The majority of 5hmC is nucleosome-associated in naïve B cells.
6 affiliations
- 1Department of Microbial Infection and Immunity, College of Medicine, Ohio State University, Columbus, OH, USA.
- 2Pelotonia Institute for Immuno-Oncology, Ohio State University, James Comprehensive Cancer Center, Columbus, OH, USA.
- 3Department of Biomedical Informatics, College of Medicine, Ohio State University, Columbus, OH, USA.
- 4Advanced Research Laboratory, Active Motif, 1914 Palomar Oaks Way STE 150, Carlsbad, CA, USA. [email protected].
- 5Department of Microbial Infection and Immunity, College of Medicine, Ohio State University, Columbus, OH, USA. [email protected].
- 6Pelotonia Institute for Immuno-Oncology, Ohio State University, James Comprehensive Cancer Center, Columbus, OH, USA. [email protected].
Abstract
Background
DNA demethylation is essential for gene activation and is primarily mediated by the Ten-Eleven-Translocation (TET) dioxygenase family. TET initiates demethylation by oxidizing 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), a chemically stable derivative that is not only an intermediate in demethylation but also an epigenetic mark. 5hmC is enriched at active gene bodies, promoters, and enhancers that exist at accessible chromatin. Yet, 75-90% of genomic DNA is physically packaged in nucleosomes, raising the question about where 5hmC resides relative to nucleosomes. To address this question, we used micrococcal nuclease (MNase) to biochemically stratify 5hmC according to nucleosome protection in naive mouse B cells.Results
Using multiple approaches, we showed that ~ 95% of 5hmC peaks are retained after MNase digest, suggesting that 5hmC is nucleosome-associated. Despite the majority of 5hmC being similar in nucleosomal DNA compared to total DNA, we identified minor subsets that were MNase-sensitive (accessible 5hmC, ~ 3%) or further MNase-resistant (protected 5hmC, ~ 1-2%). Integrative analyses revealed that the accessible 5hmC is preferentially located at promoters and enhancers, and the presence of promoter-proximal accessible 5hmC showed a stronger correlation with gene expression than total 5hmC. In contrast, protected 5hmC is enriched in chromatin states with mixed active and repressive features.Conclusion
These results shed new light on the relationship between 5hmC and accessible chromatin, suggesting 5hmC is predominantly associated with nucleosomes and revealing heterogeneous 5hmC subsets with potential distinct regulatory roles.
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BibTeX
@article{hsu2026majority,
title = {{The majority of 5hmC is nucleosome-associated in naïve B cells.}},
author = {Michael Hsu and Heng-Yi Chen and Fang-Yun Lay and Qin Ma and Benjamin Delatte and Chan-Wang Jerry Lio},
journal = {BMC genomics},
year = {2026},
month = aug,
doi = {10.1186/s12864-026-13032-y},
url = {https://europepmc.org/article/MED/42958012},
note = {CC BY-NC-ND},
}
RIS
TY - JOUR
TI - The majority of 5hmC is nucleosome-associated in naïve B cells.
AU - Michael Hsu
AU - Heng-Yi Chen
AU - Fang-Yun Lay
AU - Qin Ma
AU - Benjamin Delatte
AU - Chan-Wang Jerry Lio
PY - 2026
DA - 2026/08/04
JO - BMC genomics
DO - 10.1186/s12864-026-13032-y
UR - https://europepmc.org/article/MED/42958012
AB - Background DNA demethylation is essential for gene activation and is primarily mediated by the Ten-Eleven-Translocation (TET) dioxygenase family. TET initiates demethylation by oxidizing 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), a chemically stable derivative that is not only an intermediate in demethylation but also an epigenetic mark. 5hmC is enriched at active gene bodies, promoters, and enhancers that exist at accessible chromatin. Yet, 75-90% of genomic DNA is physically packaged in nucleosomes, raising the question about where 5hmC resides relative to nucleosomes. To address this question, we used micrococcal nuclease (MNase) to biochemically stratify 5hmC according to nucleosome protection in naive mouse B cells.Results Using multiple approaches, we showed that ~ 95% of 5hmC peaks are retained after MNase digest, suggesting that 5hmC is nucleosome-associated. Despite the majority of 5hmC being similar in nucleosomal DNA compared to total DNA, we identified minor subsets that were MNase-sensitive (accessible 5hmC, ~ 3%) or further MNase-resistant (protected 5hmC, ~ 1-2%). Integrative analyses revealed that the accessible 5hmC is preferentially located at promoters and enhancers, and the presence of promoter-proximal accessible 5hmC showed a stronger correlation with gene expression than total 5hmC. In contrast, protected 5hmC is enriched in chromatin states with mixed active and repressive features.Conclusion These results shed new light on the relationship between 5hmC and accessible chromatin, suggesting 5hmC is predominantly associated with nucleosomes and revealing heterogeneous 5hmC subsets with potential distinct regulatory roles.
SN - 1471-2164
ER -