Cell cycle-specific reactivation of an inactive X-chromosome locus by 5-azadeoxycytidine.

Jones, Peter A.; Taylor, Shirley M.; Mohandas, T.; Shapiro, Larry J.

doi:10.1073/pnas.79.4.1215

Cited by 132 publications

(57 citation statements)

References 24 publications

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“…Numerous investigations described reactivation of genes by 5-Aza-CdR by acting on various loci of the inactive X chromosome (19) and the E-cadherin gene (20). In order to detect similar methylation changes in glioma U251 cells, we also used 5-Aza-CdR.…”

Section: Discussionmentioning

confidence: 99%

Effect of 5-Aza-2'-deoxycytidine on SLC22A18 in glioma U251 cells

Chu

Feng

et al. 2011

Mol Med Report

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Abstract. SLC22A18 [solute carrier family 22 (organic cation transporter) member 18] is located within the 11p15.5 cluster, and may be a new tumor suppressor gene; evidence of SLC22A18 hypermethylation is documented in several types of human cancers. In order to determine whether SLC22A18 hypermethylation is involved in glioma, we determined the SLC22A18 gene protein expression, mRNA expression and methylation status in glioma U251 cells before and after treatment with 5-Aza-2'-deoxycytidine (5-Aza-CdR), and observed the change in growth. Glioma U251 cells treated with 5-AzaCdR were analyzed by flow cytometry to identify any change in their cell cycle profiles. Tumors induced via the injection of untreated U251 cells were measured. Immunohistochemistry, reverse transcriptase-polymerase chain reaction (RT-PCR) and PCR-based methylation assay were carried out to determine SLC22A18 gene protein expression, mRNA expression and methylation status in glioma U251 cells before and after treatment with 5-Aza-CdR. The treated cells showed an increase in their proportion in G1, from 79.2 to 83.5%, and a decrease in S phase, from 12.4 to 5.8%. The apoptotic rate increased from 6.4 to 15.8%. Tumors induced via the injection of untreated U251 cells were approximately 1.46 cm 3 in size, whereas the tumors induced by U251 cells treated with 5-Aza-CdR averaged 0.88 cm 3 in size. The expression levels of SLC22A18 protein and mRNA in U251 cells were increased following treatment with 5x10 -7 M 5-Aza-CdR. Prior to 5-Aza-CdR treatment, the SLC22A18 gene demonstrated hypermethylation and therefore could not be cleaved by HpaII and MspI. It is known that only the DNA digested with HpaII or MspI can be amplified. Following treatment with 5-Aza-CdR, the SLC22A18 gene became demethylated, and could then be cleaved by both of the enzymes, and this failed to be amplified. 5-Aza-cdR may induce glioma U251 cell division and apoptosis and enhance demethylation and protein and mRNA expression of SLC22A18. The hypermethylation of SLC22A18 may be related to the transcriptional silencing of this gene. The growth inhibitory effects of 5-Aza-CdR treatment in vivo remain recognizable.

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Section: Discussionmentioning

confidence: 99%

Effect of 5-Aza-2'-deoxycytidine on SLC22A18 in glioma U251 cells

Chu

Feng

et al. 2011

Mol Med Report

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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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“…Successful pharmacological reversal of DNA methylation has led to the proof of concept for epigenetic therapy of cancer. DNA demethylating agents lead to reexpression of silenced genes, 30,31 and have found applications in non-malignant disease such as sickle cell disease, as well as in cancer. Two agents are FDA approved, 5-Aza Cytidine and 5-Aza-2-deoxyCytidine.…”

Section: Epigenetics Is Affected By Multiple Different Mechanismsmentioning

confidence: 99%