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Methylation of nucleosomal and nuclease sensitive DNA
Abstract:
The proportion of cytosines methylated in the DNA of nucleosome oligomers and of core particles appears indistinguishable from that of total nuclear DNA from CHO cells. However the DNA in nucleoprotein which is initially released from nuclei by treatment with very low levels of micrococcal nuclease and the first 10% of material rendered acid soluble by treatment of nuclei with DNase I are enriched 2 fold in their content of 5 methylcytosine. (Cessation of hydrolysis by nuclease occurs concomitantly with precipitation of nucleosomal core particles).
Insights
DNA methylation in CHO cells is similar in nucleosomes and total DNA. However, early fractions released by micrococcal nuclease and DNase I show a two-fold increase in 5-methylcytosine, indicating specific DNA regions are preferentially methylated.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- DNA methylation is a crucial epigenetic modification regulating gene expression.
- The distribution of DNA methylation within chromatin structures like nucleosomes is not fully understood.
- Chinese Hamster Ovary (CHO) cells are a common model system for studying cellular processes.
Purpose of the Study:
- To investigate the distribution of 5-methylcytosine in different DNA fractions derived from CHO cell nuclei.
- To determine if DNA methylation levels differ between nucleosome-associated DNA and DNA released by enzymatic digestion.
Main Methods:
- Isolation of nucleosome oligomers and core particles from CHO cell nuclei.
- Enzymatic digestion of nuclei using micrococcal nuclease and DNase I.
- Quantification of 5-methylcytosine content in DNA fractions using biochemical assays.
Main Results:
- The proportion of methylated cytosines in nucleosome oligomers and core particles was comparable to total nuclear DNA.
- Early DNA fractions released by low levels of micrococcal nuclease treatment were enriched twofold in 5-methylcytosine.
- The first 10% of acid-soluble material from DNase I treatment also showed a twofold enrichment of 5-methylcytosine.
Conclusions:
- Specific, more accessible DNA regions within CHO cell nuclei exhibit higher levels of 5-methylcytosine.
- These findings suggest a non-uniform distribution of DNA methylation within the nucleus, potentially linked to chromatin accessibility.