Genome-wide association studies and type 2 diabetes

In recent years, the search for genetic determinants of type 2 diabetes (T2D) has changed dramatically. Although linkage and small-scale candidate gene studies were highly successful in the identification of genes, which, when mutated, caused monogenic forms of T2D, they were largely unsuccessful when applied to the more common forms of the disease. To date, these approaches have only identified two loci (PPARG, KCNJ11) robustly implicated in T2D susceptibility. The ability to perform large-scale association analysis, including genome-wide association studies (GWAS) in many thousands of samples from different populations, and subsequently, the shift to form large international collaborations to perform meta-analyses across many studies has taken the number of independent loci showing genome-wide significant associations with T2D to 44. This number includes six loci identified initially through the analysis of quantitative glycaemic phenotypes, illustrating the usefulness of this approach both to identify new disease genes and gain insight into the mechanisms leading to disease. Combined, these loci still only account for ～10% of the observed familial clustering in Europeans, leaving much of the variance unexplained. In this review, we will describe what GWAS have taught us about the genetic basis of T2D and discuss possible next steps to uncover the remaining heritability.     

Genome-wide analysis of copy number variation in type 1 diabetes

Type 1 diabetes (T1D) tends to cluster in families, suggesting there may be a genetic component predisposing to disease. However, a recent large-scale genome-wide association study concluded that identified genetic factors, single nucleotide polymorphisms, do not account for overall familiality. Another class of genetic variation is the amplification or deletion of >1 kilobase segments of the genome, also termed copy number variations (CNVs). We performed genome-wide CNV analysis on a cohort of 20 unrelated adults with T1D and a control (Ctrl) cohort of 20 subjects using the Affymetrix SNP Array 6.0 in combination with the Birdsuite copy number calling software. We identified 39 CNVs as enriched or depleted in T1D versus Ctrl. Additionally, we performed CNV analysis in a group of 10 monozygotic twin pairs discordant for T1D. Eleven of these 39 CNVs were also respectively enriched or depleted in the Twin cohort, suggesting that these variants may be involved in the development of islet autoimmunity, as the presently unaffected twin is at high risk for developing islet autoimmunity and T1D in his or her lifetime. These CNVs include a deletion on chromosome 6p21, near an HLA-DQ allele. CNVs were found that were both enriched or depleted in patients with or at high risk for developing T1D. These regions may represent genetic variants contributing to development of islet autoimmunity in T1D.     

Genome-wide association studies provide new insights into type 2 diabetes aetiology

Human geneticists are currently in the middle of a race. Thanks to a new technology in the form of 'genome-wide chips', investigators can potentially find many novel disease genes in one large experiment. Type 2 diabetes has been hot out of the blocks with six recent publications that together provide convincing evidence for six new gene regions involved in the condition. Together with candidate approaches, these studies have identified 11 confirmed genomic regions that alter the risk of type 2 diabetes in the European population. One of these regions, the fat mass and obesity associated gene (FTO), represents by far the best example of an association between common variation and fat mass in the general population.     

锌转运蛋白8与1型和2型糖尿病的研究进展

ZnT8(zinc transporter,member8)是锌离子转运蛋白,主要定位于胰岛β细胞,能将胞浆锌离子转运至胰岛素储存/分泌性囊泡内,其转运功能降低会影响胰岛素合成、储存和分泌,能增加2型糖尿病(T2DM)的发病风险。ZnT8蛋白也可作为抗原引起β细胞自身免疫损伤,诱发1型糖尿病(T1DM)。ZnT8基因多态性是引起其锌离子转运功能和免疫原性变化的重要因素,与糖尿病的发生、发展密切相关。该文综述了ZnT8与T1DM和T2DM的研究进展,提示ZnT8可作为糖尿病防治的新药物靶点。     

Anti-idiotypic antibody specific to GAD65 autoantibody prevents type 1 diabetes in the NOD mouse

