Obesity is heritable and predisposes to many diseases. To understand the genetic basis of obesity better, here we conduct a genome-wide association study and Metabochip meta-analysis of body mass index (BMI), a measure commonly used to define obesity and assess adiposity, in up to 339,224 individuals. This analysis identifies 97 BMI-associated loci (P < 5 × 10−8), 56 of which are novel. Five loci demonstrate clear evidence of several independent association signals, and many loci have significant effects on other metabolic phenotypes. The 97 loci account for ~2.7% of BMI variation, and genome-wide estimates suggest that common variation accounts for >20% of BMI variation. Pathway analyses provide strong support for a role of the central nervous system in obesity susceptibility and implicate new genes and pathways, including those related to synaptic function, glutamate signalling, insulin secretion/action, energy metabolism, lipid biology and adipogenesis.
The QT interval, an electrocardiographic measure reflecting myocardial repolarization, is a heritable trait. QT prolongation is a risk factor for ventricular arrhythmias and sudden cardiac death (SCD) and could indicate the presence of the potentially lethal Mendelian Long QT Syndrome (LQTS). Using a genome-wide association and replication study in up to 100,000 individuals we identified 35 common variant QT interval loci, that collectively explain ∼8-10% of QT variation and highlight the importance of calcium regulation in myocardial repolarization. Rare variant analysis of 6 novel QT loci in 298 unrelated LQTS probands identified coding variants not found in controls but of uncertain causality and therefore requiring validation. Several newly identified loci encode for proteins that physically interact with other recognized repolarization proteins. Our integration of common variant association, expression and orthogonal protein-protein interaction screens provides new insights into cardiac electrophysiology and identifies novel candidate genes for ventricular arrhythmias, LQTS,and SCD.
genome-wide association study; QT interval; Long QT Syndrome; sudden cardiac death; myocardial repolarization; arrhythmias
SIRT1 and FOXO1 interact with each other in multiple pathways regulating aging, metabolism and resistance to oxidative stress and control different pathways involved in atherosclerotic process. It is not known, if genetic polymorphisms (SNPs) at the SIRT1 and FOXO1 have an influence on carotid atherosclerosis.
Intima-media thickness (IMT) was measured on the common and internal carotid arteries. Morphological alterations of the carotid arteries and size of these alterations were included in the B-score grading on a five point scale. Eleven SNPs at SIRT1 and FOXO1 gene loci were genotyped in the SAPHIR cohort (n = 1742). The association of each SNP with common carotid IMT, internal carotid IMT and B-score was analyzed using linear regression models.
A significant association was found between common carotid IMT and two SNPs at FOXO1 - rs10507486, rs2297627 (beta = -0.00168, p = 0.0007 and beta = -0.00144, p = 0.0008 respectively) and at least a trend for rs12413112 at SIRT1 (beta = 0.00177, p = 0.0157) using an additive model adjusting for age and sex. Additional adjustment for traditional cardiovascular risk factors and markers (BMI, smoking status, hypertension, total cholesterol, HDL-cholesterol, hsCRP) even improved the strength of this association (p = 0.0037 for SIRT1 and p = 0.0002 for both SNPs at FOXO1). Analysis for internal carotis IMT and B-score did not reveal any significant association. One haplotype in FOXO1 showed a moderate effect on common carotid IMT and B-score in comparison to the reference haplotype of this gene. Several SNPs within SIRT1 showed differential effects for men and women with higher effect sizes for women: rs3740051 on all three investigated phenotypes (interaction p-value < 0.0069); rs2236319 on common and internal carotid IMT (interaction p-value < 0.0083), rs10823108, rs2273773 on common carotid IMT and rs1467568 on B-score (interaction p-value = 0.0007). The latter was significant in women only (betawomen = 0.111, pwomen = 0.00008; betamen = -0.009, pmen = 0.6464).
This study demonstrated associations of genetic variations at the SIRT1 and FOXO1 loci with carotid atherosclerosis and highlighted the need for further investigation by functional studies.
