An ABCC8 Nonsense Mutation Causing Neonatal Diabetes Through Altered Transcript Expression

Abstract

The pancreatic ATP-sensitive K+ (K-ATP) channel is a key regulator of insulin secretion. Gain-of-function mutations in the genes encoding the Kir6.2 (KCNJ11) and SUR1 (ABCC8) subunits of the channel cause neonatal diabetes, whilst loss-of-function mutations in these genes result in congenital hyperinsulinism. We report two patients with neonatal diabetes in whom we unexpectedly identified recessively inherited loss-of-function mutations. The aim of this study was to investigate how a homozygous nonsense mutation in ABCC8 could result in neonatal diabetes. The ABCC8 p.Glu747* was identified in two unrelated Vietnamese patients. This mutation is located within the in-frame exon 17 and RNA studies confirmed (a) the absence of full length SUR1 mRNA and (b) the presence of the alternatively spliced transcript lacking exon 17. Successful transfer of both patients to sulphonylurea treatment suggests that the altered transcript expression enhances the sensitivity of the K-ATP channel to Mg-ADP/ATP. This is the first report of an ABCC8 nonsense mutation causing a gain-of-channel function and these findings extend the spectrum of K-ATP channel mutations observed in patients with neonatal diabetes.

Keywords: Neonatal diabetes; nonsense mutation splicing..

Figure 1. Expression pattern of ABCC8 transcripts with and without exon 17 in human pancreas and islets. The graphs illustrate the levels of messenger RNA transcripts either containing exon 17 (black bars) or excluding exon 17 (grey bars) expressed relative to levels of the full length ABCC8 reference transcript in human islets (3 technical replicates from RNA pooled from 2 independent donors) and from human total pancreas (3 technical replicates from RNA pooled from 3 donors). Transcript levels were determined by isoform-specific qRTPCR, calculated relative to the endogenous control beta 2 microglobulin (B2M) and normalised to the amount of full length transcript present in each sample. Error bars represent the range of quantification for each sample

Figure 2. The reference genomic DNA sequence is provided. A dashed line denotes an exon-exon boundary. (A) Sequence analysis identified homozygosity for the SUR1Δ17 in patient 1 with the homozygous p.Glu747* ABCC8 variant in exon 17. (B) Sequence analysis of cDNA from the unaffected father who is heterozygous for the p.Glu747* variant identified a transcript which lacked exon 17 and two transcripts which contained exon 17. An alternate splice recognition site at the 3’ intron 16/exon 17 boundary results in two transcripts containing either 36 or 39 basepairs (+/- CAG boxed) (GenBank L78208 and L78224)