Abstract
Chromatin organization is crucial for regulating gene expression. Previously, we showed that nucleosomes form groups, termed clutches. Clutch size correlated with the pluripotency grade of mouse embryonic stem cells and human induced pluripotent stem cells. Recently, it was also shown that regions of the chromatin containing activating epigenetic marks were composed of small and dispersed chromatin nanodomains with lower DNA density compared to the larger silenced domains. Overall, these results suggest that clutch size may regulate DNA packing density and gene activity. To directly test this model, we carried out 3D, two-color super-resolution microscopy of histones and DNA with and without increased histone tail acetylation. Our results showed that lower percentage of DNA was associated with nucleosome clutches in hyperacetylated cells. We further showed that the radius and compaction level of clutch-associated DNA decreased in hyperacetylated cells, especially in regions containing several neighboring clutches. Importantly, this change was independent of clutch size but dependent on the acetylation state of the clutch. Our results directly link the epigenetic state of nucleosome clutches to their DNA packing density. Our results further provide in vivo support to previous in vitro models that showed a disruption of nucleosome-DNA interactions upon hyperacetylation.
Highlights
Each chromosome of interphase, eukaryotic nuclei occupies a specific nuclear area organized in chromosomal territories (CT) [1,2]
We previously showed that in cells treated with the histone deacetylase (HDAC) inhibitor tricostatin A (TSA), which leads to histone tail hyperacetylation, the nucleosome clutches were smaller compared to untreated cells [8]
Super-resolution microscopy has recently revealed the spatial organization of chromatin and transcriptional machinery in mammalian and bacterial cells at length scales of individual genes that were previously inaccessible to light microscopy [8,24,39,40,41,42,43]
Summary
Eukaryotic nuclei occupies a specific nuclear area organized in chromosomal territories (CT) [1,2]. The existence of this 30 nm fiber in vivo has been long debated and how the chromatin compacts and folds into higher order structures remains unclear. Several recent studies suggested that the organization of nucleosomes and higher order chromatin folding is much more heterogeneous than the regular 30 nm fiber [4,5,6,7,8,9,10,11]. The emerging picture suggests that chromatin is largely comprised of 10 nm fibers with different levels of compaction. In line with these recent studies, our previous work using super-resolution microscopy demonstrated that nucleosomes form heterogeneous groups, termed nucleosome clutches [8]. The size of nucleosome clutches was lower in cells having more open chromatin, including em-
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