Individual variation in reward sensitivity may have an important role in early substance use and subsequent development of substance abuse. This may be especially important during adolescence, a transition period marked by approach behavior and a propensity toward risk taking, novelty seeking and alteration of the social landscape. However, little is known about the relative contribution of personality, behavior, and brain responses for prediction of alcohol use in adolescents. In this study, we applied factor analyses and structural equation modeling to reward-related brain responses assessed by functional magnetic resonance imaging during a monetary incentive delay task. In addition, novelty seeking, sensation seeking, impulsivity, extraversion, and behavioral measures of risk taking were entered as predictors of early onset of drinking in a sample of 14-year-old healthy adolescents (N=324). Reward-associated behavior, personality, and brain responses all contributed to alcohol intake with personality explaining a higher proportion of the variance than behavior and brain responses. When only the ventral striatum was used, a small non-significant contribution to the prediction of early alcohol use was found. These data suggest that the role of reward-related brain activation may be more important in addiction than initiation of early drinking, where personality traits and reward-related behaviors were more significant. With up to 26% of explained variance, the interrelation of reward-related personality traits, behavior, and neural response patterns may convey risk for later alcohol abuse in adolescence, and thus may be identified as a vulnerability factor for the development of substance use disorders.

Background Variation in the complement factor H gene (CFH) is associated with risk of late age-related macular degeneration (AMD). Previous studies have been case–control studies in populations of European ancestry with little differentiation in AMD subtype, and insufficient power to confirm or refute effect modification by smoking.

Methods To precisely quantify the association of the single nucleotide polymorphism (SNP rs1061170, ‘Y402H’) with risk of AMD among studies with differing study designs, participant ancestry and AMD grade and to investigate effect modification by smoking, we report two unpublished genetic association studies (n = 2759) combined with data from 24 published studies (26 studies, 26 494 individuals, including 14 174 cases of AMD) of European ancestry, 10 of which provided individual-level data used to test gene–smoking interaction; and 16 published studies from non-European ancestry.

Results In individuals of European ancestry, there was a significant association between Y402H and late-AMD with a per-allele odds ratio (OR) of 2.27 [95% confidence interval (CI) 2.10–2.45; P = 1.1 x 10−161]. There was no evidence of effect modification by smoking (P = 0.75). The frequency of Y402H varied by ancestral origin and the association with AMD in non-Europeans was less clear, limited by paucity of studies.

Conclusion The Y402H variant confers a 2-fold higher risk of late-AMD per copy in individuals of European descent. This was stable to stratification by study design and AMD classification and not modified by smoking. The lack of association in non-Europeans requires further verification. These findings are of direct relevance for disease prediction. New research is needed to ascertain if differences in circulating levels, expression or activity of factor H protein explain the genetic association.

Primary ciliary dyskinesia (PCD) is an inherited disorder characterized by recurrent infections of the upper and lower respiratory tract, reduced fertility in males and situs inversus in about 50% of affected individuals (Kartagener syndrome). It is caused by motility defects in the respiratory cilia that are responsible for airway clearance, the flagella that propel sperm cells and the nodal monocilia that determine left-right asymmetry1. Recessive mutations that cause PCD have been identified in genes encoding components of the outer dynein arms, radial spokes and cytoplasmic pre-assembly factors of axonemal dyneins, but these mutations account for only about 50% of cases of PCD. We exploited the unique properties of dog populations to positionally clone a new PCD gene, CCDC39. We found that loss-of-function mutations in the human ortholog underlie a substantial fraction of PCD cases with axonemal disorganization and abnormal ciliary beating. Functional analyses indicated that CCDC39 localizes to ciliary axonemes and is essential for assembly of inner dynein arms and the dynein regulatory complex.