Adiponectin is strongly inversely associated with insulin resistance and type 2 diabetes, but its causal role remains controversial. We used a Mendelian randomization approach to test the hypothesis that adiponectin causally influences insulin resistance and type 2 diabetes. We used genetic variants at the ADIPOQ gene as instruments to calculate a regression slope between adiponectin levels and metabolic traits (up to 31,000 individuals) and a combination of instrumental variables and summary statistics–based genetic risk scores to test the associations with gold-standard measures of insulin sensitivity (2,969 individuals) and type 2 diabetes (15,960 case subjects and 64,731 control subjects). In conventional regression analyses, a 1-SD decrease in adiponectin levels was correlated with a 0.31-SD (95% CI 0.26–0.35) increase in fasting insulin, a 0.34-SD (0.30–0.38) decrease in insulin sensitivity, and a type 2 diabetes odds ratio (OR) of 1.75 (1.47–2.13). The instrumental variable analysis revealed no evidence of a causal association between genetically lower circulating adiponectin and higher fasting insulin (0.02 SD; 95% CI −0.07 to 0.11; N = 29,771), nominal evidence of a causal relationship with lower insulin sensitivity (−0.20 SD; 95% CI −0.38 to −0.02; N = 1,860), and no evidence of a relationship with type 2 diabetes (OR 0.94; 95% CI 0.75–1.19; N = 2,777 case subjects and 13,011 control subjects). Using the ADIPOQ summary statistics genetic risk scores, we found no evidence of an association between adiponectin-lowering alleles and insulin sensitivity (effect per weighted adiponectin-lowering allele: −0.03 SD; 95% CI −0.07 to 0.01; N = 2,969) or type 2 diabetes (OR per weighted adiponectin-lowering allele: 0.99; 95% CI 0.95–1.04; 15,960 case subjects vs. 64,731 control subjects). These results do not provide any consistent evidence that interventions aimed at increasing adiponectin levels will improve insulin sensitivity or risk of type 2 diabetes.
Structural genetic variants as short tandem repeats (STRs) are not targeted in SNP-based association studies and thus, their possible association signals are missed. We systematically searched for STRs in gene regions known to contribute to total cholesterol, HDL cholesterol, LDL cholesterol and triglyceride levels in two independent studies (KORA F4, n = 2553 and SAPHIR, n = 1648), resulting in 16 STRs that were finally evaluated. In a combined dataset of both studies, the sum of STR alleles was regressed on each phenotype, adjusted for age and sex. The association analyses were repeated for SNPs in a 200 kb region surrounding the respective STRs in the KORA F4 Study. Three STRs were significantly associated with total cholesterol (within LDLR, the APOA1/C3/A4/A5/BUD13 gene region and ABCG5/8), five with HDL cholesterol (3 within CETP, one in LPL and one inAPOA1/C3/A4/A5/BUD13), three with LDL cholesterol (LDLR, ABCG5/8 and CETP) and two with triglycerides (APOA1/C3/A4/A5/BUD13 and LPL). None of the investigated STRs, however, showed a significant association after adjusting for the lead or adjacent SNPs within that gene region. The evaluated STRs were found to be well tagged by the lead SNP within the respective gene regions. Therefore, the STRs reflect the association signals based on surrounding SNPs. In conclusion, none of the STRs contributed additionally to the SNP-based association signals identified in GWAS on lipid traits.
Oxidative stress seems to be present in patients with polycystic ovary syndrome (PCOS). The aim of this study was to evaluate the correlation between characteristics of PCOS and serum concentrations of afamin, a novel binding protein for the antioxidant vitamin E. A total of 85 patients with PCOS and 76 control subjects were investigated in a pilot cross-sectional study design between 2009 and 2013 in the University Hospital of Essen, Germany. Patients with PCOS were diagnosed according to the Rotterdam ESHRE/ASRM-sponsored PCOS Consensus Workshop Group. Afamin and diagnostic parameters of PCOS were determined at early follicular phase. Afamin concentrations were significantly higher in patients with PCOS than in controls (odds ratio (OR) for a 10 mg/ml increase in afamin=1.3, 95% CI=1.08–1.58). This difference vanished in a model adjusting for age, BMI, free testosterone index (FTI), and sex hormone-binding globulin (SHBG) (OR=1.05, 95% CI=0.80–1.38). In patients with PCOS, afamin correlated significantly with homeostatic model assessment-insulin resistance (HOMA-IR), fasting glucose, BMI, FTI, and SHBG (P<0.001), but in a multivariate linear model, only HOMA-IR remained significantly associated with afamin (P=0.001). No correlation was observed between afamin and androgens, LH, FSH, LH/FSH ratio, antral follicle count, ovarian volume, or anti-Müllerian hormone. In conclusion, elevated afamin values may indicate a state of oxidative stress and inflammation, strongly associated with IR and offering an indicator of impaired glucose tolerance in patients with PCOS irrespective of obesity.
vitamin E-binding protein; afamin; polycystic ovary syndrome; oxidative stress; insulin resistance
The relevance of lipoprotein(a) [Lp(a)] concentrations and low-molecular-weight (LMW) apo(a) phenotypes in peripheral arterial disease (PAD) has only been investigated by few studies. Therefore, we analysed this association in three independent cohorts and performed a Mendelian Randomization approach using instrumental variable regression.
