title: Detection of histone modification by chromatin immunoprecipitation combined zinc finger luciferase-based bioluminescence resonance energy transfer assay.
abstract: Epigenetic modification such as DNA methylation and histone modification have important roles in gene regulation. Epigenetic modification can be altered by environmental influences and are related to diseases. Therefore, epigenetic modifications may serve as biomarkers. In this study, we developed a convenient histone modification detection system by combining a chromatin immunoprecipitation (ChIP) and bioluminescence resonance energy transfer (BRET)-based homogeneous PCR product detection system using zinc finger fused to luciferase (ZF-luciferase) with DNA intercalating dye (ChIP-ZF-BRET assay). The ChIP-ZF-BRET assay comprises the following 3 steps: (1) ChIP, (2) PCR amplification of the target genomic region, which includes a zinc-finger recognition site, and (3) homogeneous detection of the PCR product by BRET using ZF-luciferase and fluorescent DNA intercalating dye. Using this system, we conveniently and accurately detected target histone modification at the androgen receptor gene promoter region in LNCaP and Du145 cells. The system can be applicable to DNA methylation detection using a methyl-CpG-binding domain protein or methylcytidine antibody instead of histone modification antibodies. Therefore, it may be useful and convenient for simultaneous detection of histone modification and DNA methylation in clinical diagnoses.
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture & Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Yoshida
authors.forename: Aki
authors.initials: A
authors.lastname: Kezuka
authors.forename: Koich
authors.initials: K
authors.lastname: Abe
authors.forename: Hironobu
authors.initials: H
authors.lastname: Wakeda
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Nakabayashi
authors.forename: Kenichiro
authors.initials: K
authors.lastname: Hata
authors.forename: Kazunori
authors.initials: K
authors.lastname: Ikebukuro
mesh_terms: D047369:Chromatin Immunoprecipitation / Q000379:methods*; D004735:Energy Transfer; D006367:HeLa Cells; D006657:Histones / Q000032:analysis* / Q000502:physiology; D006801:Humans; D008156:Luciferases / Q000737:chemistry*; D008163:Luminescent Measurements / Q000379:methods*; D016335:Zinc Fingers
chemical_list: D006657:Histones; D008156:Luciferases