Overt autoantibodies to the smaller isoform of glutamate decarboxylase (GAD65Ab) are a characteristic in patients with Type 1 diabetes (T1D). Anti-idiotypic antibodies (anti-Id) directed to GAD65Ab effectively prevent the binding of GAD65 to GAD65Ab in healthy individuals. Levels of GAD65Ab-specific anti-Id are significantly lower in patients with T1D, leading to overt GAD65Ab in these patients. To determine the possible protective role of GAD65Ab-specific anti-Id in T1D pathogenesis, we developed the monoclonal anti-Id MAb 8E6G4 specifically targeting human monoclonal GAD65Ab b96.11. MAb 8E6G4 was demonstrated as a specific anti-Id directed to the antigen binding site of b96.11. MAb 8E6G4 recognized human antibodies in sera from healthy individuals, T2D patients, and T1D patients as established by ELISA. We confirmed these MAb 8E6G4-bound human antibodies to contain GAD65Ab by testing the eluted antibodies for binding to GAD65 in radioligand binding assays. These findings confirm that GAD65Ab are present in sera of individuals, who test GAD65Ab-negative in conventional detection assays. To test our hypothesis that GAD65Ab-specific anti-Id have an immune modulatory role in T1D, we injected young Non Obese Diabetic (NOD) mice with MAb 8E6G4. The animals were carefully monitored for development of T1D for 40 weeks. Infiltration of pancreatic islets by mononuclear cells (insulitis) was determined to establish the extent of an autoimmune attack on the pancreatic islets. Administration of MAb 8E6G4 significantly reduced the cumulative incidence rate of T1D and delayed the time of onset. Insulitis was significantly less severe in animals that received MAb 8E6G4 as compared to control animals. These results support our hypothesis that anti-Id specific to GAD65Ab have a protective role in T1D.     

Genome-wide association analysis for feed efficiency in Angus cattle

Estimated breeding values for average daily feed intake (AFI; kg/day), residual feed intake (RFI; kg/day) and average daily gain (ADG; kg/day) were generated using a mixed linear model incorporating genomic relationships for 698 Angus steers genotyped with the Illumina BovineSNP50 assay. Association analyses of estimated breeding values (EBVs) were performed for 41,028 single nucleotide polymorphisms (SNPs), and permutation analysis was used to empirically establish the genome-wide significance threshold (P < 0.05) for each trait. SNPs significantly associated with each trait were used in a forward selection algorithm to identify genomic regions putatively harbouring genes with effects on each trait. A total of 53, 66 and 68 SNPs explained 54.12% (24.10%), 62.69% (29.85%) and 55.13% (26.54%) of the additive genetic variation (when accounting for the genomic relationships) in steer breeding values for AFI, RFI and ADG, respectively, within this population. Evaluation by pathway analysis revealed that many of these SNPs are in genomic regions that harbour genes with metabolic functions. The presence of genetic correlations between traits resulted in 13.2% of SNPs selected for AFI and 4.5% of SNPs selected for RFI also being selected for ADG in the analysis of breeding values. While our study identifies panels of SNPs significant for efficiency traits in our population, validation of all SNPs in independent populations will be necessary before commercialization.     

拷贝数变异的全基因组关联分析

基因组拷贝数变异(copy number variations,CNVs)是指与基因组参考序列相比,基因组中≥1 kb的DNA片段插入、缺失和/或扩增,及其互相组合衍生出的复杂变异．由于其具有分布范围广、可遗传、相对稳定和高度异质性等特点,目前认为,CNVs是一种新的可以作为疾病易感标志的基因组DNA多态性,其变异引起的基因剂量改变可以导致表型改变．最近,一种基于CNVs的新的疾病易感基因鉴定策略——CNV全基因组关联分析开始出现,这一策略和传统的基于单核苷酸多态性的关联分析具有互补性,通过认识基因组结构变异可以认识复杂疾病的分子机制和遗传基础．     

Genome-wide association analysis of oxidative stress resistance in Drosophila melanogaster

Background

Aerobic organisms are susceptible to damage by reactive oxygen species. Oxidative stress resistance is a quantitative trait with population variation attributable to the interplay between genetic and environmental factors. Drosophila melanogaster provides an ideal system to study the genetics of variation for resistance to oxidative stress.

