title: DNA methylated alleles of the phenylalanine hydroxylase promoter remodeled at elevated phenylalanine levels in newborns with hyperphenylalaninemia.
abstract: OBJECTIVE
  Although high phenylalanine (phe) exposure has been shown to influence the DNA methylation status of leukocytes in hyperphenylalaninemia (HPA), the potential of DNA methylation changes as a biomarker of pretreatment high phe exposure in diet free newborns with HPA has not been explored. We therefore investigated the DNA methylation pattern of the phenylalanine hydroxylase (PAH) gene promoter at different phe levels, and the possibility of DNA methylation pattern changes being a biomarker of high phe exposure in diet free newborns with HPA.
  
  
  METHODS
  With a combination of methylated PCR, high resolution melting, and sequencing, the cytosine phosphodiester bond guanine (CpG) dinucleotides in the 5' untranslated region of the PAH gene were analysed 2-15days after birth using leukocyte DNA from diet free 16 newborns with HPA and 16 healthy controls.
  
  
  RESULTS
  In 2-3days blood cards, GTGTG and GTGC/TG alleles were both detected at similar low mean phe levels in healthy controls (59.39±14.62 and 55.33±13.43μmol/L) and non-phenylketonuria (PKU) HPA (265.00 and 244.25±73.73μmol/L). In HPA with PKU, the GTGTG and GTGC/TG alleles were both detected at dissimilar elevated mean phe levels (380.80±64.62 and 589.00±191.96μmol/L). In ≥7day blood cards, GTGTG and GTGC/TG alleles were both detected at similar excess mean phe levels in HPA with PKU (2297±374.38 and 1562.66±718.23μmol/L).
  
  
  CONCLUSIONS
  The demethylated GTGTG and partial methylated GTGC/TG alleles are not pathogenic alleles. Our results suggest a specific remodeling of the DNA methylated alleles of the PAH promoter at elevated, but not excess phe levels in diet free newborns with PKU.
authors.affiliation: Department of Pediatrics and Adolescent Medicine, Medical University of Vienna, Währinger Gürtel 18-20, Vienna, Austria. Electronic address: bellarmine.item@meduniwien.ac.at.
authors.forename: Chike Bellarmine
authors.initials: CB
authors.lastname: Item
authors.affiliation: Department of Pediatrics and Adolescent Medicine, Medical University of Vienna, Währinger Gürtel 18-20, Vienna, Austria.
authors.forename: Somayeh
authors.initials: S
authors.lastname: Farhadi
authors.affiliation: Department of Pediatrics and Adolescent Medicine, Medical University of Vienna, Währinger Gürtel 18-20, Vienna, Austria.
authors.forename: Andrea
authors.initials: A
authors.lastname: Schanzer
authors.affiliation: Department of Pediatrics and Adolescent Medicine, Medical University of Vienna, Währinger Gürtel 18-20, Vienna, Austria.
authors.forename: Susanne
authors.initials: S
authors.lastname: Greber-Platzer
mesh_terms: D020121:5' Untranslated Regions; D000483:Alleles; D016022:Case-Control Studies; D018899:CpG Islands; D019175:DNA Methylation; D005260:Female; D006801:Humans; D007231:Infant, Newborn; D008297:Male; D009691:Nucleic Acid Denaturation; D010649:Phenylalanine / Q000097:blood*; D010651:Phenylalanine Hydroxylase / Q000097:blood / Q000235:genetics*; D010661:Phenylketonurias / Q000097:blood / Q000175:diagnosis / Q000235:genetics* / Q000473:pathology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA
chemical_list: D020121:5' Untranslated Regions; D010649:Phenylalanine; D010651:Phenylalanine Hydroxylase
keywords: DNA methylation; Epigenetics; Hyperphenylalaninemia; Newborn; Phenylalanine hydroxylase