title: Both polymorphic variable number of tandem repeats and autoimmune regulator modulate differential expression of insulin in human thymic epithelial cells.
abstract: OBJECTIVE
  Polymorphic INS-VNTR plays an important role in regulating insulin transcript expression in the human thymus that leads to either insulin autoimmunity or tolerance. The molecular mechanisms underlying the INS-VNTR haplotype-dependent insulin expression are still unclear. In this study, we determined the mechanistic components underlying the differential insulin gene expression in human thymic epithelial cells, which should have profound effects on the insulin autoimmune tolerance induction.
  
  
  METHODS
  A repetitive DNA region designated as a variable number of tandem repeats (VNTR) is located upstream of the human insulin gene and correlates with the incidence of type 1 diabetes. We generated six class I and two class III VNTR constructs linked to the human insulin basal promoter or SV40 heterologous promoter/enhancer and demonstrated that AIRE protein modulates the insulin promoter activities differentially through binding to the VNTR region.
  
  
  RESULTS
  Here we show that in the presence of the autoimmune regulator (AIRE), the class III VNTR haplotype is responsible for an average of three-fold higher insulin expression than class I VNTR in thymic epithelial cells. In a protein-DNA pull-down experiment, AIRE protein is capable of binding to VNTR class I and III probes. Further, the transcriptional activation of the INS-VNTR by AIRE requires the insulin basal promoter. The VNTR sequence loses its activation activity when linked to a heterologous promoter and/or enhancer.
  
  
  CONCLUSIONS
  These findings demonstrate a type 1 diabetes predisposition encoded by the INS-VNTR locus and a critical function played by AIRE, which constitute a dual control mechanisms regulating quantitative expression of insulin in human thymic epithelial cells.
authors.affiliation: The Research Institute for Children, Children’s Hospital, New Orleans, Louisiana, USA.
authors.forename: Chuan Qi
authors.initials: CQ
authors.lastname: Cai
authors.forename: Tao
authors.initials: T
authors.lastname: Zhang
authors.forename: Mary B
authors.initials: MB
authors.lastname: Breslin
authors.forename: Matthieu
authors.initials: M
authors.lastname: Giraud
authors.forename: Michael S
authors.initials: MS
authors.lastname: Lan
mesh_terms: D015551:Autoimmunity / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics*; D004742:Enhancer Elements, Genetic / Q000235:genetics; D004847:Epithelial Cells / Q000502:physiology*; D005091:Exons / Q000235:genetics; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D007328:Insulin / Q000235:genetics*; D008156:Luciferases / Q000235:genetics; D018598:Minisatellite Repeats / Q000235:genetics; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic / Q000235:genetics; D015183:Restriction Mapping; D020080:Tandem Repeat Sequences / Q000235:genetics*; D013950:Thymus Gland / Q000502:physiology*; D014158:Transcription, Genetic
chemical_list: D017931:DNA Primers; D007328:Insulin; D008156:Luciferases