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 hypothesised that the variant site at the 3′ end of exon 4 might influence splicing and characterised mRNA transcripts to investigate the mutational mechanism underlying this susceptibility to diabetes.

Research design and methods: We established lymphoblastoid cell lines from a G319S homozygote and controls. HNF1A transcripts were characterised 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 up-regulated (55% of mRNA transcripts) compared to 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 7 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 HNF-1α 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.