title: Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes.
abstract: Methylation-specific fluorescence in situ hybridization (MeFISH) was developed for microscopic visualization of DNA methylation status at specific repeat sequences in individual cells. MeFISH is based on the differential reactivity of 5-methylcytosine and cytosine in target DNA for interstrand complex formation with osmium and bipyridine-containing nucleic acids (ICON). Cell nuclei and chromosomes hybridized with fluorescence-labeled ICON probes for mouse major and minor satellite repeats were treated with osmium for crosslinking. After denaturation, fluorescent signals were retained specifically at satellite repeats in wild-type, but not in DNA methyltransferase triple-knockout (negative control) mouse embryonic stem cells. Moreover, using MeFISH, we successfully detected hypomethylated satellite repeats in cells from patients with immunodeficiency, centromeric instability and facial anomalies syndrome and 5-hydroxymethylated satellite repeats in male germ cells, the latter of which had been considered to be unmethylated based on anti-5-methylcytosine antibody staining. MeFISH will be suitable for a wide range of applications in epigenetics research and medical diagnosis.
authors.affiliation: Division of Epigenomics and Development, Medical Institute of Bioregulation, and Epigenome Network Research Center, Kyushu University, Fukuoka 812-8582, Japan, The Cancer Institute, Tangshan People's Hospital, Hebei 063001, China, Institut de Génétique et de Biologie Moléculaire et Cellulaire, Université de Strasbourg, F-67404 Illkirch, Cité Universitaire de Strasbourg, France, Department of Epigenetics Medicine, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi 409-3898, Japan, Division of Medical Genetics, Saitama Children's Medical Center, Saitama 339-8551, Japan and Research Center for Advanced Science and Technology, The University of Tokyo, Tokyo 113-8656, Japan.
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Li
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Miyanari
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Shirane
authors.forename: Hirohisa
authors.initials: H
authors.lastname: Nitta
authors.forename: Takeo
authors.initials: T
authors.lastname: Kubota
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Ohashi
authors.forename: Akimitsu
authors.initials: A
authors.lastname: Okamoto
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Sasaki
mesh_terms: D044503:5-Methylcytosine / Q000031:analogs & derivatives; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000235:genetics*; D002478:Cells, Cultured; D033481:Chromosomes, Mammalian / Q000737:chemistry*; D003596:Cytosine / Q000031:analogs & derivatives / Q000032:analysis; D019175:DNA Methylation; D004276:DNA, Satellite; D053595:Embryonic Stem Cells / Q000737:chemistry; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008297:Male; D051379:Mice; D013094:Spermatozoa / Q000737:chemistry
chemical_list: D004276:DNA, Satellite; C011865:5-hydroxymethylcytosine; D044503:5-Methylcytosine; D003596:Cytosine