The combination of DNA methylation and H1 histone binding inhibits the action of a restriction nuclease on plasmid DNA.
To investigate the potentials of DNA methylation and H1 histone in regulating the action of DNA binding proteins, well ordered complexes were formed by slow salt gradient dialysis of mixtures of H1 histone with either methylated or nonmethylated DNA. The sites methylated in the plasmids were CCGG. Methylation of cytosine in this site protects the DNA against HpaII endonuclease but not against MspI. However, when the methylated DNA was complexed to H1, it was protected against MspI. The protection was only effective for a subset of the MspI restriction sites. The protection of DNA afforded by the combination of H1 binding and DNA methylation did not apply to EcoRI, PstI, or BamHI sites and so did not seem to be due to aggregation of the DNA by H1 histone. Gel retardation assays indicated that the affinity of H1 for methylated DNA was not detectably different from its affinity for nonmethylated DNA. Probably methylated DNA when bound to H1 is in a conformation that is resistant to MspI endonuclease. Such conformational changes induced by DNA methylation and H1 binding might affect the action of other DNA binding proteins, perhaps in chromatin as well as in H1.DNA complexes.
Higurashi
M
M
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Cole
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RD
D000818:Animals; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism; D019003:Deoxyribonuclease HpaII; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D008745:Methylation; D051379:Mice; D010957:Plasmids
D006657:Histones; D004247:DNA; D019003:Deoxyribonuclease HpaII; D015252:Deoxyribonucleases, Type II Site-Specific