Emmanuel Vaillant

Variation in FTO contributes to childhood obesity and severe adult obesity

We identified a set of SNPs in the first intron of the FTO (fat mass and obesity associated) gene on chromosome 16q12.2 that is consistently strongly associated with early-onset and severe obesity in both adults and children of European ancestry with an experiment-wise P value of 1.67 × 10−26 in 2,900 affected individuals and 5,100 controls. The at-risk haplotype yields a proportion of attributable risk of 22% for common obesity. We conclude that FTO contributes to human obesity and hence may be a target for subsequent functional analyses.

A genome-wide scan for human obesity genes reveals a major susceptibility locus on chromosome 10

Obesity, a common multifactorial disorder, is a major risk factor for type 2 diabetes, hypertension and coronary heart disease (CHD). According to the definition of the World Health Organization (WHO), approximately 6-10% of the population in Westernized countries are considered obese. Epidemiological studies have shown that 30-70% of the variation in body weight may be attributable to genetic factors. To date, two genome-wide scans using different obesity-related quantitative traits have provided candidate regions for obesity. We have undertaken a genome-wide scan in affected sibpairs to identify chromosomal regions linked to obesity in a collection of French families. Model-free multipoint linkage analyses revealed evidence for linkage to a region on chromosome 10p (MLS=4.85). Two further loci on chromosomes 5cen–q and 2p showed suggestive evidence for linkage of serum leptin levels in a genome-wide context. The peak on chromosome 2 coincided with the region containing the gene (POMC) encoding pro-opiomelanocortin, a locus previously linked to leptin levels and fat mass in a Mexican-American population and shown to be mutated in obese humans. Our results suggest that there is a major gene on chromosome 10p implicated in the development of human obesity, and the existence of two further loci influencing leptin levels.

Rare MTNR1B variants impairing melatonin receptor 1B function contribute to type 2 diabetes

Genome-wide association studies have revealed that common noncoding variants in MTNR1B (encoding melatonin receptor 1B, also known as MT2) increase type 2 diabetes (T2D) risk. Although the strongest association signal was highly significant (P < 1 × 10−20), its contribution to T2D risk was modest (odds ratio (OR) of ∼1.10–1.15). We performed large-scale exon resequencing in 7,632 Europeans, including 2,186 individuals with T2D, and identified 40 nonsynonymous variants, including 36 very rare variants (minor allele frequency (MAF) <0.1%), associated with T2D (OR = 3.31, 95% confidence interval (CI) = 1.78–6.18; P = 1.64 × 10−4). A four-tiered functional investigation of all 40 mutants revealed that 14 were non-functional and rare (MAF < 1%), and 4 were very rare with complete loss of melatonin binding and signaling capabilities. Among the very rare variants, the partial- or total-loss-of-function variants but not the neutral ones contributed to T2D (OR = 5.67, CI = 2.17–14.82; P = 4.09 × 10−4). Genotyping the four complete loss-of-function variants in 11,854 additional individuals revealed their association with T2D risk (8,153 individuals with T2D and 10,100 controls; OR = 3.88, CI = 1.49–10.07; P = 5.37 × 10−3). This study establishes a firm functional link between MTNR1B and T2D risk.

Low copy number of the salivary amylase gene predisposes to obesity

Common multi-allelic copy number variants (CNVs) appear enriched for phenotypic associations compared to their biallelic counterparts. Here we investigated the influence of gene dosage effects on adiposity through a CNV association study of gene expression levels in adipose tissue. We identified significant association of a multi-allelic CNV encompassing the salivary amylase gene (AMY1) with body mass index (BMI) and obesity, and we replicated this finding in 6,200 subjects. Increased AMY1 copy number was positively associated with both amylase gene expression (P = 2.31 × 10−14) and serum enzyme levels (P < 2.20 × 10−16), whereas reduced AMY1 copy number was associated with increased BMI (change in BMI per estimated copy = −0.15 (0.02) kg/m2; P = 6.93 × 10−10) and obesity risk (odds ratio (OR) per estimated copy = 1.19, 95% confidence interval (CI) = 1.13–1.26; P = 1.46 × 10−10). The OR value of 1.19 per copy of AMY1 translates into about an eightfold difference in risk of obesity between subjects in the top (copy number > 9) and bottom (copy number < 4) 10% of the copy number distribution. Our study provides a first genetic link between carbohydrate metabolism and BMI and demonstrates the power of integrated genomic approaches beyond genome-wide association studies.