Methods and results
Lp(a) concentrations, apo(a) phenotypes, and one SNP in the LPA gene (rs10455872) were measured in the CAVASIC study, including 241 male patients with intermittent claudication and 246 age- and diabetes-matched controls as well as in the two population-based studies KORA F3 (n = 3184) and KORA F4 (n = 3080). In KORA F3/F4, 109/80 persons suffered from intermittent claudication, 200/144 from PAD, and 128/103 showed an ankle–brachial index (ABI) <0.9. In CAVASIC, adjusted logistic regression analyses revealed significant associations between an increase of log-Lp(a) per one standard deviation (SD) (OR = 1.28, P = 0.02) as well as LMW apo(a) phenotypes and symptomatic PAD (OR = 1.65, P = 0.03). Linear regression models with continuous ABI showed a significant association in the combined analyses of KORA F3/F4: an increase in log-Lp(a) per one SD (β = −0.006, P = 0.005) and the presence of LMW apo(a) phenotypes (β = −0.011, P = 0.02) or the minor allele of rs10455872 (ß = −0.016, P = 0.03) were associated with a decrease in ABI in the fully adjusted linear and instrumental variable regression models.
Analyses in three independent populations showed significant associations of Lp(a) concentrations, LMW apo(a) phenotypes, and rs10455872 with PAD. This points to a causal relationship between Lp(a) and PAD since the genetically determined apo(a) phenotypes and SNP alleles are indeed associated with PAD.
Lp(a) concentrations; Apolipoprotein(a) phenotypes; Peripheral arterial disease; Ankle-brachial index; Mendelian randomization; Causality
Both genome-wide association studies and candidate gene studies have reported that the major determinant of plasma levels of the Lipoprotein (a) [Lp(a)] reside within the LPA locus on chromosome 6. We have used data from the Human CVD bead chip to explore the contribution of other candidate genes determining Lp(a) levels.
48,032 single nucleotide polymorphisms (SNPs) from the Illumina Human CVD bead chip were genotyped in 5,059 participants of the Whitehall II study (WHII) of randomly ascertained healthy men and women. SNPs showing association with Lp(a) levels of p< 10−4 outside the LPA locus were selected for replication in a total of an additional 9,463 participants of five European based studies (EAS, EPIC-Norfolk, NPHSII, PROCARDIS, and SAPHIR)
In Whitehall II, apart from the LPA locus (where p values for several SNPs were < 10−30) there was significant association at four loci GALNT2, FABP1, PPARGC1A and TNFRSFF11A. However, a meta-analysis of the six studies did not confirm any of these findings.
Results from this meta analysis of 14,522 participants revealed no candidate genes from the Human CVD bead chip outside the LPA locus to have an effect on Lp(a) levels. Further studies with genome-wide and denser SNP coverage are required to confirm or refute this finding.
Lipoprotein(a); LPA; Illumina Human CVD bead chip; genetic association
Approaches exploiting extremes of the trait distribution may reveal novel loci for common traits, but it is unknown whether such loci are generalizable to the general population. In a genome-wide search for loci associated with upper vs. lower 5th percentiles of body mass index, height and waist-hip ratio, as well as clinical classes of obesity including up to 263,407 European individuals, we identified four new loci (IGFBP4, H6PD, RSRC1, PPP2R2A) influencing height detected in the tails and seven new loci (HNF4G, RPTOR, GNAT2, MRPS33P4, ADCY9, HS6ST3, ZZZ3) for clinical classes of obesity. Further, we show that there is large overlap in terms of genetic structure and distribution of variants between traits based on extremes and the general population and little etiologic heterogeneity between obesity subgroups.
High serum uric acid levels are associated with gout, atherosclerosis and cardiovascular disease. Three genes (SLC2A9, ABCG2, and SLC17A3) were reported to be involved in the regulation of uric acid levels.
Design and Methods: SNPs rs2231142 (ABCG2) and rs1165205 (SLC17A3) were genotyped in three cohorts (n = 4492) and combined with previously genotyped SNPs within SLC2A9 (rs6855911, rs7442295, rs6449213, rs12510549).
Each copy of the minor allele decreased uric acid levels by 0.30–0.38 mg/dL for SLC2A9 (p values: 10−20–10−36) and increased levels by 0.34 mg/dL for ABCG2 (p = 1.1×10−16). SLC17A3 influenced uric acid levels only modestly. Together the SNPs showed graded associations with uric acid levels of 0.111 mg/dL per risk allele (p = 3.8×10−42). In addition, we observed a sex-specific interaction of age with the association of SLC2A9 SNPs with uric acid levels, where increasing age strengthened the association of SNPs in women and decreased the association in men.
Genetic variants within SLC2A9, ABCG2 and SLC17A3 show highly significant associations with uric acid levels, and for SNPs within SLC2A9 this association is strongly modified by age and sex.
Epidemiology; Genetics; Uric acid; Copy number variation; Sex-specific effect; Genetic risk score
Two recent genome-wide association studies identified the liver-expressed transmembrane protein adiponutrin to be associated with liver-related phenotypes such as nonalcoholic fatty liver disease and liver function enzymes. These associations were not uniformly reported for various ethnicities. The aim of this study was to investigate a common nonsynonymous variant within adiponutrin (rs738409, exon 3) with parameters of liver function in three independent West-Eurasian study populations including a total of 4290 participants.
