title: Autism associated gene, engrailed2, and flanking gene levels are altered in post-mortem cerebellum.
abstract: BACKGROUND
  Previous genetic studies demonstrated association between the transcription factor engrailed2 (EN2) and Autism Spectrum Disorder (ASD). Subsequent molecular analysis determined that the EN2 ASD-associated haplotype (rs1861972-rs1861973 A-C) functions as a transcriptional activator to increase gene expression. EN2 is flanked by 5 genes, serotonin receptor5a (HTR5A), insulin induced gene1 (INSIG1), canopy1 homolog (CNPY1), RNA binding motif protein33 (RBM33), and sonic hedgehog (SHH). These flanking genes are co-expressed with EN2 during development and coordinate similar developmental processes. To investigate if mRNA levels for these genes are altered in individuals with autism, post-mortem analysis was performed.
  
  
  METHODS
  qRT-PCR quantified mRNA levels for EN2 and the 5 flanking genes in 78 post-mortem cerebellar samples. mRNA levels were correlated with both affection status and rs1861972-rs1861973 genotype. Molecular analysis investigated whether EN2 regulates flanking gene expression.
  
  
  RESULTS
  EN2 levels are increased in affected A-C/G-T individuals (p = .0077). Affected individuals also display a significant increase in SHH and a decrease in INSIG1 levels. Rs1861972-rs1861973 genotype is correlated with significant increases for SHH (A-C/G-T) and CNPY1 (G-T/G-T) levels. Human cell line over-expression and knock-down as well as mouse knock-out analysis are consistent with EN2 and SHH being co-regulated, which provides a possible mechanism for increased SHH post-mortem levels.
  
  
  CONCLUSIONS
  EN2 levels are increased in affected individuals with an A-C/G-T genotype, supporting EN2 as an ASD susceptibility gene. SHH, CNPY1, and INSIG1 levels are also significantly altered depending upon affection status or rs1861972-rs1861973 genotype. Increased EN2 levels likely contribute to elevated SHH expression observed in the post-mortem samples.
authors: Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, United States of America.
authors: Jiyeon
authors: J
authors: Choi
authors: Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, United States of America.
authors: Myka R
authors: MR
authors: Ababon
authors: Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, United States of America.
authors: Mai
authors: M
authors: Soliman
authors: Cancer Institute of New Jersey, Piscataway, New Jersey, United States of America.
authors: Yong
authors: Y
authors: Lin
authors: Department of Genetics, Rutgers University, Piscataway, New Jersey, United States of America.
authors: Linda M
authors: LM
authors: Brzustowicz
authors: Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, United States of America.
authors: Paul G
authors: PG
authors: Matteson
authors: Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey, United States of America ; Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Piscataway, New Jersey, United States of America ; Department of Genetics, Rutgers University, Piscataway, New Jersey, United States of America.
authors: James H
authors: JH
authors: Millonig
mesh_terms: D000818:Animals; D001321:Autistic Disorder / Q000235:genetics*; D002531:Cerebellum / Q000378:metabolism* / Q000473:pathology; D021901:DNA, Intergenic / Q000235:genetics*; D005786:Gene Expression Regulation; D020022:Genetic Predisposition to Disease; D015894:Genome, Human / Q000235:genetics; D057809:HEK293 Cells; D006239:Haplotypes / Q000235:genetics; D053823:Hedgehog Proteins / Q000235:genetics; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D011180:Postmortem Changes; D000099266:Engrailed 2 Protein
chemical_list: D021901:DNA, Intergenic; D053823:Hedgehog Proteins; D018398:Homeodomain Proteins; D009419:Nerve Tissue Proteins; D000099266:Engrailed 2 Protein; C509239:SHH protein, human