title: Methylome analysis for spina bifida showsSOX18hypomethylation as a risk factor with evidence for a complex (epi)genetic interplay to affect neural tube development.
abstract: Neural tube defects (NTDs) are severe congenital malformations that arise from failure of neurulation during early embryonic development. The molecular basis underlying most human NTDs still remains largely unknown. Based on the hypothesis that folic acid prevents NTDs by stimulating methylation reactions, DNA methylation changes could play a role in NTDs. We performed a methylome analysis for patients with myelomeningocele (MMC). Using a candidate CpG analysis forHOXgenes, a significant association betweenHOXB7hypomethylation and MMC was found.
  
  
  
  In the current study, we analyzed leukocyte methylome data of ten patients with MMC and six controls using Illumina Methylation Analyzer and WateRmelon R-packages and performed validation studies using larger MMC and control cohorts with Sequenom EpiTYPER.
  
  
  
  The methylome analysis showed 75 CpGs in 45 genes that are significantly differentially methylated in MMC patients. CpG-specific methylation differences were next replicated for the top six candidate genesABAT,CNTNAP1,SLC1A6,SNED1,SOX18, andTEPPbut only for theSOX18locus a significant overall hypomethylation was observed (Pvalue = 0.0003). Chemically induced DNA demethylation in HEK cells resulted inSOX18hypomethylation and increased expression. Injection ofsox18mRNA in zebrafish resulted in abnormal neural tube formation. Quantification of DNA methylation for theSOX18locus was also determined for five families where parents had normal methylation values compared to significant lower values for both the MMC as their non-affected child.SOX18methylation studies were performed for a MMC patient with a paternally inherited chromosomal deletion that includesBMP4. The patient showed extremeSOX18hypomethylation similar to his healthy mother while his father had normal methylation values.
  
  
  
  This is the first genome-wide methylation study in leukocytes for patients with NTDs. We reportSOX18as a novel MMC risk gene but our findings also suggest thatSOX18hypomethylation must interplay with environmental and (epi)genetic factors to cause NTDs. Further studies are needed that combine methylome data with next-generation sequencing approaches to unravel NTD etiology.
authors.affiliation: Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Campus Gasthuisberg, O&N1, Herestraat 49, Box 911, 3000 Leuven, Belgium ; Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium.
authors.forename: Anne
authors.initials: A
authors.lastname: Rochtus
authors.affiliation: Department of Electrical Engineering ESAT-SCD, University of Leuven, Leuven, Belgium.
authors.forename: Raf
authors.initials: R
authors.lastname: Winand
authors.affiliation: Department of Electrical Engineering ESAT-SCD, University of Leuven, Leuven, Belgium.
authors.forename: Griet
authors.initials: G
authors.lastname: Laenen
authors.affiliation: Genetic Research About Stress and Psychiatry (GRASP), University of Leuven, Leuven, Belgium.
authors.forename: Elise
authors.initials: E
authors.lastname: Vangeel
authors.affiliation: Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Campus Gasthuisberg, O&N1, Herestraat 49, Box 911, 3000 Leuven, Belgium.
authors.forename: Benedetta
authors.initials: B
authors.lastname: Izzi
authors.affiliation: Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Campus Gasthuisberg, O&N1, Herestraat 49, Box 911, 3000 Leuven, Belgium.
authors.forename: Christine
authors.initials: C
authors.lastname: Wittevrongel
authors.affiliation: Department of Electrical Engineering ESAT-SCD, University of Leuven, Leuven, Belgium.
authors.forename: Yves
authors.initials: Y
authors.lastname: Moreau
authors.affiliation: Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium.
authors.forename: Carla
authors.initials: C
authors.lastname: Verpoorten
authors.affiliation: Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium.
authors.forename: Katrien
authors.initials: K
authors.lastname: Jansen
authors.affiliation: Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Campus Gasthuisberg, O&N1, Herestraat 49, Box 911, 3000 Leuven, Belgium ; Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium.
authors.forename: Chris
authors.initials: C
authors.lastname: Van Geet
authors.affiliation: Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Campus Gasthuisberg, O&N1, Herestraat 49, Box 911, 3000 Leuven, Belgium.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Freson
mesh_terms: D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D005786:Gene Expression Regulation; D020022:Genetic Predisposition to Disease; D055106:Genome-Wide Association Study; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D008591:Meningomyelocele / Q000235:genetics* / Q000473:pathology; D054259:Neural Tube / Q000002:abnormalities* / Q000254:growth & development; D055760:SOXF Transcription Factors / Q000235:genetics*
chemical_list: C095473:SOX18 protein, human; D055760:SOXF Transcription Factors
keywords: BMP4; DNA methylation; Epigenetics; Myelomeningocele; Neural tube defects; SOX18; Spina bifida