Abstract
The nucleosome is the fundamental packing unit of the eukaryotic genome, and CpG methylation is an epigenetic modification associated with gene repression and silencing. We investigated nucleosome assembly mediated by histone chaperone Nap1 and the effects of CpG methylation based on three-color single molecule FRET measurements, which enabled direct monitoring of histone binding in the context of DNA wrapping. According to our observation, (H3-H4)2 tetramer incorporation must precede H2A-H2B dimer binding, which is independent of DNA termini wrapping. Upon CpG methylation, (H3-H4)2 tetramer incorporation and DNA termini wrapping are facilitated, whereas proper incorporation of H2A-H2B dimers is inhibited. We suggest that these changes are due to rigidified DNA and increased random binding of histones to DNA. According to the results, CpG methylation expedites nucleosome assembly in the presence of abundant DNA and histones, which may help facilitate gene packaging in chromatin. The results also indicate that the slowest steps in nucleosome assembly are DNA termini wrapping and tetramer positioning, both of which are affected heavily by changes in the physical properties of DNA.
Highlights
Nucleosome assembly mediated by histone chaperones and DNA methylation play important roles in gene regulation
We investigated nucleosome assembly mediated by histone chaperone Nucleosome assembly protein 1 (Nap1) and the effects of CpG methylation based on threecolor single molecule FRET measurements, which enabled direct monitoring of histone binding in the context of DNA wrapping
Similar effects have been reported with human and mouse nucleosomes [32]. These results suggest that CpG methylation may induce changes in the interactions between histones and DNA, which would affect the dynamics of nucleosome assembly
Summary
Nucleosome assembly mediated by histone chaperones and DNA methylation play important roles in gene regulation. We investigated nucleosome assembly mediated by histone chaperone Nap and the effects of CpG methylation based on threecolor single molecule FRET measurements, which enabled direct monitoring of histone binding in the context of DNA wrapping. Similar effects have been reported with human and mouse nucleosomes [32] These results suggest that CpG methylation may induce changes in the interactions between histones and DNA, which would affect the dynamics of nucleosome assembly. We revealed that DNA wrapping and (H3-H4) tetramer binding are facilitated and that canonical dimer binding is inhibited upon CpG methylation, and these results are in line with rigidified DNA and increased histone-DNA affinity Based on these results, we suggest that CpG methylation expedites nucleosome assembly, which may help facilitate gene packaging in chromatin. Our results confirm that the physical properties of DNA are critical factors controlling the efficiency of nucleosome assembly
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