Standard cancer cell lines do not model the intratumoural heterogeneity situation sufficiently. Clonal selection leads to a homogeneous population of cells by genetic drift. Heterogeneity of tumour cells, however, is particularly critical for therapeutically relevant studies, since it is a prerequisite for acquiring drug resistance and reoccurrence of tumours. Here, we report the isolation of a highly tumourigenic primary pancreatic cancer cell line, called JoPaca-1 and its detailed characterization at multiple levels. Implantation of as few as 100 JoPaca-1 cells into immunodeficient mice gave rise to tumours that were histologically very similar to the primary tumour. The high heterogeneity of JoPaca-1 was reflected by diverse cell morphology and a substantial number of chromosomal aberrations. Comparative whole-genome sequencing of JoPaca-1 and BxPC-3 revealed mutations in genes frequently altered in pancreatic cancer. Exceptionally high expression of cancer stem cell markers and a high clonogenic potential in vitro and in vivo was observed. All of these attributes make this cell line an extremely valuable model to study the biology of and pharmaceutical effects on pancreatic cancer.

Altered mitochondrial DNA (mtDNA) levels have been associated with common diseases in humans. We investigated the genetic mechanism that controls mtDNA levels using genome-wide linkage analyses in families from the Genetic Analysis of Idiopathic Thrombophilia Project (GAIT). We measure mtDNA levels by quantitative real-time PCR in 386 subjects from 21 extended Spanish families. A variance component linkage method using 485 microsatellites was conducted to evaluate linkage and to detect quantitative trait loci (QTLs) involved in the control of mtDNA levels. The heritalibility of mtDNA levels was 0.33 (p = 1.82e-05). We identified a QTL on Chromosome 2 (LOD = 2.21) using all of the subjects, independently on their sex. When females and males were analysed separately, three QTLs were identified. Females showed the same QTL on Chromosome 2 (LOD = 3.09), indicating that the QTL identified in the analysis using all of the subjects was a strong female QTL, and another one on Chromosome 3 (LOD = 2.67), whereas in males a QTL was identified on Chromosome 1 (LOD = 2.81). These QTLs were fine-mapped to find associations with mtDNA levels. The most significant SNP association was for the rs10888838 on Chromosome 1 in males. This SNP mapped to the gene MRPL37, involved in mitochondrial protein translation. The rs2140855 on Chromosome 2 showed association in the analysis using all of the subjects. It was near the gene CMPK2, which encodes a mitochondrial enzyme of the salvage pathway of deoxyribonucleotide synthesis. Our results provide evidence of a sex-specific genetic mechanism for the control of mtDNA levels and provide a framework to identify new genes that influence mtDNA levels.

While gliomas are the most common primary brain tumors, their etiology is largely unknown. To identify novel risk loci for glioma, we conducted genome-wide association (GWA) analysis of two case–control series from France and Germany (2269 cases and 2500 controls). Pooling these data with previously reported UK and US GWA studies provided data on 4147 glioma cases and 7435 controls genotyped for 424 460 common tagging single-nucleotide polymorphisms. Using these data, we demonstrate two statistically independent associations between glioma and rs11979158 and rs2252586, at 7p11.2 which encompasses the EGFR gene (population-corrected statistics, Pc = 7.72 × 10−8 and 2.09 × 10−8, respectively). Both associations were independent of tumor subtype, and were independent of EGFR amplification, p16INK4a deletion and IDH1 mutation status in tumors; compatible with driver effects of the variants on glioma development. These findings show that variation in 7p11.2 is a determinant of inherited glioma risk.

The manifestation of coronary artery disease (CAD) follows a well-choreographed series of events that includes damage of arterial endothelial cells and deposition of lipids in the sub-endothelial layers. Genome-wide association studies (GWAS) of multiple populations with distinctive genetic and lifestyle backgrounds are a crucial step in understanding global CAD pathophysiology. In this study, we report a GWAS on the genetic basis of arterial stenosis as measured by cardiac catheterization in a Lebanese population. The locus of the phosphatase and actin regulator 1 gene (PHACTR1) showed association with coronary stenosis in a discovery experiment with genome wide data in 1,949 individuals (rs9349379, OR = 1.37, p = 1.57×10−5). The association was replicated in an additional 2,547 individuals (OR = 1.31, p = 8.85×10−6), leading to genome-wide significant association in a combined analysis (OR = 1.34, p = 8.02×10−10). Results from this GWAS support a central role of PHACTR1 in CAD susceptibility irrespective of lifestyle and ethnic divergences. This association provides a plausible component for understanding molecular mechanisms involved in the formation of stenosis in cardiac vessels and a potential drug target against CAD.

