title: Epigenetic mechanism causes Wnt9b deficiency and nonsyndromic cleft lip and palate in the A/WySn mouse strain.
abstract: BACKGROUND
  The heritable multifactorial etiology of human nonsyndromic cleft lip with or without cleft palate (CL ± P) is not understood. CL ± P occurs in 15% of neonates in the homozygous A/WySn mouse strain, with a multifactorial genetic etiology, the clf1 and clf2 variant genes. Clf1 acts as a mutant allele of Wnt9b but its coding sequence is normal. An IAP (intracisternal A particle) retrotransposon inserted near the Wnt9b gene is associated with clf1.
  
  
  METHODS
  Transcription of noncoding sequence between the IAP and the Wnt9b gene was examined in A/WySn embryos. The levels of Wnt9b transcript and of an "IAP antisense" transcript initiated in the IAP and extending into the noncoding interval were assayed in A/WySn and C57BL/6J whole embryos or heads across embryonic days 8 to 12. Methylation of the 5' LTR of the IAP was examined in E12 A/WySn embryo heads.
  
  
  RESULTS
  Mean Wnt9b transcript levels were lower in A/WySn than in C57BL/6J at all ages examined and lower in CL ± P embryos than in their normal littermates. The "IAP antisense" transcript was found in all A/WySn embryos and was highest in CL ± P embryos. The IAP at Wnt9b was generally unmethylated in CL ± P embryos and approximately 50% methylated in normal littermates.
  
  
  CONCLUSIONS
  The clf1 mutation in A/WySn is a "metastable epiallele", in which stochastic deficiency in some individuals of DNA methylation of a retrotransposon uniquely inserted near the Wnt9b gene allows transcriptional activity of the retrotransposon and interference with transcription from Wnt9b. Methylation of metastable epialleles should be investigated in human nonsyndromic CL ± P.
authors.affiliation: Department of Medical Genetics, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Diana M
authors.initials: DM
authors.lastname: Juriloff
authors.forename: Muriel J
authors.initials: MJ
authors.lastname: Harris
authors.forename: Dixie L
authors.initials: DL
authors.lastname: Mager
authors.forename: Liane
authors.initials: L
authors.lastname: Gagnier
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D001483:Base Sequence; D052160:Benzothiazoles; D002971:Cleft Lip / Q000235:genetics*; D002972:Cleft Palate / Q000235:genetics*; D019175:DNA Methylation / Q000235:genetics / Q000502:physiology*; D003959:Diamines; D004622:Embryo, Mammalian / Q000196:embryology* / Q000648:ultrastructure; D007426:Genes, Intracisternal A-Particle / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains; D008855:Microscopy, Electron, Scanning; D008969:Molecular Sequence Data; D009930:Organic Chemicals; D011804:Quinolines; D060888:Real-Time Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D051153:Wnt Proteins / Q000172:deficiency*
chemical_list: D052160:Benzothiazoles; D003959:Diamines; D009930:Organic Chemicals; D011804:Quinolines; D051153:Wnt Proteins; C515303:Wnt9b protein, mouse; C098022:SYBR Green I
keywords: Wnt9b; cleft lip; epigenetic; genetic; metastable epiallele; methylation; mouse; retrotransposon