Mouse DNA methylase: Methylation of native DNA

Adams, Roger; McKay, E.L.; Craig, Linda; Burdon, Roy H.

doi:10.1016/0005-2787(79)90143-6

Cited by 87 publications

(37 citation statements)

References 22 publications

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“…These results may be related to the reproducible lag observed in de novo methylation (Fig. 2) and possibly also to different extents of aggregation of the enzyme under different conditions (29).…”

Section: Resultsmentioning

confidence: 87%

Human placental DNA methyltransferase: DNA substrate and DNA binding specificity

1984

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We have partially purified a DNA methyltransferase from human placenta using a novel substrate for a highly sensitive assay of methylation of hemimethylated DNA. This substrate was prepared by extensive nick translation of bacteriophage XP12 DNA, which normally5 has virtually all of its cytosine residues replaced by 5-methylcytosine (m C). Micrococcus luteus DNA wgs just as good a substrate if it was first similarly nick translated with m dCTP instead of dCTP in the polymerization mixture. At different stages in purification and under various conditions (including in the _presence or absence of high mobility group proteins), the methylation of m C-deficient DVA and that of hemimethylated DNA were compared. Although hemimethylated, m C-rich DNAs were much better substrates than were m C-deficient DNAs and normal XP12 DNA could not be methylated, all of these DNAs were bound equally well by the enzyme. In contrast, from the same placental extraSt, a DNAbinding protein of unknown functon was isolated which binds to m C-rich DNA in preference to the analogous m C-poor DNA.

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Section: Resultsmentioning

confidence: 87%

Human placental DNA methyltransferase: DNA substrate and DNA binding specificity

1984

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“…The enzyme DNA 5-cytosine methyltransferase (DNA MeTase) 1 catalyzes the transfer of a methyl group from Sadenosyl methionine to the 5Ј-position of cytosines residing in the dinucleotide sequence CpG (8). Thus far three DNA MeTases have been identified in somatic tissues of vertebrates.…”

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confidence: 99%

Characterization of the Human DNA Methyltransferase Splice Variant Dnmt1b

Bonfils¹,

Beaulieu²,

Chan

et al. 2000

Journal of Biological Chemistry

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“…1). The substrate efficiency of DNA from a variety of organisms was also tested (data not shown) and varied widely, as reported by others (20,(27)(28)(29)(30). In general, G+C-rich DNAs were better substrates than G+C-poor DNAs.…”

Section: Methodsmentioning

confidence: 65%

Ethylation of poly(dC-dG).poly(dC-dG) by ethyl methanesulfonate stimulates the activity of mammalian DNA methyltransferase in vitro.

Farrance¹,

Ivarie²

1985

Proc. Natl. Acad. Sci. U.S.A.

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Ethylation of poly(dC-dG) poly(dC-dG) with ethyl methanesulfonate (EtMes), a known carcinogen, at increasing molar ratios of EtMes/C-G base pairs progressively stimulated the methyl-accepting ability of the DNA during in vitro methylation by partially purified rat DNA (cytosine-5)-methyltransferase (EC 2.1.1.37). Maximum stimulation was 2-fold over mock-treated DNA when 2.7% of the guanines were modified at the N-7 position, the major site of ethylation by EtMes in DNA. If a CpG site "hemiethylated" at guanine N-7 mimics a hemimethylated CpG site, we calculate that the enzyme has a relative affinity for hemiethylated CpG 18-fold above unmodified CpG. If ethylation of a dioxyphosphate oxygen of the phosphodiester bond is responsible for stimulation, the relative affinity could be much higher, up to 370-fold.DNA methylation at cytosine C-5 in the rare CpG dinucleotide appears to play an important role in regulating the expression of many mammalian genes (reviewed in refs. 1-7) and is being extensively studied as a potential target for chemical and radiation-induced carcinogenesis. A large body of evidence now exists showing that chemical carcinogens and UV irradiation, both of which chemically modify DNA, significantly decrease enzymatic methylation of DNA in vivo and in vitro (8)(9)(10)(11)(12)(13)(14). Demethylation may, therefore, play a role in activation of the cancer cell phenotype as proposed by Holliday (15).By contrast, however, we recently found that one widely used chemical carcinogen, ethyl methanesulfonate (EtMes), inactivated specific gene expression (16) apparently by promoting an increase in enzymatic methylation of cultured cell DNA (17). Methyl methanesulfonate has also been reported to increase the level of 5-methylcytidine in DNA in regenerating rat liver (18). Because EtMes introduces ethyl groups primarily at guanine N-7 and secondarily at the phosphodiester bond (19), it was proposed that EtMes might promote enzymatic methylation (17) by creating a "fraudulent" hemimethylated site by rotation of an ethyl group at guanine N-7 or the phosphodiester in close proximity to cytosine C-5 in CpG sequences in B-DNA. This ethylated site might be recognized as hemimethylated by DNA (cytosine-5)-methyltransferase (MeTase; EC 2.1.1.37) and subsequently undergo methylation. We have tested a prediction of the model that unmethylated CpG sites modified by EtMes will be more efficient substrates for DNA methyltransferase in vitro than unmodified sites, and we report the results here. MATERIALS AND METHODSDNA MeTase. The enzyme was extracted with 0.8 M KCl from purified rat liver and spleen nuclei as described (20,21). The crude nuclear extract in 20 mM NaCi in buffer A (50 mM Tris HCl, pH 7.8/1 mM dithiothreitol) was loaded onto a S-adenosyl-L-homocysteine agarose column (3 ml; Bethesda Research Laboratories); the column was washed with 30 ml of 60 mM KCI/buffer A/1 mM EDTA and 15 ml of the same buffer with 20 ,uM S-adenosylmethionine (AdoMet). Enzyme was eluted with 0.3 M KCI in buffer A/1 mM EDTA a...

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Mouse DNA methylase: Methylation of native DNA

Cited by 87 publications

References 22 publications

Human placental DNA methyltransferase: DNA substrate and DNA binding specificity

Human placental DNA methyltransferase: DNA substrate and DNA binding specificity

Characterization of the Human DNA Methyltransferase Splice Variant Dnmt1b

Ethylation of poly(dC-dG).poly(dC-dG) by ethyl methanesulfonate stimulates the activity of mammalian DNA methyltransferase in vitro.

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