The study was performed in 1) the population-based Bruneck Study (n=783), 2) the SAPHIR Study from Austria based on a healthy working population (n=1705), and the Utah Obesity Case-Control Study including a group of 1019 severely obese individuals (average BMI 46.0 kg/m2) and 783 controls from the same geographical region of Utah. Liver enzymes measured were alanine-aminotransferase (ALT), aspartate-aminotransferase (AST) and gamma-glutamyl transferase (GGT).
Results and Discussion
We found a strong recessive association of this polymorphism with age- and gender-adjusted ALT and AST levels: being homozygous for the minor allele resulted in a highly significant increase of ALT levels of 3.53 U/L (p=1.86×10−9) and of AST levels of 2.07 U/L (p=9.58×10−6), respectively. The associations were consistently found in all three study populations. In conclusion, the highly significant associations of this transversion polymorphism within the adiponutrin gene with increased ALT and AST levels support a role for adiponutrin as a susceptibility gene for hepatic dysfunction.
PNPLA3; rs738409; genetic association; hepatic dysfunction
Given the anthropometric differences between men and women and previous evidence of sex-difference in genetic effects, we conducted a genome-wide search for sexually dimorphic associations with height, weight, body mass index, waist circumference, hip circumference, and waist-to-hip-ratio (133,723 individuals) and took forward 348 SNPs into follow-up (additional 137,052 individuals) in a total of 94 studies. Seven loci displayed significant sex-difference (FDR<5%), including four previously established (near GRB14/COBLL1, LYPLAL1/SLC30A10, VEGFA, ADAMTS9) and three novel anthropometric trait loci (near MAP3K1, HSD17B4, PPARG), all of which were genome-wide significant in women (P<5×10−8), but not in men. Sex-differences were apparent only for waist phenotypes, not for height, weight, BMI, or hip circumference. Moreover, we found no evidence for genetic effects with opposite directions in men versus women. The PPARG locus is of specific interest due to its role in diabetes genetics and therapy. Our results demonstrate the value of sex-specific GWAS to unravel the sexually dimorphic genetic underpinning of complex traits.
Men and women differ substantially regarding height, weight, and body fat. Interestingly, previous work detecting genetic effects for waist-to-hip ratio, to assess body fat distribution, has found that many of these showed sex-differences. However, systematic searches for sex-differences in genetic effects have not yet been conducted. Therefore, we undertook a genome-wide search for sexually dimorphic genetic effects for anthropometric traits including 133,723 individuals in a large meta-analysis and followed promising variants in further 137,052 individuals, including a total of 94 studies. We identified seven loci with significant sex-difference including four previously established (near GRB14/COBLL1, LYPLAL1/SLC30A10, VEGFA, ADAMTS9) and three novel anthropometric trait loci (near MAP3K1, HSD17B4, PPARG), all of which were significant in women, but not in men. Of interest is that sex-difference was only observed for waist phenotypes, but not for height or body-mass-index. We found no evidence for sex-differences with opposite effect direction for men and women. The PPARG locus is of specific interest due to its link to diabetes genetics and therapy. Our findings demonstrate the importance of investigating sex differences, which may lead to a better understanding of disease mechanisms with a potential relevance to treatment options.
Candidate gene association studies for peripheral artery disease (PAD), including subclinical disease assessed with the ankle-brachial index (ABI), have been limited by the modest number of genes examined. We conducted a two stage meta-analysis of ~50,000 SNPs across ~2100 candidate genes to identify genetic variants for ABI.
Methods and results
We studied subjects of European ancestry from 8 studies (n = 21,547, 55% women, mean age 44–73 years) and African American ancestry from 5 studies (n = 7267, 60% women, mean age 41–73 years) involved in the candidate gene association resource (CARe) consortium. In each ethnic group, additive genetic models were used (with each additional copy of the minor allele corresponding to the given beta) to test each SNP for association with continuous ABI (excluding ABI > 1.40) and PAD (defined as ABI < 0.90) using linear or logistic regression with adjustment for known PAD risk factors and population stratification. We then conducted a fixed-effects inverse-variance weighted meta-analyses considering a p < 2 × 10−6 to denote statistical significance.
In the European ancestry discovery meta-analyses, rs2171209 in SYTL3 (β = −0.007, p = 6.02 × 10−7) and rs290481 in TCF7L2 (β = −0.008, p = 7.01 × 10−7) were significantly associated with ABI. None of the SNP associations for PAD were significant, though a SNP in CYP2B6 (p = 4.99 × 10−5) was among the strongest associations. These 3 genes are linked to key PAD risk factors (lipoprotein(a), type 2 diabetes, and smoking behavior, respectively). We sought replication in 6 population-based and 3 clinical samples (n = 15,440) for rs290481 and rs2171209. However, in the replication stage (rs2171209, p = 0.75; rs290481, p = 0.19) and in the combined discovery and replication analysis the SNP–ABI associations were no longer significant (rs2171209, p = 1.14 × 10−3; rs290481, p = 8.88 × 10−5). In African Americans, none of the SNP associations for ABI or PAD achieved an experiment-wide level of significance.
