title: Effect of alcohol consumption on CpG methylation in the differentially methylated regions of H19 and IG-DMR in male gametes: implications for fetal alcohol spectrum disorders.
abstract: BACKGROUND
  Exposure to alcohol in utero is the main attributable cause of fetal alcohol spectrum disorders (FASD) which in its most severe form is characterized by irreversible behavioral and cognitive disability. Paternal preconception drinking is not considered to be a significant risk factor, even though animal studies have demonstrated that chronic paternal alcohol consumption has a detrimental effect on the physical and mental development of offspring even in the absence of in utero alcohol exposure. It has been documented that alcohol can reduce the levels and activity of DNA methyltransferases resulting in DNA hypomethylation and that reduced methyltransferase activity can cause activation of normally silenced genes. The aim of this study was to establish a link between alcohol use in men and hypomethylation of paternally imprinted loci in sperm DNA in genomic regions critical for embryonic development, thus providing a mechanism for paternal effects in the aetiology of FASD.
  
  
  METHODS
  Sperm DNA from male volunteers was bisulfite treated and the methylation patterns of 2 differentially methylated regions (DMRs), H19 and IG-DMR, analyzed following sequencing of individual clones. The methylation patterns were correlated with the alcohol consumption levels of the volunteer males.
  
  
  RESULTS
  There was a pattern of increased demethylation with alcohol consumption at the 2 imprinted loci with a significant difference observed at the IG-DMR between the nondrinking and heavy alcohol consuming groups. Greater inter-individual variation in average methylation was observed at the H19 DMR and individual clones were more extensively demethylated than those of the IG-DMR. CpG site #4 in the IG-DMR was preferentially demethylated among all individuals and along with the H19 DMR CpG site #7 located within the CTCF binding site 6 showed significant demethylation in the alcohol consuming groups compared with the control group.
  
  
  CONCLUSIONS
  This study demonstrates a correlation between chronic alcohol use and demethylation of normally hypermethylated imprinted regions in sperm DNA. We hypothesize that, should these epigenetic changes in imprinted genes be transmitted through fertilization, they would alter the critical gene expression dosages required for normal prenatal development resulting in offspring with features of FASD.
authors.affiliation: Division of Human Genetics, National Health Laboratory Service and School of Pathology, University of the Witwatersrand, Johannesburg, South Africa.
authors.forename: Lillian A
authors.initials: LA
authors.lastname: Ouko
authors.forename: Katpaham
authors.initials: K
authors.lastname: Shantikumar
authors.forename: Jaysen
authors.initials: J
authors.lastname: Knezovich
authors.forename: Philip
authors.initials: P
authors.lastname: Haycock
authors.forename: Desmond J
authors.initials: DJ
authors.lastname: Schnugh
authors.forename: Michèle
authors.initials: M
authors.lastname: Ramsay
mesh_terms: D000328:Adult; D000428:Alcohol Drinking / Q000378:metabolism*; D002492:Central Nervous System Depressants / Q000494:pharmacology*; D003001:Cloning, Molecular; D018899:CpG Islands / Q000187:drug effects / Q000235:genetics*; D004247:DNA / Q000187:drug effects / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D000431:Ethanol / Q000494:pharmacology*; D005260:Female; D063647:Fetal Alcohol Spectrum Disorders / Q000235:genetics*; D005854:Germ Cells / Q000187:drug effects*; D006801:Humans; D008297:Male; D008745:Methylation / Q000187:drug effects; D011247:Pregnancy; D012313:RNA / Q000235:genetics* / Q000378:metabolism*; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated / Q000235:genetics* / Q000378:metabolism*; D013076:Sperm Count; D013094:Spermatozoa / Q000737:chemistry / Q000187:drug effects / Q000648:ultrastructure; D019966:Substance-Related Disorders / Q000235:genetics; D013447:Sulfites / Q000737:chemistry; D055815:Young Adult
chemical_list: D002492:Central Nervous System Depressants; C094335:H19 long non-coding RNA; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; D013447:Sulfites; D000431:Ethanol; D012313:RNA; D004247:DNA