force decreased the abundance of H4 acetylation modifications, supporting our in vitro results.

Finally, motivated to understand the functional relevance of mechanically sensitive H4 acetylation, we created cell lines that mimicked specific histone modifications in a hyperacetylated and hypoacetylated state. We hypothesized that hypoacetylated mimic overexpression would create a rigid chromatin environment vulnerable to DNA damage. We observed that overexpressing this hypoacetylated histone mimic led to increased DNA damage following both mechanical stress and senescence induction.

Collectively, these experiments demonstrate that histone H4 acetylation is central to the multiphase organization of regulatory chromatin. These in vitro and in vivo approaches can be generalized to investigate the multifaceted role of histone modifications in controlling gene regulation and cell fate.