Contribution of type 2 diabetes associated loci in the Arabic population from Tunisia: a case-control study

BackgroundCandidate gene and genome-wide association studies have both reproducibly identified several common Single Nucleotide Polymorphisms (SNPs) that confer type 2 diabetes (T2D) risk in European populations. Our aim was to evaluate the contribution to T2D of five of these established T2D-associated loci in the Arabic population from Tunisia.MethodsA case-control design comprising 884 type 2 diabetic patients and 513 control subjects living in the East-Center of Tunisia was used to analyze the contribution to T2D of the following SNPs: E23K in KCNJ11/Kir6.2, K121Q in ENPP1, the -30G/A variant in the pancreatic β-cell specific promoter of Glucokinase, rs7903146 in TCF7L2 encoding transcription factor 7-like2, and rs7923837 in HHEX encoding the homeobox, hematopoietically expressed transcription factor.ResultsTCF7L2-rs7903146 T allele increased susceptibility to T2D (OR = 1.25 [1.06–1.47], P = 0.006) in our study population. This risk was 56% higher among subjects carrying the TT genotype in comparison to those carrying the CC genotype (OR = 1.56 [1.13–2.16], P = 0.002). No allelic or genotypic association with T2D was detected for the other studied polymorphisms.ConclusionIn the Tunisian population, TCF7L2-rs7903146 T allele confers an increased risk of developing T2D as previously reported in the European population and many other ethnic groups. In contrast, none of the other tested SNPs that influence T2D risk in the European population was associated with T2D in the Tunisian Arabic population. An insufficient power to detect minor allelic contributions or genetic heterogeneity of T2D between different ethnic groups can explain these findings.

Disruption of a Novel Krüppel-like Transcription Factor p300-regulated Pathway for Insulin Biosynthesis Revealed by Studies of the c.-331 INS Mutation Found in Neonatal Diabetes Mellitus

Krüppel-like transcription factors (KLFs) have elicited significant attention because of their regulation of essential biochemical pathways and, more recently, because of their fundamental role in the mechanisms of human diseases. Neonatal diabetes mellitus is a monogenic disorder with primary alterations in insulin secretion. We here describe a key biochemical mechanism that underlies neonatal diabetes mellitus insulin biosynthesis impairment, namely a homozygous mutation within the insulin gene (INS) promoter, c.-331C>G, which affects a novel KLF-binding site. The combination of careful expression profiling, electromobility shift assays, reporter experiments, and chromatin immunoprecipitation demonstrates that, among 16 different KLF proteins tested, KLF11 is the most reliable activator of this site. Congruently, the c.-331C>G INS mutation fails to bind KLF11, thus inhibiting activation by this transcription factor. Klf11−/− mice recapitulate the disruption in insulin production and blood levels observed in patients. Thus, these data demonstrate an important role for KLF11 in the regulation of INS transcription via the novel c.-331 KLF site. Lastly, our screening data raised the possibility that other members of the KLF family may also regulate this promoter under distinct, yet unidentified, cellular contexts. Collectively, this study underscores a key role for KLF proteins in biochemical mechanisms of human diseases, in particular, early infancy onset diabetes mellitus.

Contribution of 24 obesity-associated genetic variants to insulin resistance, pancreatic beta-cell function and type 2 diabetes risk in the French population

Context:Obesity is the major determinant of type 2 diabetes (T2D), presumably through its effect on insulin resistance. Genome-wide association studies reported many single-nucleotide polymorphisms (SNPs) that increase obesity risk and body mass index (BMI), but their impact on T2D-related traits and risk is unclear.Objective:We aimed at analyzing the effect of 24 obesity risk alleles, separately and in combination, on variation of both insulin resistance and β-cell dysfunction, and on T2D risk.Design:We genotyped 24 obesity-associated SNPs and calculated an obesity genotype score (sum of the obesity risk alleles per individual). We analyzed the contribution of each SNP and this score to the variation of four metabolic indices: homeostasis model assessment of insulin resistance (HOMA-IR), homeostasis model assessment of the pancreatic β-cell function (HOMA-B), insulin sensitivity index (ISI) and insulinogenic index (II) (in up to 8050 nondiabetic French individuals) and to T2D risk (in 2077 T2D cases and 3085 controls).Results:We found a highly significant effect of the obesity genotype score on increased insulin resistance adjusted for age and gender (β=0.02; P-value=7.16 × 10−9 for HOMA-IR). Individually, we identified nominal or significant association between increased insulin resistance and risk alleles in FAIM2, FTO, GNPDA2, MC4R, NPC1, PTER and SH2B1. Most signals, including the obesity genotype score and FTO SNP, were also associated with increased β-cell function (β=0.01; P-value=1.05 × 10−6 and β=0.04; P-value=3.45 × 10−4, respectively). In our T2D case–control study, only the obesity genotype score and the well-known FTO locus significantly contributed to T2D risk (OR=1.03; P-value=9.99 × 10−3 and OR=1.15; P-value=9.46 × 10−4, respectively). Adjustment for BMI abolished all significant associations.Conclusions:Genetic predisposition to obesity contributes to increased insulin resistance and to its compensation through increased β-cell function, and weakly increases the T2D risk. These associations are mediated by BMI.

