title: Identification of a novel functional deletion variant in the 5'-UTR of the DJ-1 gene.
abstract: BACKGROUND
  DJ-1 forms part of the neuronal cellular defence mechanism against oxidative insults, due to its ability to undergo self-oxidation. Oxidative stress has been implicated in the pathogenesis of central nervous system damage in different neurodegenerative disorders including Alzheimer's disease and Parkinson's disease (PD). Various mutations in the DJ-1 (PARK7) gene have been shown to cause the autosomal recessive form of PD. In the present study South African PD patients were screened for mutations in DJ-1 and we aimed to investigate the functional significance of a novel 16 bp deletion variant identified in one patient.
  
  
  METHODS
  The possible effect of the deletion on promoter activity was investigated using a Dual-Luciferase Reporter assay. The DJ-1 5'-UTR region containing the sequence flanking the 16 bp deletion was cloned into a pGL4.10-Basic luciferase-reporter vector and transfected into HEK293 and BE(2)-M17 neuroblastoma cells. Promoter activity under hydrogen peroxide-induced oxidative stress conditions was also investigated. Computational (in silico) cis-regulatory analysis of DJ-1 promoter sequence was performed using the transcription factor-binding site database, TRANSFAC via the PATCH and rVISTA platforms.
  
  
  RESULTS
  A novel 16 bp deletion variant (g.-6_+10del) was identified in DJ-1 which spans the transcription start site and is situated 93 bp 3' from a Sp1 site. The deletion caused a reduction in luciferase activity of approximately 47% in HEK293 cells and 60% in BE(2)-M17 cells compared to the wild-type (P < 0.0001), indicating the importance of the 16 bp sequence in transcription regulation. The activity of both constructs was up-regulated during oxidative stress. Bioinformatic analysis revealed putative binding sites for three transcription factors AhR, ARNT, HIF-1 within the 16 bp sequence. The frequency of the g.-6_+10del variant was determined to be 0.7% in South African PD patients (2 heterozygotes in 148 individuals).
  
  
  CONCLUSIONS
  This is the first report of a functional DJ-1 promoter variant, which has the potential to influence transcript stability or translation efficiency. Further work is necessary to determine the extent to which the g.-6_+10del variant affects the normal function of the DJ-1 promoter and whether this variant confers a risk for PD.
authors.affiliation: Division of Molecular Biology and Human Genetics, Faculty of Health Sciences, University of Stellenbosch, Cape Town, 8001, South Africa. rowenak@sun.ac.za
authors.forename: Rowena J
authors.initials: RJ
authors.lastname: Keyser
authors.forename: Lize
authors.initials: L
authors.lastname: van der Merwe
authors.forename: Mauritz
authors.initials: M
authors.lastname: Venter
authors.forename: Craig
authors.initials: C
authors.lastname: Kinnear
authors.forename: Louise
authors.initials: L
authors.lastname: Warnich
authors.forename: Jonathan
authors.initials: J
authors.lastname: Carr
authors.forename: Soraya
authors.initials: S
authors.lastname: Bardien
mesh_terms: D020121:5' Untranslated Regions / Q000235:genetics*; D000293:Adolescent; D000328:Adult; D000368:Aged; D001483:Base Sequence; D002460:Cell Line; D017124:Conserved Sequence; D005260:Female; D017353:Gene Deletion; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D015513:Oncogene Proteins / Q000235:genetics*; D010300:Parkinson Disease / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D011401:Promoter Regions, Genetic; D000071617:Protein Deglycase DJ-1; D014158:Transcription, Genetic; D055815:Young Adult
chemical_list: D020121:5' Untranslated Regions; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; C105131:PARK7 protein, human; D000071617:Protein Deglycase DJ-1