title: Diabetes susceptibility in the Canadian Oji-Cree population is moderated by abnormal mRNA processing of HNF1A G319S transcripts.
abstract: OBJECTIVE
  The G319S HNF1A variant is associated with an increased risk of type 2 diabetes in the Canadian Oji-Cree population. We hypothesized that the variant site at the 3' end of exon 4 might influence splicing and characterized mRNA transcripts to investigate the mutational mechanism underlying this susceptibility to diabetes.
  
  
  METHODS
  We established lymphoblastoid cell lines from a G319S homozygote and controls. HNF1A transcripts were characterized in the cell lines and pancreatic tissue by sequence analysis of RT-PCR products and quantification using real-time PCR. Susceptibility to mRNA surveillance was investigated using cycloheximide.
  
  
  RESULTS
  Full-length G319S mRNA accounted for 24% of mRNA transcripts in the homozygous G319S cell line. A novel isoform lacking the terminal 12 bases of exon 4 was upregulated (55% of mRNA transcripts) compared with control cell lines (33%) and human pancreatic tissue (17%). Two abnormal transcripts present only in the G319S cell line included premature termination codons as a result of the inclusion of seven nucleotides from intron 4 or the deletion of exon 8. Cycloheximide treatment increased the levels of both transcripts.
  
  
  CONCLUSIONS
  The G319S variant results in the production of two abnormal transcripts and an alteration in the relative balance of normal splicing products. This is predicted to lead to a reduction in total HNF1A transcript levels, but residual hepatocyte nuclear factor-1alpha protein activity in G319S homozygotes may still reach up to 66% of normal levels. A combination of abnormal splicing and reduced activity of the G319S protein may explain the diabetes susceptibility.
authors.affiliation: Institute of Biomedical and Clinical Sciences, Peninsula Medical School, Exeter, U.K. l.w.harries@exeter.ac.uk
authors.forename: Lorna W
authors.initials: LW
authors.lastname: Harries
authors.forename: Melissa J
authors.initials: MJ
authors.lastname: Sloman
authors.forename: Elizabeth A C
authors.initials: EA
authors.lastname: Sellers
authors.forename: Andrew T
authors.initials: AT
authors.lastname: Hattersley
authors.forename: Sian
authors.initials: S
authors.lastname: Ellard
mesh_terms: D019943:Amino Acid Substitution; D002170:Canada / Q000453:epidemiology; D002460:Cell Line; D003920:Diabetes Mellitus / Q000453:epidemiology / Q000235:genetics*; D005091:Exons; D005786:Gene Expression Regulation; D020022:Genetic Predisposition to Disease; D014644:Genetic Variation; D051538:Hepatocyte Nuclear Factor 1-alpha / Q000235:genetics*; D006720:Homozygote; D006801:Humans; D007198:Indians, North American / Q000235:genetics*; D008214:Lymphocytes / Q000502:physiology; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics* / Q000302:isolation & purification; D014158:Transcription, Genetic
chemical_list: C496487:HNF1A protein, human; D051538:Hepatocyte Nuclear Factor 1-alpha; D012333:RNA, Messenger