By applying an imputation strategy based on the 1000 Genomes project to two genome-wide association studies (GWAS), we detected a susceptibility locus for venous thrombosis on chromosome 11p11.2 that was missed by previous GWAS analyses that had been conducted on the same datasets. A comprehensive linkage disequilibrium and haplotype analysis of the whole locus where twelve SNPs exhibited association p-values lower than 2.23 10−11 and the use of independent case-control samples demonstrated that the culprit variant was a rare variant located ∼1 Mb away from the original hits, not tagged by current genome-wide genotyping arrays and even not well imputed in the original GWAS samples. This variant was in fact the rs1799963, also known as the FII G20210A prothrombin mutation. This work may be of major interest not only for its scientific impact but also for its methodological findings.

Our specific aims were to evaluate the power of bivariate analysis and to compare its performance with traditional univariate analysis in samples of unrelated subjects under varying sampling selection designs. Bivariate association analysis was based on the seemingly unrelated regression (SUR) model that allows different genetic models for different traits. We conducted extensive simulations for the case of two correlated quantitative phenotypes, with the quantitative trait locus making equal or unequal contributions to each phenotype. Our simulation results confirmed that the power of bivariate analysis is affected by the size, direction and source of the phenotypic correlations between traits. They also showed that the optimal sampling scheme depends on the size and direction of the induced genetic correlation. In addition, we demonstrated the efficacy of SUR-based bivariate test by applying it to a real Genome-Wide Association Study (GWAS) of Bone Mineral Density (BMD) values measured at the lumbar spine (LS) and at the femoral neck (FN) in a sample of unrelated males with low BMD (LS Z-scores ≤−2) and with high BMD (LS and FN Z-scores >0.5). A substantial amount of top hits in bivariate analysis did not reach nominal significance in any of the two single-trait analyses. Altogether, our studies suggest that bivariate analysis is of practical significance for GWAS of correlated phenotypes.

Background

The nitric oxide (NO) pathway is involved in asthma, and eosinophils participate in the regulation of the NO pool in pulmonary tissues. We investigated associations between single nucleotide polymorphisms (SNPs) of NO synthase genes (NOS) and biological NO-related phenotypes measured in two compartments (exhaled breath condensate and plasma) and blood eosinophil counts.

Methodology

SNPs (N = 121) belonging to NOS1, NOS2 and NOS3 genes were genotyped in 1277 adults from the French Epidemiological study on the Genetics and Environment of Asthma (EGEA). Association analyses were conducted on four quantitative phenotypes: the exhaled fraction of NO (FeNO), plasma and exhaled breath condensate (EBC) nitrite-nitrate levels (NO2–NO3) and blood eosinophils in asthmatics and non-asthmatics separately. Genetic heterogeneity of these phenotypes between asthmatics and non-asthmatics was also investigated.

Principal Findings

In non-asthmatics, after correction for multiple comparisons, we found significant associations of FeNO levels with three SNPs in NOS3 and NOS2 (P≤0.002), and of EBC NO2–NO3 level with NOS2 (P = 0.002). In asthmatics, a single significant association was detected between FeNO levels and one SNP in NOS3 (P = 0.004). Moreover, there was significant heterogeneity of NOS3 SNP effect on FeNO between asthmatics and non-asthmatics (P = 0.0002 to 0.005). No significant association was found between any SNP and NO2–NO3 plasma levels or blood eosinophil counts.