Genetic determinants of ABI and PAD remain elusive. Follow-up of these preliminary findings may uncover important biology given the known gene-risk factor associations. New and more powerful approaches to PAD gene discovery are warranted.
Ankle brachial index; Peripheral artery disease; Genetics; Candidate gene array; Meta-analysis; Ethnicity
Smoking is a leading global cause of disease and mortality1. We performed a genomewide meta-analytic association study of smoking-related behavioral traits in a total sample of 41,150 individuals drawn from 20 disease, population, and control cohorts. Our analysis confirmed an effect on smoking quantity (SQ) at a locus on 15q25 (P=9.45e-19) that includes three genes encoding neuronal nicotinic acetylcholine receptor subunits (CHRNA5, CHRNA3, CHRNB4). We used data from the 1000 Genomes project to investigate the region using imputation, which allowed analysis of virtually all common variants in the region and offered a five-fold increase in coverage over the HapMap. This increased the spectrum of potentially causal single nucleotide polymorphisms (SNPs), which included a novel SNP that showed the highest significance, rs55853698, located within the promoter region of CHRNA5. Conditional analysis also identified a secondary locus (rs6495308) in CHRNA3.
Genetic determinants of peripheral arterial disease (PAD) remain largely unknown. To identify genetic variants associated with the ankle-brachial index (ABI), a noninvasive measure of PAD, we conducted a meta-analysis of genome-wide association study data from 21 population-based cohorts.
Methods and Results
Continuous ABI and PAD (ABI≤0.9) phenotypes adjusted for age and sex were examined. Each study conducted genotyping and imputed data to the ~2.5 million SNPs in HapMap. Linear and logistic regression models were used to test each SNP for association with ABI and PAD using additive genetic models. Study-specific data were combined using fixed-effects inverse variance weighted meta-analyses. There were a total of 41,692 participants of European ancestry (~60% women, mean ABI 1.02 to 1.19), including 3,409 participants with PAD and with GWAS data available. In the discovery meta-analysis, rs10757269 on chromosome 9 near CDKN2B had the strongest association with ABI (β= −0.006, p=2.46x10−8). We sought replication of the 6 strongest SNP associations in 5 population-based studies and 3 clinical samples (n=16,717). The association for rs10757269 strengthened in the combined discovery and replication analysis (p=2.65x10−9). No other SNP associations for ABI or PAD achieved genome-wide significance. However, two previously reported candidate genes for PAD and one SNP associated with coronary artery disease (CAD) were associated with ABI : DAB21P (rs13290547, p=3.6x10−5); CYBA (rs3794624, p=6.3x10−5); and rs1122608 (LDLR, p=0.0026).
GWAS in more than 40,000 individuals identified one genome-wide significant association on chromosome 9p21 with ABI. Two candidate genes for PAD and 1 SNP for CAD are associated with ABI.
cohort study; genetic association; genome-wide association study; meta-analysis; peripheral vascular disease
The late endosomal LAMTOR complex serves as a convergence point for both the RAF/MEK/ERK and the PI3K/AKT/mTOR pathways. Interestingly, both of these signalling cascades play a significant role in the aetiology of breast cancer. Our aim was to address the possible role of genetic polymorphisms in LAMTOR2 and LAMTOR3 as genetic risk factors for breast cancer.
We sequenced the exons and exon–intron boundaries of LAMTOR2 (p14) and LAMTOR3 (MP1) in 50 prospectively collected pairs of cancerous tissue and blood samples from breast cancer patients and compared their genetic variability. We found one single nucleotide polymorphism (SNP) in LAMTOR2 (rs7541) and two SNPs in LAMTOR3 (rs2298735 and rs148972953) in both tumour and blood samples, but no somatic mutations in cancerous tissues. In addition, we genotyped all three SNPs in 296 samples from the Risk Prediction of Breast Cancer Metastasis Study and found evidence of a genetic association between rs148972953 and oestrogen (ER) and progesterone receptor negative status (PR) (ER: OR = 3.60 (1.15–11.28); PR: OR = 4.27 (1.43–12.72)). However, when we additionally genotyped rs148972953 in the MARIE study including 2,715 breast cancer cases and 5,216 controls, we observed neither a difference in genotype frequencies between patients and controls nor was the SNP associated with ER or PR. Finally, all three SNPs were equally frequent in breast cancer samples and female participants (n = 640) of the population-based SAPHIR Study.
The identified polymorphisms in LAMTOR2 and LAMTOR3 do not seem to play a relevant role in breast cancer. Our work does not exclude a role of other not yet identified SNPs or that the here annotated polymorphism may in fact play a relevant role in other diseases. Our results underscore the importance of replication in association studies.
