Global DNA Methylation Level Monitoring by methyl-CpG Binding Domain-Fused Luciferase. (24th March 2019)

• Record Type: Journal Article
• Authors: Baba, Yuji; Karube, Isao; Yoshida, Wataru
• Is Part Of: Analytical letters. Volume 52:Number 5(2019)
• Page Start: 754
• Page End: 760
• Publication Date: 2019-03-24
• Subjects: 5-Aza-2′-deoxycytidine -- bioluminescence resonance energy transfer -- DNA methylation -- folic acid -- luciferase -- methyl-CpG binding domain; Chemistry, Analytic -- Periodicals; Chemistry, Analytic -- Abstracts; 543
• DOI: 10.1080/00032719.2018.1494739
• Languages: English
• ISSNs: 0003-2719

Abstract

Abstract: DNA methylation is an epigenetic modification that represses gene expression. In cancer cells, alterations of the DNA methylation state in promoter regions and repetitive DNA sequences are observed; therefore, DNA methyltransferase inhibitors have been the focus of interest as potential anticancer drugs. We previously reported a simple global DNA methylation level-sensing assay using methyl-CpG binding domain (MBD) fused to luciferase (MBD-luciferase). In the assay, the MBD-luciferase binds to methyl-CpG sites on genomic DNA. Subsequently, bioluminescence resonance energy transfer (BRET) between the luciferase and a fluorescent DNA intercalating dye generates a signal that is dependent on DNA methylation level. In this study, we investigated whether global DNA hypomethylation induced by a DNA methyltransferase inhibitor or nutrient can be monitored by the BRET assay. 5-Aza-2′-deoxycytidine and folic acid were utilized as the DNA-methyltransferase inhibitor and nutrient that affect DNA methylation in cells. The HeLa cells were cultured with the inhibitor or in folic acid-deficient medium and their global DNA methylation levels measured. Both time- and concentration-dependent hypomethylation were detected by the BRET assay. These results demonstrate that global DNA hypomethylation can be monitored by the BRET assay, indicating that the assay is applicable to cell-based screening of DNA-methyltransferase inhibitors.