Conclusions

Variants in NO synthase genes influence FeNO and EBC NO2–NO3 levels in adults. These genetic determinants differ according to asthma status. Significant associations were only detected for exhaled phenotypes, highlighting the critical relevance to have access to specific phenotypes measured in relevant biological fluid.

The insulin (INS) region is the second most important locus associated with Type 1 Diabetes (T1D). The study of the DNA methylation pattern of the 7 CpGs proximal to the TSS in the INS gene promoter revealed that T1D patients have a lower level of methylation of CpG -19, -135 and -234 (p = 2.10−16) and a higher methylation of CpG -180 than controls, while methylation was comparable for CpG -69, -102, -206. The magnitude of the hypomethylation relative to a control population was 8–15% of the corresponding levels in controls and was correlated in CpGs -19 and -135 (r = 0.77) and CpG -135 and -234 (r = 0.65). 70/485 (14%) of T1D patients had a simultaneous decrease in methylation of CpG -19, -135, -234 versus none in 317 controls. CpG methylation did not correlate with glycated hemoglobin or with T1D duration. The methylation of CpG -69, -102, -180, -206, but not CpG -19, -135, -234 was strongly influenced by the cis-genotype at rs689, a SNP known to show a strong association with T1D. We hypothesize that part of this genetic association could in fact be mediated at the statistical and functional level by the underlying changes in neighboring CpG methylation. Our observation of a CpG-specific, locus-specific methylation pattern, although it can provide an epigenetic biomarker of a multifactorial disease, does not indicate whether the reported epigenetic pattern preexists or follows the establishment of T1D. To explore the effect of chronic hyperglycemia on CpG methylation, we studied non obese patients with type 2 diabetes (T2D) who were found to have decreased CpG-19 methylation versus age-matched controls, similar to T1D (p = 2.10−6) but increased CpG-234 methylation (p = 5.10−8), the opposite of T1D. The causality and natural history of the different epigenetic changes associated with T1D or T2D remain to be determined.

Alcohol abuse and dependence have proven to be complex genetic traits that are influenced by environmental factors. Primate and human studies have shown that early life stress increases the propensity for alcohol abuse in later life. The reinforcing properties of alcohol are mediated by dopaminergic signaling; however, there is little evidence to indicate how stress alters alcohol reinforcement. KCNJ6 (the gene encoding G-protein-coupled inwardly rectifying potassium channel 2 (GIRK2)) is a brain expressed potassium channel with inhibitory effects on dopaminergic tone. The properties of GIRK2 have been shown to be enhanced by the stress peptide corticotrophin-releasing hormone. Therefore, we sought to examine the role of KCNJ6 polymorphisms in adult alcohol dependence and stress-related alcohol abuse in adolescents. We selected 11 SNPs in the promoter region of KCNJ6, which were genotyped in 1152 adult alcohol dependents and 1203 controls. One SNP, rs2836016, was found to be associated with alcohol dependence (p=0.01, false discovery rate). We then assessed rs2836016 in an adolescent sample of 261 subjects, which were characterized for early life stress and adolescent hazardous drinking, defined using the Alcohol Use Disorders Identification Test (AUDIT), to examine gene–environment interactions. In the adolescent sample, the risk genotype of rs2836016 was significantly associated with increased AUDIT scores, but only in those individuals exposed to high levels of psychosocial stress in early life (p=0.01). Our findings show that KCNJ6 is associated with alcohol dependence and may moderate the effect of early psychosocial stress on risky alcohol drinking in adolescents. We have identified a candidate gene for future studies investigating a possible functional link between the response to stress and alcohol reinforcement.

Background

Transient receptor potential (TRP) vanilloid and ankyrin cation channels are activated by various noxious chemicals and may play an important role in the pathogenesis of cough. The aim was to study the influence of single nucleotide polymorphisms (SNPs) in TRP genes and irritant exposures on cough.

