title: Comprehensive and quantitative multilocus methylation analysis reveals the susceptibility of specific imprinted differentially methylated regions to aberrant methylation in Beckwith-Wiedemann syndrome with epimutations.
abstract: OBJECTIVE
  Expression of imprinted genes is regulated by DNA methylation of differentially methylated regions (DMRs). Beckwith-Wiedemann syndrome is an imprinting disorder caused by epimutations of DMRs at 11p15.5. To date, multiple methylation defects have been reported in Beckwith-Wiedemann syndrome patients with epimutations; however, limited numbers of DMRs have been analyzed. The susceptibility of DMRs to aberrant methylation, alteration of gene expression due to aberrant methylation, and causative factors for multiple methylation defects remain undetermined.
  
  
  METHODS
  Comprehensive methylation analysis with two quantitative methods, matrix-assisted laser desorption/ionization mass spectrometry and bisulfite pyrosequencing, was conducted across 29 DMRs in 54 Beckwith-Wiedemann syndrome patients with epimutations. Allelic expressions of three genes with aberrant methylation were analyzed. All DMRs with aberrant methylation were sequenced.
  
  
  RESULTS
  Thirty-four percent of KvDMR1-loss of methylation patients and 30% of H19DMR-gain of methylation patients showed multiple methylation defects. Maternally methylated DMRs were susceptible to aberrant hypomethylation in KvDMR1-loss of methylation patients. Biallelic expression of the genes was associated with aberrant methylation. Cis-acting pathological variations were not found in any aberrantly methylated DMR.
  
  
  CONCLUSIONS
  Maternally methylated DMRs may be vulnerable to DNA demethylation during the preimplantation stage, when hypomethylation of KvDMR1 occurs, and aberrant methylation of DMRs affects imprinted gene expression. Cis-acting variations of the DMRs are not involved in the multiple methylation defects.
authors.affiliation: 1] Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan [2] Department of Pediatrics, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Maeda
authors.affiliation: Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Higashimoto
authors.affiliation: Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Jozaki
authors.affiliation: Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Yatsuki
authors.affiliation: Department of Maternal-Fetal Biology, National Research Institute for Child Health and Development, Tokyo, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Nakabayashi
authors.affiliation: Education Center, Asahikawa Medical University, Asahikawa, Japan.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Makita
authors.affiliation: Department of Pediatrics, Maternal, Perinatal, and Child Medical Center, Tenshi Hospital, Sapporo, Japan.
authors.forename: Hidefumi
authors.initials: H
authors.lastname: Tonoki
authors.affiliation: Department of Medical Genetics, Osaka Medical Center and Research Institute for Maternal and Child Health, Izumi, Japan.
authors.forename: Nobuhiko
authors.initials: N
authors.lastname: Okamoto
authors.affiliation: Department of Medical Genetics, Kitasato University Graduate School of Medical Sciences, Kanagawa, Japan.
authors.forename: Fumio
authors.initials: F
authors.lastname: Takada
authors.affiliation: Division of Medical Genetics, Saitama Children's Medical Center, Saitama, Japan.
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Ohashi
authors.affiliation: Department of Pediatrics, Nippon Medical School, Tokyo, Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Migita
authors.affiliation: Division of Medical Genetics, National Center for Child Health and Development, Tokyo, Japan.
authors.forename: Rika
authors.initials: R
authors.lastname: Kosaki
authors.affiliation: Department of Molecular Endocrinology, National Research Institute for Child Health and Development, Tokyo, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Matsubara
authors.affiliation: Department of Pediatrics, Hamamatsu University School of Medicine, Hamamatsu, Japan.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Ogata
authors.affiliation: Department of Pediatrics, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Muneaki
authors.initials: M
authors.lastname: Matsuo
authors.affiliation: Department of Pediatrics, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Yuhei
authors.initials: Y
authors.lastname: Hamasaki
authors.affiliation: 1] Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan [2] Department of Pediatrics, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Ohtsuka
authors.affiliation: Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Kenichi
authors.initials: K
authors.lastname: Nishioka
authors.affiliation: Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Keiichiro
authors.initials: K
authors.lastname: Joh
authors.affiliation: Nishikyushu University, Saga, Japan.
authors.forename: Tsunehiro
authors.initials: T
authors.lastname: Mukai
authors.affiliation: Department of Maternal-Fetal Biology, National Research Institute for Child Health and Development, Tokyo, Japan.
authors.forename: Kenichiro
authors.initials: K
authors.lastname: Hata
authors.affiliation: Division of Molecular Genetics and Epigenetics, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, Saga, Japan.
authors.forename: Hidenobu
authors.initials: H
authors.lastname: Soejima
mesh_terms: D000293:Adolescent; D000483:Alleles; D001506:Beckwith-Wiedemann Syndrome / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D004247:DNA / Q000737:chemistry; D019175:DNA Methylation; D005260:Female; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D020022:Genetic Predisposition to Disease; D018392:Genomic Imprinting; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008297:Male; D009154:Mutation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D004247:DNA