Adiponectin serum concentrations are an important biomarker in cardiovascular epidemiology with heritability etimates of 30–70%. However, known genetic variants in the adiponectin gene locus (ADIPOQ) account for only 2%–8% of its variance. As transcription factors are thought to play an under-acknowledged role in carrying functional variants, we hypothesized that genetic polymorphisms in genes coding for the main transcription factors for the ADIPOQ promoter influence adiponectin levels. Single nucleotide polymorphisms (SNPs) at these genes were selected based on the haplotype block structure and previously published evidence to be associated with adiponectin levels. We performed association analyses of the 24 selected SNPs at forkhead box O1 (FOXO1), sterol-regulatory-element-binding transcription factor 1 (SREBF1), sirtuin 1 (SIRT1), peroxisome-proliferator-activated receptor gamma (PPARG) and transcription factor activating enhancer binding protein 2 beta (TFAP2B) gene loci with adiponectin levels in three different European cohorts: SAPHIR (n = 1742), KORA F3 (n = 1636) and CoLaus (n = 5355). In each study population, the association of SNPs with adiponectin levels on log-scale was tested using linear regression adjusted for age, sex and body mass index, applying both an additive and a recessive genetic model. A pooled effect size was obtained by meta-analysis assuming a fixed effects model. We applied a significance threshold of 0.0033 accounting for the multiple testing situation. A significant association was only found for variants within SREBF1 applying an additive genetic model (smallest p-value for rs1889018 on log(adiponectin) = 0.002, β on original scale = −0.217 µg/ml), explaining ∼0.4% of variation of adiponectin levels. Recessive genetic models or haplotype analyses of the FOXO1, SREBF1, SIRT1, TFAPB2B genes or sex-stratified analyses did not reveal additional information on the regulation of adiponectin levels. The role of genetic variations at the SREBF1 gene in regulating adiponectin needs further investigation by functional studies.
Group 14 of Genetic Analysis Workshop 17 examined several issues related to analysis of complex traits using DNA sequence data. These issues included novel methods for analyzing rare genetic variants in an aggregated manner (often termed collapsing rare variants), evaluation of various study designs to increase power to detect effects of rare variants, and the use of machine learning approaches to model highly complex heterogeneous traits. Various published and novel methods for analyzing traits with extreme locus and allelic heterogeneity were applied to the simulated quantitative and disease phenotypes. Overall, we conclude that power is (as expected) dependent on locus-specific heritability or contribution to disease risk, large samples will be required to detect rare causal variants with small effect sizes, extreme phenotype sampling designs may increase power for smaller laboratory costs, methods that allow joint analysis of multiple variants per gene or pathway are more powerful in general than analyses of individual rare variants, population-specific analyses can be optimal when different subpopulations harbor private causal mutations, and machine learning methods may be useful for selecting subsets of predictors for follow-up in the presence of extreme locus heterogeneity and large numbers of potential predictors.
rare variants; LASSO; machine learning; random forests; logic regression; binary trees; Poisson regression; ISIS; classification trees; meta-analysis; extreme sampling
Iron-refractory iron deficiency anaemia (IRIDA) is a rare disorder which was linked to mutations in two genes (SLC11A2 and TMPRSS6). Common polymorphisms within these genes were associated with serum iron levels. We identified a family of Serbian origin with asymptomatic non-consanguineous parents with three of four children presenting with IRIDA not responding to oral but to intravenous iron supplementation. After excluding all known causes responsible for iron deficiency anaemia we searched for mutations in SLC11A2 and TMPRSS6 that could explain the severe anaemia in these children.
We sequenced the exons and exon–intron boundaries of SLC11A2 and TMPRSS6 in all six family members. Thereby, we found seven known and fairly common SNPs, but no new mutation. We then genotyped these seven SNPs in the population-based SAPHIR study (n = 1,726) and performed genetic association analysis on iron and ferritin levels. Only two SNPs, which were top-hits from recent GWAS on iron and ferritin, exhibited an effect on iron and ferritin levels in SAPHIR. Six SAPHIR participants carrying the same TMPRSS6 genotypes and haplotype-pairs as one anaemic son showed lower ferritin and iron levels than the average. One individual exhibiting the joint SLC11A2/TMPRSS6 profile of the anaemic son had iron and ferritin levels lying below the 5th percentile of the population's iron and ferritin level distribution. We then checked the genotype constellations in the Nijmegen Biomedical Study (n = 1,832), but the profile of the anaemic son did not occur in this population.
We cannot exclude a gene-gene interaction between SLC11A2 and TMPRSS6, but we can also not confirm it. As in this case candidate gene sequencing did not reveal causative rare mutations, the samples will be subjected to whole exome sequencing.