Methods

Nocturnal, usual, and chronic cough, smoking, and job history were obtained by questionnaire in 844 asthmatic and 2046 non-asthmatic adults from the Epidemiological study on the Genetics and Environment of Asthma (EGEA) and the European Community Respiratory Health Survey (ECRHS). Occupational exposures to vapors, gases, dusts, and/or fumes were assessed by a job-exposure matrix. Fifty-eight tagging SNPs in TRPV1, TRPV4, and TRPA1 were tested under an additive model.

Results

Statistically significant associations of 6 TRPV1 SNPs with cough symptoms were found in non-asthmatics after correction for multiple comparisons. Results were consistent across the eight countries examined. Haplotype-based association analysis confirmed the single SNP analyses for nocturnal cough (7-SNP haplotype: p-global = 4.8 × 10-6) and usual cough (9-SNP haplotype: p-global = 4.5 × 10-6). Cough symptoms were associated with exposure to irritants such as cigarette smoke and occupational exposures (p < 0.05). Four polymorphisms in TRPV1 further increased the risk of cough symptoms from irritant exposures in asthmatics and non-asthmatics (interaction p < 0.05).

Conclusions

TRPV1 SNPs were associated with cough among subjects without asthma from two independent studies in eight European countries. TRPV1 SNPs may enhance susceptibility to cough in current smokers and in subjects with a history of workplace exposures.

Robert Clarke and colleagues conduct a meta-analysis of unpublished datasets to examine the causal relationship between elevation of homocysteine levels in the blood and the risk of coronary heart disease. Their data suggest that an increase in homocysteine levels is not likely to result in an increase in risk of coronary heart disease.

Background

Moderately elevated blood levels of homocysteine are weakly correlated with coronary heart disease (CHD) risk, but causality remains uncertain. When folate levels are low, the TT genotype of the common C677T polymorphism (rs1801133) of the methylene tetrahydrofolate reductase gene (MTHFR) appreciably increases homocysteine levels, so “Mendelian randomization” studies using this variant as an instrumental variable could help test causality.

Methods and Findings

Nineteen unpublished datasets were obtained (total 48,175 CHD cases and 67,961 controls) in which multiple genetic variants had been measured, including MTHFR C677T. These datasets did not include measurements of blood homocysteine, but homocysteine levels would be expected to be about 20% higher with TT than with CC genotype in the populations studied. In meta-analyses of these unpublished datasets, the case-control CHD odds ratio (OR) and 95% CI comparing TT versus CC homozygotes was 1.02 (0.98–1.07; p = 0.28) overall, and 1.01 (0.95–1.07) in unsupplemented low-folate populations. By contrast, in a slightly updated meta-analysis of the 86 published studies (28,617 CHD cases and 41,857 controls), the OR was 1.15 (1.09–1.21), significantly discrepant (p = 0.001) with the OR in the unpublished datasets. Within the meta-analysis of published studies, the OR was 1.12 (1.04–1.21) in the 14 larger studies (those with variance of log OR<0.05; total 13,119 cases) and 1.18 (1.09–1.28) in the 72 smaller ones (total 15,498 cases).

Conclusions

The CI for the overall result from large unpublished datasets shows lifelong moderate homocysteine elevation has little or no effect on CHD. The discrepant overall result from previously published studies reflects publication bias or methodological problems.

Please see later in the article for the Editors' Summary

Editors' Summary

Background

Coronary heart disease (CHD) is the leading cause of death among adults in developed countries. With age, fatty deposits (atherosclerotic plaques) coat the walls of the coronary arteries, the blood vessels that supply the heart with oxygen and nutrients. The resultant restriction of the heart's blood supply causes shortness of breath, angina (chest pains that are usually relieved by rest), and sometimes fatal heart attacks. Many established risk factors for CHD, including smoking, physical inactivity, being overweight, and eating a fat-rich diet, can be modified by lifestyle changes. Another possible modifiable risk factor for CHD is a high blood level of the amino acid homocysteine. Methylene tetrahydofolate reductase, which is encoded by the MTHFR gene, uses folate to break down and remove homocysteine so fortification of cereals with folate can reduce population homocysteine blood levels. Pooled results from prospective observational studies that have looked for an association between homocysteine levels and later development of CHD suggest that the reduction in homocysteine levels that can be achieved by folate supplementation is associated with an 11% lower CHD risk.