Methods and Findings

We used 167 wild-derived inbred lines of the Drosophila Genetic Reference Panel for a genome-wide association study of acute oxidative stress resistance to two oxidizing agents, paraquat and menadione sodium bisulfite. We found significant genetic variation for both stressors. Single nucleotide polymorphisms (SNPs) associated with variation in oxidative stress resistance were often sex-specific and agent-dependent, with a small subset common for both sexes or treatments. Associated SNPs had moderately large effects, with an inverse relationship between effect size and allele frequency. Linear models with up to 12 SNPs explained 67–79% and 56–66% of the phenotypic variance for resistance to paraquat and menadione sodium bisulfite, respectively. Many genes implicated were novel with no known role in oxidative stress resistance. Bioinformatics analyses revealed a cellular network comprising DNA metabolism and neuronal development, consistent with targets of oxidative stress-inducing agents. We confirmed associations of seven candidate genes associated with natural variation in oxidative stress resistance through mutational analysis.

Conclusions

We identified novel candidate genes associated with variation in resistance to oxidative stress that have context-dependent effects. These results form the basis for future translational studies to identify oxidative stress susceptibility/resistance genes that are evolutionary conserved and might play a role in human disease.     

Glutamate decarboxylase (GAD) autoantibody epitope shift during the first year of type 1 diabetes.

Autoantibodies in Type 1 diabetes patients may differentiate between glutamate decarboxylase (GAD65) cloned from human, mouse and rat with a significant better binding to the human antigen. A subgroup of 15% (27/183) patients showed significantly better binding to rodent than to human GAD65. The aim of this study was to determine whether the autoantibody specificity would remain anti-rodent during longitudinal follow-up for one year. We observed 1) that the average slope of the difference between human and mouse GAD65 autoantibodies binding increased between onset and after one year, which demonstrates reduced binding to rodent GAD65 and 2) that, in a group followed every third month, 9/11 (80%) children with rodent specific GAD65 autoantibodies at onset converted within one year to preference against human GAD65. This shift in preference was confirmed by significantly lower EC50 values in the initially anti-rodent GAD65 autoantibodies compared to samples taken one year after clinical diagnosis as determined in displacement studies with unlabeled human GAD65. We speculate that the evolution of GAD65 autoantibodies in Type 1 diabetes includes reactivity to a non-human GAD65 N-terminal end conformation. Progression towards Type 1 diabetes is, however, associated with a maturation of the immune response towards human GAD65 autoreactivity.     

Gender-dependent association of type 2 diabetes with the vasoactive intestinal peptide receptor 1

Type 2 diabetes is characterized by an inadequate pancreatic beta-cell response to the progressive insulin resistance. Its pathogenesis is complex and has been connected with a state of preclinical chronic inflammation. Vasoactive intestinal peptide (VIP) and its receptors play a relevant role in the homeostasis of insulin secretion as well as in the control of inflammation. In particular, VIP receptor 1 (VPAC1) has been found to be down-modulated during inflammation, and to be associated with several diseases. The objective of this study was to compare the distribution of SNPs mapping in the VIP receptor 1 gene in cases with type 2 diabetes and matched controls. Seven hundred cases with type 2 diabetes (423 males and 277 females) and 830 random controls (419 males and 411 females) were analyzed for the distribution of three common SNPs mapping in the VPAC1 gene. The results show a significantly different genotype distribution of the SNP rs9677 in the 3’-UTR of VPAC1 in female cases with type 2 diabetes compared to gender-matched controls (ptrend = 6 × 10^− 4). The rs9677 CC genotype confers the highest risk (OR: 2.1) and correlates with worse clinical parameters such as higher level of total cholesterol, higher LDL/HDL ratio and a higher HbA1c concentration. The genetic association reported here indicates that VIP/VPAC1 signaling can be a relevant pathway in the pathogenesis of type 2 diabetes in females suggesting that at least some aspects of the genetic predisposition to this disease can be gender-specific.     