Circulating levels of adiponectin, a hormone produced predominantly by adipocytes, are highly heritable and are inversely associated with type 2 diabetes mellitus (T2D) and other metabolic traits. We conducted a meta-analysis of genome-wide association studies in 39,883 individuals of European ancestry to identify genes associated with metabolic disease. We identified 8 novel loci associated with adiponectin levels and confirmed 2 previously reported loci (P = 4.5×10−8–1.2×10−43). Using a novel method to combine data across ethnicities (N = 4,232 African Americans, N = 1,776 Asians, and N = 29,347 Europeans), we identified two additional novel loci. Expression analyses of 436 human adipocyte samples revealed that mRNA levels of 18 genes at candidate regions were associated with adiponectin concentrations after accounting for multiple testing (p<3×10−4). We next developed a multi-SNP genotypic risk score to test the association of adiponectin decreasing risk alleles on metabolic traits and diseases using consortia-level meta-analytic data. This risk score was associated with increased risk of T2D (p = 4.3×10−3, n = 22,044), increased triglycerides (p = 2.6×10−14, n = 93,440), increased waist-to-hip ratio (p = 1.8×10−5, n = 77,167), increased glucose two hours post oral glucose tolerance testing (p = 4.4×10−3, n = 15,234), increased fasting insulin (p = 0.015, n = 48,238), but with lower in HDL-cholesterol concentrations (p = 4.5×10−13, n = 96,748) and decreased BMI (p = 1.4×10−4, n = 121,335). These findings identify novel genetic determinants of adiponectin levels, which, taken together, influence risk of T2D and markers of insulin resistance.
Serum adiponectin levels are highly heritable and are inversely correlated with the risk of type 2 diabetes (T2D), coronary artery disease, stroke, and several metabolic traits. To identify common genetic variants associated with adiponectin levels and risk of T2D and metabolic traits, we conducted a meta-analysis of genome-wide association studies of 45,891 multi-ethnic individuals. In addition to confirming that variants at the ADIPOQ and CDH13 loci influence adiponectin levels, our analyses revealed that 10 new loci also affecting circulating adiponectin levels. We demonstrated that expression levels of several genes in these candidate regions are associated with serum adiponectin levels. Using a powerful novel method to assess the contribution of the identified variants with other traits using summary-level results from large-scale GWAS consortia, we provide evidence that the risk alleles for adiponectin are associated with deleterious changes in T2D risk and metabolic syndrome traits (triglycerides, HDL, post-prandial glucose, insulin, and waist-to-hip ratio), demonstrating that the identified loci, taken together, impact upon metabolic disease.
The common disease/rare variant hypothesis predicts that rare variants with large effects will have a strong impact on corresponding phenotypes. Therefore it is assumed that rare functional variants are enriched in the extremes of the phenotype distribution. In this analysis of the Genetic Analysis Workshop 17 data set, my aim is to detect genes with rare variants that are associated with quantitative traits using two general approaches: analyzing the association with the complete distribution of values by means of linear regression and using statistical tests based on the tails of the distribution (bottom 10% of values versus top 10%). Three methods are used for this extreme phenotype approach: Fisher’s exact test, weighted-sum method, and beta method. Rare variants were collapsed on the gene level. Linear regression including all values provided the highest power to detect rare variants. Of the three methods used in the extreme phenotype approach, the beta method performed best. Furthermore, the sample size was enriched in this approach by adding additional samples with extreme phenotype values. Doubling the sample size using this approach, which corresponds to only 40% of sample size of the original continuous trait, yielded a comparable or even higher power than linear regression. If samples are selected primarily for sequencing, enriching the analysis by gathering a greater proportion of individuals with extreme values in the phenotype of interest rather than in the general population leads to a higher power to detect rare variants compared to analyzing a population-based sample with equivalent sample size.
Recent genome-wide association (GWA) studies described 95 loci controlling serum lipid levels. These common variants explain ∼25% of the heritability of the phenotypes. To date, no unbiased screen for gene–environment interactions for circulating lipids has been reported. We screened for variants that modify the relationship between known epidemiological risk factors and circulating lipid levels in a meta-analysis of genome-wide association (GWA) data from 18 population-based cohorts with European ancestry (maximum N = 32,225). We collected 8 further cohorts (N = 17,102) for replication, and rs6448771 on 4p15 demonstrated genome-wide significant interaction with waist-to-hip-ratio (WHR) on total cholesterol (TC) with a combined P-value of 4.79×10−9. There were two potential candidate genes in the region, PCDH7 and CCKAR, with differential expression levels for rs6448771 genotypes in adipose tissue. The effect of WHR on TC was strongest for individuals carrying two copies of G allele, for whom a one standard deviation (sd) difference in WHR corresponds to 0.19 sd difference in TC concentration, while for A allele homozygous the difference was 0.12 sd. Our findings may open up possibilities for targeted intervention strategies for people characterized by specific genomic profiles. However, more refined measures of both body-fat distribution and metabolic measures are needed to understand how their joint dynamics are modified by the newly found locus.
