Analysis of putative RNase sensitivity and protease insensitivity of demethylation activity in extracts from rat myoblasts

Swisher, Jennifer F.A.; Pyle, Anna Marie; Rand, Eyal; Cedar, Howard

doi:10.1093/nar/26.24.5573

Cited by 39 publications

(26 citation statements)

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“…Initial indications that the reaction was RNA-dependent were not sustained upon further enrichment of the activity (Swisher et al 1998). An RNA-containing demethylating complex was, however, reported in chicken cells (Jost et al 1997(Jost et al , 1999.…”

Section: Active Demethylation Of Dnamentioning

confidence: 99%

DNA methylation patterns and epigenetic memory

Bird¹

2002

Genes Dev.

6,153

4,723

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Section: Active Demethylation Of Dnamentioning

confidence: 99%

DNA methylation patterns and epigenetic memory

Bird¹

2002

Genes Dev.

6,153

4,723

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“…Other investigators suggested that 5-methylcytosine methyl was transferred to an RNAse-sensitive intermediate [11]. However, the involvement of RNA was later ruled out [12]. In the most recent report of a demethylating activity, Battacharya, et al [13] provided evidence for direct demethylation of the cytosine ring.…”

Section: Cytosine Methylation In Gene Regulationmentioning

confidence: 99%

DNA methylation in eukaryotic chromosome stability revisited: DNA methyltransferase in the management of DNA conformation space

Smith

Crocitto

1999

Mol. Carcinog.

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“…Several demethylases have been described previously (2,13,21,24). Unfortunately, the activities of some of these reported demethylases cannot be reproduced in other laboratories (16,22), and in one case they cannot be reproduced in the laboratory reporting the activity (20). It has been found that genes lose their CpG methylation within the promoter when they become activated, whereas genes acquire CpG methylation after they are no longer transcribed (for reviews, see references 4 and 18).…”

mentioning

confidence: 99%

Modulation of DNA Binding Protein Affinity Directly Affects Target Site Demethylation

Lin

Tomzynski

et al. 2000

Molecular and Cellular Biology

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It has recently been shown that in Xenopus, DNA demethylation at promoter regions may involve protein-DNA interactions, based on the specificity of the demethylated sites. Utilizing a stable episomal system in human cells, we recently mapped the sites and dissected the steps of demethylation at oriP sites bound by EBNA1 protein. Although it is clear that protein binding is required for demethylation of the oriP sites, it is uncertain whether this is a unique feature of the replication origin or whether it is a general phenomenon for all DNA sequences to which sequence-specific proteins are bound. In the present study, we utilize the well-defined Escherichia coli lac repressor/operator system in human cells to determine whether protein binding to methylated DNA, in a region that is neither a replication origin nor a promoter, can also lead to demethylation of the binding sites. We found that demethylation specified by protein binding is not unique to the replication origin or to the promoter. We also found that transcriptional activity does not influence demethylation of the lac operator. Isopropyl-␤-D-thiogalactopyranoside (IPTG), an inhibitor of the lac repressor, can prevent demethylation of the lac operator DNA sites and can modulate demethylation of the lac operator by affecting the binding affinity of the lac repressor. Using this system, a titration of protein binding can be done. This titration permits one to infer that protein binding site occupancy is the determinant of demethylation at DNA sites and permits a determination of how this process progresses over time.CpG methylation is tightly regulated during DNA replication and differentiation of somatic cells. The preexisting DNA methylation pattern is maintained by DNA methyltransferase 1 (DNMT1) during DNA replication (14). Changes in the basic methylation pattern occur throughout development through the dynamic processes of de novo methylation and demethylation. Two gene products, DNMT3A and DNMT3B, have been recently reported to have methyltransferase activity both in vitro and in vivo (11, 17). These de novo methyltransferases are believed to be involved in gene regulation, X-inactivation, genomic imprinting, and methylation of endogenous retroviruses and transposable elements (for a review, see reference 27). Several demethylases have been described previously (2,13,21,24). Unfortunately, the activities of some of these reported demethylases cannot be reproduced in other laboratories (16,22), and in one case they cannot be reproduced in the laboratory reporting the activity (20). It has been found that genes lose their CpG methylation within the promoter when they become activated, whereas genes acquire CpG methylation after they are no longer transcribed (for reviews, see references 4 and 18). Matsuo et al. (15) suggested that demethylation may involve protein-DNA interactions in Xenopus embryos, based on the specificity of the demethylated sites. We have further demonstrated in an episomal system that binding of proteins specifies sites of demet...

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Analysis of putative RNase sensitivity and protease insensitivity of demethylation activity in extracts from rat myoblasts

Cited by 39 publications

References 0 publications

DNA methylation patterns and epigenetic memory

DNA methylation patterns and epigenetic memory

DNA methylation in eukaryotic chromosome stability revisited: DNA methyltransferase in the management of DNA conformation space

Modulation of DNA Binding Protein Affinity Directly Affects Target Site Demethylation

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