Genome-wide association analysis for lethal brachycephalic-like facial dysmorphia in Labrador Retrievers

A GWAS was performed for inborn X-linked facial dysmorphia with severe growth retardation in Labrador Retrievers. This lethal condition was mapped on the X chromosome at 17–21 Mb and supported by eight SNPs in complete LD. Dams of affected male puppies were heterozygous for the significantly associated SNPs and male affected puppies carried the associated alleles hemizygously. In the near vicinity to the associated region, RPS6KA3 was identified as a candidate gene causing facial dysmorphia in humans and mice known as Coffin–Lowry syndrome. Haplotype analysis showed significant association with the phenotypes of all 18 animals under study. This haplotype was validated through normal male progeny from a dam with the not-associated haplotype on both X chromosomes but male affected full-sibs with the associated haplotype.     

Identification and functional analysis of CBLB mutations in type 1 diabetes

Casitas B-lineage lymphoma b (Cblb) is a negative regulator of T-cell activation and dysfunction of Cblb in rats and mice results in autoimmunity. In particular, a nonsense mutation in Cblb has been identified in a rat model of autoimmune type 1 diabetes. To clarify the possible involvement of CBLB mutation in type 1 diabetes in humans, we performed mutation screening of CBLB and characterized functional properties of the mutations in Japanese subjects. Six missense mutations (A155V, F328L, N466D, K837R, T882A, and R968L) were identified in one diabetic subject each, excepting N466D. Of these mutations, F328L showed impaired suppression of T-cell activation and was a loss-of-function mutation. These data suggest that the F328L mutation is involved in the development of autoimmune diseases including type 1 diabetes, and also provide insight into the structure-function relationship of CBLB protein.     

Genome-wide association analysis of circulating vitamin D levels in children with asthma

Vitamin D deficiency is becoming more apparent in many populations. Genetic factors may play a role in the maintenance of vitamin D levels. The objective of this study was to perform a genome-wide analysis (GWAS) of vitamin D levels, including replication of prior GWAS results. We measured 25-hydroxyvitamin D (25(OH)D) levels in serum collected at the time of enrollment and at year 4 in 572 Caucasian children with asthma, who were part of a multi-center clinical trial, the Childhood Asthma Management Program. Replication was performed in a second cohort of 592 asthmatics from Costa Rica and a third cohort of 516 Puerto Rican asthmatics. In addition, we attempted replication of three SNPs that were previously identified in a large GWAS of Caucasian individuals. The setting included data from a clinical trial of childhood asthmatics and two cohorts of asthmatics recruited for genetic studies of asthma. The main outcome measure was circulating 25(OH)D levels. The 25(OH)D levels at the two time-points were only modestly correlated with each other (intraclass correlation coefficient?=?0.33) in the CAMP population. We identified SNPs that were nominally associated with 25(OH)D levels at two time-points in CAMP, and replicated four SNPs in the Costa Rican cohort: rs11002969, rs163221, rs1678849, and rs4864976. However, these SNPs were not significantly associated with 25(OH)D levels in a third population of Puerto Rican asthmatics. We were able to replicate the SNP with the strongest effect, previously reported in a large GWAS: rs2282679 (GC), and we were able to replicate another SNP, rs10741657 (CYP2R1), to a lesser degree. We were able to replicate two of three prior significant findings in a GWAS of 25(OH)D levels. Other SNPs may be additionally associated with 25(OH)D levels in certain populations.