DNA hypomethylation causes an increase in DNase-I sensitivity and an advance in the time of replication of the entire inactive X chromosome

Jablonka, Eva; Goitein, Ruth; Marcus, Menashe; Cedar, Howard

doi:10.1007/bf00293162

Cited by 79 publications

(47 citation statements)

References 20 publications

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“…However to date, the mechanisms of cellular responses of this drug are not well understood. Treatment with 5-aza-CR causes a variety of changes in cells, including decondensation of chromatin (Haaf and Schmidt, 1989), the activation of silenced genes and global genomic hypomethylation (Christman, 2002), and alterations in DNA replication timing (Jablonka et al, 1985), all of which are believed to be consequences of drug-induced demethylation. 5-aza-CR-induced cytotoxicity may be also related to enzyme adduct formation (Juttermann et al, 1994) whereby the incorporation of 5-aza-CR into DNA leads to the irreversible binding of DNMT1 to incorporated 5-aza-CR residues and the rapid loss of DNMT1 activity (Christman, 2002).…”

Section: Discussionmentioning

confidence: 99%

RETRACTION: 5-aza-Cytidine Is a Potent Inhibitor of DNA Methyltransferase 3a and Induces Apoptosis in HCT-116 Colon Cancer Cells via Gadd45- and p53-Dependent Mechanisms

Schneider‐Stock

Diab-Assef²,

Rohrbeck³

et al. 2004

J Pharmacol Exp Ther

109

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Methyltransferase inhibitors commonly used in clinical trials promote tumor cell death, but their detailed cytotoxic action is not yet fully understood. A deeper knowledge about their apotosis-inducing mechanisms and their interaction with DNA methyltransferases (DNMTs) DNMT1, DNMT3a, and DNMT3b might allow the design of more effective drugs with lower cytotoxicity. 5-aza-cytidine (5-aza-CR), a potent inhibitor of DNMT1, is known to induce demethylation and reactivation of silenced genes. In this study, we investigated the p53 dependence of apoptotic, cell cycle, and growth inhibitory effects of 5-aza-CR, as well as the influence on the expression level of DNMT1, DNMT3a, and DNMT3b in the colon cancer cell line HCT-116. Exposure to 5-aza-CR induced the up-regulation of genes promoting cell cycle arrest and DNA repair (p21 WAF1 and GADD45) or apoptosis (p53, RIPK2, Bak1, caspase 5, and caspase 6). In parallel, there was a down-regulation of antiapoptotic Bcl2 protein and the G 2 /M-mediator cyclin B1. Coincubation with pifithrin-alpha (PFT-␣), a selective p53 inhibitor, restored GADD45, Bcl2, cyclin B1, and p21 WAF1 expression levels and almost completely reversed the growth inhibitory, cell cycle, and apoptotic effects of 5-aza-CR. 5-aza-CR treatment caused global demethylation and reactivation of p16 expression. There was a marked decrease in DNMT1 and DNMT3a mRNA expression, with PFT-␣ reversing these effects. However, 5-aza-CR treatment did not modulate DNMT3b expression. Our data demonstrate that 5-aza-CR action in HCT-116 is mediated by p53 and its downstream effectors p21 WAF1 and GADD45. This is the first report to show a link between p53 and regulation of DNMT1 and de novo methyltransferase DNMT3a.

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Section: Discussionmentioning

confidence: 99%

RETRACTION: 5-aza-Cytidine Is a Potent Inhibitor of DNA Methyltransferase 3a and Induces Apoptosis in HCT-116 Colon Cancer Cells via Gadd45- and p53-Dependent Mechanisms

Schneider‐Stock

Diab-Assef²,

Rohrbeck³

et al. 2004

J Pharmacol Exp Ther

109

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“…While this appears to be accomplished through a straightforward binary histone modification code, it should be noted that the relationship between histone acetylation levels and replication timing may actually be quite complex (Figs. 2, 3), and other epigenetic signals, such as DNA methylation, could also play a role in this program (Jablonka et al 1985;Hansen et al 2000). It has been suggested that replication timing may merely reflect local transcription, with active regions being early replicating (Gilbert 2002).…”

Section: Histone Modification Controls Replication Timing Genes and Devmentioning

confidence: 99%

DNA replication timing of the human β-globin domain is controlled by histone modification at the origin

Goren¹,

Tabib²,

Hecht³

et al. 2008

Genes Dev.

