title: Functional characterization of a -100_-102delAAG deletion-insertion polymorphism in the promoter region of the HTR3B gene.
abstract: OBJECTIVE
  The HTR3B gene encodes the B-subunit of the type 3 serotonin receptor (5-HT3). A -100_-102delAAG deletion in the promoter region has been associated with poor response to antiemetic medication and susceptibility to bipolar affective disorders. The molecular mechanisms underlying these associations, however, remained unclear.
  
  
  METHODS
  We performed electrophoretic mobility shift and luciferase reporter gene assays to elucidate the effect of this polymorphism on the HTR3B promoter activity in PC-12 and HEK293 cells. The reporter constructs carried a 2171 bp fragment of the native HTR3B promoter or 30 bp of the polymorphic locus in tandem triplication upstream of the thymidine kinase minimal promoter.
  
  
  RESULTS
  Deletion mapping indicated that the sequence around the -100_-102delAAG polymorphism had significant promoter activity. Electrophoretic mobility shift assays indicated differential binding of nuclear proteins to the polymorphic DNA region with stronger binding to the insertion than to the deletion allele. The activity of the native promoter carrying the deletion allele was 25% higher in PC-12 (P=0.016) and 40% higher in HEK cells (P=0.016) compared with the respective insertion construct. Constructs carrying the deletion allele in tandem triplicates showed 43% (PC-12 cells, P=0.002) and 28% (HEK293 cells, P=0.015) higher activity than those carrying the insertion allele. The polymorphism was not linked with known amino acid substitutions in HTR3A and HTR3B.
  
  
  CONCLUSIONS
  The -100_-102delAAG 3 bp deletion increases the HTR3B promoter activity in vitro. The consequences of this for the structure and the function of the resulting 5-HT3 receptors remain to be elucidated.
authors.affiliation: Department of Clinical Pharmacology, Georg-August-University Göttingen, Göttingen, Germany.
authors.forename: Cornelia
authors.initials: C
authors.lastname: Meineke
authors.forename: Mladen Vassilev
authors.initials: MV
authors.lastname: Tzvetkov
authors.forename: Kristin
authors.initials: K
authors.lastname: Bokelmann
authors.forename: Elke
authors.initials: E
authors.lastname: Oetjen
authors.forename: Karen
authors.initials: K
authors.lastname: Hirsch-Ernst
authors.forename: Rolf
authors.initials: R
authors.lastname: Kaiser
authors.forename: Jürgen
authors.initials: J
authors.lastname: Brockmöller
mesh_terms: D000818:Animals; D001483:Base Sequence; D002477:Cells; D004247:DNA / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D017930:Genes, Reporter; D006239:Haplotypes; D006801:Humans; D015810:Linkage Disequilibrium; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D016716:PC12 Cells; D010597:Pharmacogenetics; D011401:Promoter Regions, Genetic; D051381:Rats; D011985:Receptors, Serotonin / Q000235:genetics* / Q000378:metabolism; D044406:Receptors, Serotonin, 5-HT3; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D014162:Transfection
chemical_list: D017931:DNA Primers; C509410:HTR3B protein, human; D011985:Receptors, Serotonin; D044406:Receptors, Serotonin, 5-HT3; D011994:Recombinant Proteins; D004247:DNA; D008156:Luciferases