Differential coexpression analysis of obesity-associated networks in human subcutaneous adipose tissue.

Objective:To use a unique obesity-discordant sib-pair study design to combine differential expression analysis, expression quantitative trait loci (eQTLs) mapping and a coexpression regulatory network approach in subcutaneous human adipose tissue to identify genes relevant to the obese state.Study design:Genome-wide transcript expression in subcutaneous human adipose tissue was measured using Affymetrix U133 Plus 2.0 microarrays (Affymetrix, Santa Clara, CA, USA), and genome-wide genotyping data was obtained using an Applied Biosystems (Applied Biosystems; Life Technologies, Carlsbad, CA, USA) SNPlex linkage panel.Subjects:A total of 154 Swedish families ascertained through an obese proband (body mass index (BMI) >30 kg m−2) with a discordant sibling (BMI>10 kg m−2 less than proband).Results:Approximately one-third of the transcripts were differentially expressed between lean and obese siblings. The cellular adhesion molecules (CAMs) KEGG grouping contained the largest number of differentially expressed genes under cis-acting genetic control. By using a novel approach to contrast CAMs coexpression networks between lean and obese siblings, a subset of differentially regulated genes was identified, with the previously GWAS obesity-associated neuronal growth regulator 1 (NEGR1) as a central hub. Independent analysis using mouse data demonstrated that this finding of NEGR1 is conserved across species.Conclusion:Our data suggest that in addition to its reported role in the brain, NEGR1 is also expressed in subcutaneous adipose tissue and acts as a central ‘hub’ in an obesity-related transcript network.

RFX6 Regulates Insulin Secretion by Modulating Ca2+ Homeostasis in Human β Cells

Development and function of pancreatic β cells involve the regulated activity of specific transcription factors. RFX6 is a transcription factor essential for mouse β cell differentiation that is mutated in monogenic forms of neonatal diabetes. However, the expression and functional roles of RFX6 in human β cells, especially in pathophysiological conditions, are poorly explored. We demonstrate the presence of RFX6 in adult human pancreatic endocrine cells. Using the recently developed human β cell line EndoC-βH2, we show that RFX6 regulates insulin gene transcription, insulin content, and secretion. Knockdown of RFX6 causes downregulation of Ca(2+)-channel genes resulting in the reduction in L-type Ca(2+)-channel activity that leads to suppression of depolarization-evoked insulin exocytosis. We also describe a previously unreported homozygous missense RFX6 mutation (p.V506G) that is associated with neonatal diabetes, which lacks the capacity to activate the insulin promoter and to increase Ca(2+)-channel expression. Our data therefore provide insights for understanding certain forms of neonatal diabetes.

Contribution of the low-frequency, loss-of-function p.R270H mutation in FFAR4 ( GPR120 ) to increased fasting plasma glucose levels

Background We previously reported that the low-frequency, loss-of-function variant p.R270H in FFAR4 encoding the lipid sensor GPR120 was associated with obesity. Gpr120-deficient mice develop obesity and both impaired fasting glucose and glucose intolerance under a high-fat diet. We aimed to assess the contribution of p.R270H to type 2 diabetes (T2D) risk and the variation of glucose-related traits. Methods We genotyped p.R270H in 8996 non-diabetic individuals (among whom 4523 had an oral glucose tolerance test (OGTT)) and in a T2D case–control study including 4725 cases and 4339 controls. The regression models were adjusted for age, sex and body mass index (BMI). Results We found a significant association between p.R270H and increased fasting glucose levels (β=0.092±0.05 mmol/L; p=4.13×10−4). Furthermore, p.R270H nominally contributed to decreased homeostasis model of pancreatic β-cell function (HOMA-B; β=−0.090±0.06; p=6.01×10−3). Despite a high statistical power, we did not find any significant association between p.R270H and T2D risk or the variation of fasting insulin levels, the homeostasis model of insulin resistance or OGTT-derived indices. Conclusions These results suggest that the low-frequency p.R270H variant which inhibits GPR120 activity might influence fasting glucose levels in a normal physiological range. This study does not exclude that other coding mutations in FFAR4 with stronger functional effect than p.R270H may be associated with T2D.