Self Cite

130

113

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The human ␤-globin genes constitute a large chromosomal domain that is developmentally regulated. In nonerythroid cells, these genes replicate late in S phase, while in erythroid cells, replication is early. The replication origin is packaged with acetylated histones in erythroid cells, yet is associated with deacetylated histones in nonerythroid cells. Recruitment of histone acetylases to this origin brings about a transcription-independent shift to early replication in lymphocytes. In contrast, tethering of a histone deacetylase in erythroblasts causes a shift to late replication. These results suggest that histone modification at the origin serves as a binary switch for controlling replication timing.Supplemental material is available at http://www.genesdev.org.Received December 20, 2007; revised version accepted March 17, 2008. Studies in both yeast and animal cells demonstrate that the time at which replication origins fire during S phase is established by a preset marking system that operates in early G1 at about the same point that nuclear structure is restored following cell division (Raghuraman et al. 1997;Dimitrova and Gilbert 1999). This epigenetic signal may involve histone modification, as suggested by genetic manipulation experiments in yeast showing that normally early replicating origins can actually be made to fire aberrantly in late S phase simply by causing forced local histone deacetylation (Vogelauer et al. 2002). Whereas in yeast cells each origin replicates at an invariable fixed time within S phase, replication timing in higher eukaryotes is often subject to developmental regulation, and this is accomplished by altering the firing schedule of fixed origins. The human ␤-globin domain represents a good example of this phenomenon. Genetic and biochemical studies have demonstrated that this entire ∼100-kb region is copied by employing a single bidirectional origin that fires early during S phase in erythroid cells, but is converted to a late replication mode in all nonerythroid cells (Kitsberg et al. 1993). Nothing, however, is known about how this timing profile is set up at the molecular level and, in particular, whether this is dependent on local transcription (Gilbert 2002). Results and DiscussionIn order to assess whether this epigenetic switch may be mediated by changes in histone modification, we used chromatin immunoprecipitation (ChIP) analysis to map the mean histone acetylation pattern over the entire ␤-globin domain in different cell types (Fig. 1). In proerythroblast cells, we observed a large enrichment of histone H3 and H4 acetylation over the ␤-globin promoter and coding sequence in correlation with the transcription profile of this gene in these erythroid cells. Mapping studies indicate that the replication origin in this region is actually composed of two separate functional units (Wang et al. 2004), and it appears from our data that these sequences (see map) are also prepackaged with a combination of hyperacetylated histones and me-H3(K4) in the proerythroblast cells (F...

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“…Hypomethylation caused by aza-C can lead to alteration of gene expression, replication timing, and decondensation of chromatin (14)(15)(16)(17)(18). Significantly, tumor-suppressor genes can be reactivated by aza-C treatment, and synergistic effects have been seen with a histone deacetylase inhibitor (19).…”

Section: Introductionmentioning

confidence: 99%

5-Azacytidine–Induced Methyltransferase-DNA Adducts Block DNA Replication In vivo

2007

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5-Azacytidine (aza-C) and its derivatives are cytidine analogues used for leukemia chemotherapy. The primary effect of aza-C is the prohibition of cytosine methylation, which results in covalent methyltransferase-DNA (MTase-DNA) adducts at cytosine methylation sites. These adducts have been suggested to cause chromosomal rearrangements and contribute to cytotoxicity, but the detailed mechanisms have not been elucidated. We used two-dimensional agarose gel electrophoresis and electron microscopy to analyze plasmid pBR322 replication dynamics in Escherichia coli cells grown in the presence of aza-C. Two-dimensional gel analysis revealed the accumulation of specific bubble and Y molecules, dependent on overproduction of the cytosine MTase EcoRII (M.EcoRII) and treatment with aza-C. Furthermore, a point mutation that eliminates a particular EcoRII methylation site resulted in disappearance of the corresponding bubble and Y molecules. These results imply that aza-C-induced MTase-DNA adducts block DNA replication in vivo. RecA-dependent X structures were also observed after aza-C treatment. These molecules may be generated from blocked forks by recombinational repair and/or replication fork regression. In addition, electron microscopy analysis revealed both bubbles and rolling circles (RC) after aza-C treatment. These results suggest that replication can switch from theta to RC mode after a replication fork is stalled by an MTase-DNA adduct. The simplest model for the conversion of theta to RC mode is that the blocked replication fork is cleaved by a branchspecific endonuclease. Such replication-dependent DNA breaks may represent an important pathway that contributes to genome rearrangement and/or cytotoxicity. [Cancer Res 2007;67(17):8248-54]

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DNA hypomethylation causes an increase in DNase-I sensitivity and an advance in the time of replication of the entire inactive X chromosome

Cited by 79 publications

References 20 publications

RETRACTION: 5-aza-Cytidine Is a Potent Inhibitor of DNA Methyltransferase 3a and Induces Apoptosis in HCT-116 Colon Cancer Cells via Gadd45- and p53-Dependent Mechanisms

RETRACTION: 5-aza-Cytidine Is a Potent Inhibitor of DNA Methyltransferase 3a and Induces Apoptosis in HCT-116 Colon Cancer Cells via Gadd45- and p53-Dependent Mechanisms

DNA replication timing of the human β-globin domain is controlled by histone modification at the origin

5-Azacytidine–Induced Methyltransferase-DNA Adducts Block DNA Replication In vivo

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