Methods for the design and analysis of oligodeoxynucleotide-based DNA (cytosine-5) methyltransferase inhibitors

Clark, Jarrod; Шевчук, Т. В.; Kho, Mark R; Smith, Steven S.

doi:10.1016/s0003-2697(03)00402-0

Cited by 12 publications

(7 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, no difference in methylation patterning at CG sites was observed (Figure 7) when C m CWGG was introduced at the APC promoter (a promoter that is not methylated at CG sites in this cell line). Both findings are consistent with the enzymology of wild-type Dnmt1 isolated from human placenta as reported previously (1,48,64–66). This makes it unlikely that the selection process we used has selected for an altered form of Dnmt1.…”

Section: Discussionsupporting

confidence: 93%

Transgene-induced CCWGG methylation does not alter CG methylation patterning in human kidney cells

Шевчук¹

2005

Nucleic Acids Research

View full text Add to dashboard Cite

Several reports suggest that CmCWGG methylation tends not to co-exist with mCG methylation in human cells. We have asked whether or not methylation at CCWGG sites can influence CG methylation. DNA from cells expressing an M.EcoRII–GFP fusion was actively methylated at CCWGG sites. CG methylation as measured by R.HpaII/R.MspI ratios was unchanged in cells expressing the transgene. Cloned representatives of CmCWGG methylated DNA often contained, or were adjacent to an ALU repeat, suggesting that M.EcoRII-GFP actively methylated gene-rich R-band DNA. The transgenic methyltransferase applied CmCWGG methylation to a representative human promoter that was heavily methylated at CG dinucleotides (the SERPINB5 promoter) and to a representative promoter that was essentially unmethylated at CG dinucleotides (the APC promoter). In each case, the CG methylation pattern remained in its original state, unchanged by the presence of neighboring CmCWGG sites. Q-PCR measurements showed that RNA expression from the APC gene was not significantly altered by the presence of CmCWGG in its promoter. Kinetic studies suggested that an adjacent CmCWGG methylation site influences neither the maintenance nor the de novo methylation activities of purified human Dnmt1. We conclude that CmCWGG methylation does not exert a significant effect on CG methylation in human kidney cells.

show abstract

Section: Discussionsupporting

confidence: 93%

Transgene-induced CCWGG methylation does not alter CG methylation patterning in human kidney cells

Шевчук¹

2005

Nucleic Acids Research

View full text Add to dashboard Cite

show abstract

“…Several lines of evidence argue against this model. First, the active site of DNMT1 is essentially impervious to water as demonstrated by the complete lack of cytosine deamination during catalytic methylation (35). Consequently, anhydrous nucleophilic attack at C6 by the active site cysteine is expected to produce C6-DNMT dihydroazacytidine in equilibrium with a ring-open covalent intermediate (see Supplementary Figure S1).…”

Section: Discussionmentioning

confidence: 99%

2′-Deoxyriboguanylurea, the primary breakdown product of 5-aza-2′-deoxyribocytidine, is a mutagen, an epimutagen, an inhibitor of DNA methyltransferases and an inducer of 5-azacytidine-type fragile sites

Lamparska

Clark

Babilonia

et al. 2012

Nucleic Acids Research

Self Cite

View full text Add to dashboard Cite

5-Aza-2′-deoxycytidine (5azaC-dR) has been employed as an inhibitor of DNA methylation, a chemotherapeutic agent, a clastogen, a mutagen, an inducer of fragile sites and a carcinogen. However, its effects are difficult to quantify because it rapidly breaks down in aqueous solution to the stable compound 2′-deoxyriboguanylurea (GuaUre-dR). Here, we used a phosphoramidite that permits the introduction of GuaUre-dR at defined positions in synthetic oligodeoxynucleotides to demonstrate that it is a potent inhibitor of human DNA methyltransferase 1 (hDNMT1) and the bacterial DNA methyltransferase (M.EcoRII) and that it is a mutagen that can form productive base pairs with either Guanine or Cytosine. Pure GuaUre-dR was found to be an effective demethylating agent and was able to induce 5azaC-dR type fragile sites FRA1J and FRA9E in human cells. Moreover, we report that demethylation associated with C:G → G:C transversion and C:G → T:A transition mutations was observed in human cells exposed to pure GuaUre-dR. The data suggest that most of the effects attributed to 5azaC-dR are exhibited by its stable primary breakdown product.

show abstract

“…Catalysis with this DNA and WT M.HhaI is significantly slower than the normal cytosine-containing substrate (Table 1), 11 most likely due to the slower rate of attack on the methylsulfonium from the covalent intermediate. 18,30,31 The use of this inhibitor provides kinetic information about steps before the elimination step ( Figure 1). The inactivation kinetic constants with the three mutants are at least 10 3 -fold slower than that observed with the WT (Table 1).…”

Section: Discussionmentioning

confidence: 99%

AdoMet-dependent Methyl-transfer: Glu119 Is Essential for DNA C5-Cytosine Methyltransferase M.HhaI

Shieh¹,

Reich²

2007

Journal of Molecular Biology

View full text Add to dashboard Cite

Methods for the design and analysis of oligodeoxynucleotide-based DNA (cytosine-5) methyltransferase inhibitors

Cited by 12 publications

References 34 publications

Transgene-induced CCWGG methylation does not alter CG methylation patterning in human kidney cells

Transgene-induced CCWGG methylation does not alter CG methylation patterning in human kidney cells

2′-Deoxyriboguanylurea, the primary breakdown product of 5-aza-2′-deoxyribocytidine, is a mutagen, an epimutagen, an inhibitor of DNA methyltransferases and an inducer of 5-azacytidine-type fragile sites

AdoMet-dependent Methyl-transfer: Glu119 Is Essential for DNA C5-Cytosine Methyltransferase M.HhaI

Contact Info

Product

Resources

About