Circulating serum lipids contribute greatly to the global health by affecting the risk for cardiovascular diseases. Serum lipid levels are partly inherited, and already 95 loci affecting high- and low-density lipoprotein cholesterol, total cholesterol, and triglycerides have been found. Serum lipids are also known to be affected by multiple epidemiological risk factors like body composition, lifestyle, and sex. It has been hypothesized that there are loci modifying the effects between risk factors and serum lipids, but to date only candidate gene studies for interactions have been reported. We conducted a genome-wide screen with meta-analysis approach to identify loci having interactions with epidemiological risk factors on serum lipids with over 30,000 population-based samples. When combining results from our initial datasets and 8 additional replication cohorts (maximum N = 17,102), we found a genome-wide significant locus in chromosome 4p15 with a joint P-value of 4.79×10−9 modifying the effect of waist-to-hip ratio on total cholesterol. In the area surrounding this genetic variant, there were two genes having association between the genotypes and the gene expression in adipose tissue, and we also found enrichment of association in genes belonging to lipid metabolism related functions.
Improved glycemic control reduces complications in patients with diabetes mellitus (DM). However, it is discussed controversially whether patients with diabetes mellitus and end-stage renal disease benefit from strict glycemic control.
We followed 78 patients with DM initiating dialysis treatment of the region of Vorarlberg in a prospective cohort study applying a time-dependent Cox regression analysis using all measured laboratory values for up to more than seven years. This resulted in 880 HbA1c measurements (with one measurement every 3.16 patient months on average) during the entire observation period. Non-linear P-splines were used to allow flexible modeling of the association with mortality and cardiovascular disease (CVD) events.
We observed a decreased mortality risk with increasing HbA1c values (HR = 0.72 per 1% increase, p = 0.024). Adjustment for age and sex and additional adjustment for other CVD risk factors only slightly attenuated the association (HR = 0.71, p = 0.044). A non-linear P-spline showed that the association did not follow a fully linear pattern with a highly significant non-linear component (p = 0.001) with an increased risk of all-cause mortality for HbA1c values up to 6–7%. Causes of death were associated with HbA1c values. The risk for CVD events, however, increased with increasing HbA1c values (HR = 1.24 per 1% increase, p = 0.048) but vanished after extended adjustments.
This study considered the entire information collected on HbA1c over a period of more than seven years. Besides the methodological advantages our data indicate a significant inverse association between HbA1c levels and all-cause mortality. However, for CVD events no significant association could be found.
Waist-hip ratio (WHR) is a measure of body fat distribution and a predictor of metabolic consequences independent of overall adiposity. WHR is heritable, but few genetic variants influencing this trait have been identified. We conducted a meta-analysis of 32 genome-wide association studies for WHR adjusted for body-mass-index (up to 77,167 participants), following up 16 loci in an additional 29 studies (up to 113,636 subjects). We identified 13 novel loci in or near RSPO3, VEGFA, TBX15-WARS2, NFE2L3, GRB14, DNM3-PIGC, ITPR2-SSPN, LY86, HOXC13, ADAMTS9, ZNRF3-KREMEN1, NISCH-STAB1, and CPEB4 (P 1.9 × 10−9 to 1.8 × 10−40), and the known signal at LYPLAL1. Seven of these loci exhibited marked sexual dimorphism, all with a stronger effect on WHR in women than men (P for sex-difference 1.9 × 10−3 to 1.2 × 10−13). These findings provide evidence for multiple loci that modulate body fat distribution, independent of overall adiposity, and reveal powerful gene-by-sex interactions.
genome-wide association; waist-hip-ratio; body fat distribution; central obesity; meta-analysis; genetics; visceral adipose tissue; metabolism; body composition; Expression Quantitative Trait Loci; sex difference
Waist-hip ratio (WHR) is a measure of body fat distribution and a predictor of metabolic consequences independent of overall adiposity. WHR is heritable, but few genetic variants influencing this trait have been identified. We conducted a meta-analysis of 32 genome-wide association studies for WHR adjusted for body mass index (comprising up to 77,167 participants), following up 16 loci in an additional 29 studies (comprising up to 113,636 subjects). We identified 13 new loci in or near RSPO3, VEGFA, TBX15-WARS2, NFE2L3, GRB14, DNM3-PIGC, ITPR2-SSPN, LY86, HOXC13, ADAMTS9, ZNRF3-KREMEN1, NISCH-STAB1 and CPEB4 (P = 1.9 × 10−9 to P = 1.8 × 10−40) and the known signal at LYPLAL1. Seven of these loci exhibited marked sexual dimorphism, all with a stronger effect on WHR in women than men (P for sex difference = 1.9 × 10−3 to P = 1.2 × 10−13). These findings provide evidence for multiple loci that modulate body fat distribution independent of overall adiposity and reveal strong gene-by-sex interactions.