The Gly(972)Arg variant of human IRS1 gene is associated with variation in glomerular filtration rate likely through impaired insulin receptor signaling

Abstract

The objective of this study is to identify and characterize the genetic variants related to the glomerular filtration rate (GFR) linkage on 2q37. Of the positional candidate genes, we selected IRS1 and resequenced its 2-kb promoter region and exons for sequence variants in 32 subjects. A total of 11 single nucleotide polymorphisms (SNPs) were identified. To comprehensively cover the 59-kb-long intron-1, eight additional tagging SNPs were selected from the HapMap. All the 19 SNPs were genotyped by TaqMan Assay in the entire data set (N = 670; 39 families). Association analyses between the SNPs and GFR and type 2 diabetes-related traits were performed using the measured genotype approach. Of the SNPs examined for association, only the Gly(972)Arg variant of IRS1 exhibited a significant association with GFR (P = 0.0006) and serum triglycerides levels (P = 0.003), after accounting for trait-specific covariate effects. Carriers of Arg972 had significantly decreased GFR values. Gly(972)Arg contributed to 26% of the linkage signal on 2q. Expression of IRS1 mutant Arg972 in human mesangial cells significantly reduced the insulin-stimulated phosphorylation of IRS1 and Akt kinase. Taken together, the data provide the first evidence that genetic variation in IRS1 may influence variation in GFR probably through impaired insulin receptor signaling.

FIG. 1.

Schematic diagram of human IRS1 (NM_005544) gene structure on 2q36 and the location of the SNPs identified and genotyped in SAFDGS. The exons of the IRS1 are represented by solid box and the intron by a thin line. SNPs with the base change and their reference sequence (rs) numbers are indicated by the vertical arrow.

FIG. 2.

LD between SNP pairs within the IRS1 gene. SNPs are labeled on the y-axis, and the locations (bp) within the gene are shown on the x-axis. Pairwise LD is estimated using r² values and depicted in the figure by the color intensity of the shaded box, as shown in the legend. The diagonal represents a comparison of each SNP against itself (i.e., r² = 1.0). (A high-quality color representation of this figure is available in the online issue.)

FIG. 3.

Linkage analysis conditional on the Gly(972)Arg polymorphism on chromosome 2q.

FIG. 4.

Effect of Arg972 on the insulin-induced phosphorylation of IRS1 and Akt in human mesangial cells. A: Tyrosine (941) phosphorylation of IRS1. Human mesangial cells were transfected with plasmids carrying wild type (WT; Gly972) and mutant (Mut; Arg972) IRS1 and treated with 100 nM insulin for 0, 5, and 10 min. Equal amount of total cellular proteins were separated on SDS-PAGE gel and probed with anti-IRS1 and antiphospho-IRS1. B: Threonine (308) phosphorylation of Akt kinase. Human mesangial cells were transfected with plasmids carrying wild-type (Gly972) and mutant (Arg972) IRS1 and treated with 100 nM insulin for 0, 5, and 10 min. Equal amount of total cellular proteins were immunoprecipitated with anti-Akt kinase and separated on SDS-PAGE gel and probed with anti-Akt and antiphospho-Akt kinase. Representative images of three independent experiments are shown.