Differential methylation of the type 2 diabetes susceptibility locus KCNQ1 is associated with insulin sensitivity and is predicted by CpG site specific genetic variation

Abstract

Aims: Epigenetic mechanisms regulate gene expression and may influence the pathogenesis of type 2 diabetes through the loss of insulin sensitivity. The aims of this study were to measure variation in DNA methylation at the type 2 diabetes locus KCNQ1 and assess its relationship with metabolic measures and with genotype.

Methods: DNA methylation from whole blood DNA was quantified using pyrosequencing at 5 CpG sites at the KCNQ1 locus in 510 individuals without diabetes from the 'Relationship between Insulin Sensitivity and Cardiovascular disease' (RISC) cohort. Genotype data was analysed at the same locus in 1119 individuals in the same cohort. Insulin sensitivity was assessed by euglycaemic-hyperinsulinaemic clamp.

Results: DNA methylation at the KCNQ1 locus was inversely associated with insulin sensitivity and serum adiponectin. This association was driven by a methylation-altering Single Nucleotide Polymorphism (SNP) (rs231840) which ablated a methylation site and reduced methylation levels. A second SNP (rs231357), in weak Linkage Disequilibrium (LD) with rs231840, was also associated with insulin sensitivity and DNA methylation. These SNPs have not been previously reported to be associated with type 2 diabetes risk or insulin sensitivity.

Conclusion: Evidence indicates that genetic and epigenetic determinants at the KCNQ1 locus influence insulin sensitivity.

Keywords: Insulin sensitivity; KCNQ1; Methylation; SNP; Type 2 diabetes.

Fig. 1

Chromosome 11p15 locus. A: Black bar represents the amplicon generated for Pyrosequencing®. CpG sites analysed are numbered and shown as black triangles. The grey bar represents the location of the Pyrosequencing® primer. Location of rs231840 is shown. B: Topography of region of interest. Gene annotations were taken from the University of California Santa Cruz genome browser (http://genome.ucsc.edu/). C: Linkage disequilibrium plot created from Haploview v4.2. Values indicated are D′ (1 = complete linkage disequilibrium). Colour scheme represents r² values: r² = 0, white; 0 < r² < 1, shades of grey; r² = 1, black. Grey squares around SNP identifiers indicate the blocks as defined by solid spine linkage disequilibrium (SSLD) or singleton SNPs. Type 2 diabetes associated SNPs are shown at top of the panel (rs2334499, rs231362, rs2237892).

Fig. 2

Methylation vs genotype at rs231840. Boxplot showing the relationship between percentage methylation and genotype. Boxes show the median and interquartile range, whiskers represent the minimum and maximum. Closed circles and star are possible outliers.

Fig. 3

Average methylation at adjacent 4 CpG sites vs genotype at rs231840. Boxplot showing relationship between percentage methylation and genotype. Boxes show the median and interquartile range, whiskers represent the minimum and maximum. Closed circles are possible outliers.

Fig. 4

Pairwise comparisons of KCNQ1 CpGs between blood and other tissues using publicly available array data. Six tissue types are shown (blood: n = 11; muscle, omentum, and subcutaneous fat: n = 6; liver: n = 5; pancreas: n = 4). The upper panel shows the Pearson correlation coefficient and P values; the lower panel shows the pairwise scatterplot (trend line shown in red). Data are a subset of Gene Expression Omnibus data entry GSE48472 . (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)