Why Was This Study Done?

Prospective observational studies cannot prove that high homocysteine levels cause CHD because of confounding, the potential presence of other unknown shared characteristics that really cause CHD. However, an approach called “Mendelian randomization” can test whether high blood homocysteine causes CHD. A common genetic variant of the MTHFR gene—the C677T polymorphism—reduces MTHFR efficiency so TT homozygotes (individuals in whom both copies of the MTHFR gene have the nucleotide thymine at position 677; the human genome contains two copies of most genes) have 25% higher blood homocysteine levels than CC homozygotes. In meta-analyses (statistical pooling of the results of several studies) of published Mendelian randomized studies, TT homozygotes have a higher CHD risk than CC homozygotes. Because gene variants are inherited randomly, they are not subject to confounding, so this result suggests that high blood homocysteine causes CHD. But what if only Mendelian randomization studies that found an association have been published? Such publication bias would affect this aggregate result. Here, the researchers investigate the association of the MTHFR C677T polymorphism with CHD in unpublished datasets that have analyzed this polymorphism incidentally during other genetic studies.

What Did the Researchers Do and Find?

The researchers obtained 19 unpublished datasets that contained data on the MTHFR C677T polymorphism in thousands of people with and without CHD. Meta-analysis of these datasets indicates that the excess CHD risk in TT homozygotes compared to CC homozygotes was 2% (much lower than predicted from the prospective observational studies), a nonsignificant difference (that is, it could have occurred by chance). When the probable folate status of the study populations (based on when national folic acid fortification legislation came into effect) was taken into account, there was still no evidence that TT homozygotes had an excess CHD risk. By contrast, in an updated meta-analysis of 86 published studies of the association of the polymorphism with CHD, the excess CHD risk in TT homozygotes compared to CC homozygotes was 15%. Finally, in a meta-analysis of randomized trials on the use of vitamin B supplements for homocysteine reduction, folate supplementation had no significant effect on the 5-year incidence of CHD.

What Do These Findings Mean?

These analyses of unpublished datasets are consistent with lifelong moderate elevation of homocysteine levels having no significant effect on CHD risk. In other words, these findings indicate that circulating homocysteine levels within the normal range are not causally related to CHD risk. The meta-analysis of the randomized trials of folate supplementation also supports this conclusion. So why is there a discrepancy between these findings and those of meta-analyses of published Mendelian randomization studies? The discrepancy is too large to be dismissed as a chance finding, suggest the researchers, but could be the result of publication bias—some studies might have been prioritized for publication because of the positive nature of their results whereas the unpublished datasets used in this study would not have been affected by any failure to publish null results. Overall, these findings reveal a serious example of publication bias and argue against the use of folate supplements as a means of reducing CHD risk.

Additional Information

Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001177.

The American Heart Association provides information about CHD and tips on keeping the heart healthy; it also provides information on homocysteine, folic acid, and CHD, general information on supplements and heart health, and personal stories about CHD

The UK National Health Service Choices website provides information about CHD, including personal stories about CHD

Information is available from the British Heart Foundation on heart disease and keeping the heart healthy

The US National Heart Lung and Blood Institute also provides information on CHD (in English and Spanish)

MedlinePlus provides links to many other sources of information on CHD (in English and Spanish)

Wikipedia has a page on Mendelian randomization (note: Wikipedia is a free online encyclopedia that anyone can edit; available in several languages)

Cerebral malaria (CM) is a lethal neurological complication of malaria. We implemented a genome-wide screen in mutagenized mice to identify host proteins involved in CM pathogenesis and whose inhibition may be of therapeutic value. One pedigree (P48) segregated a resistance trait whose CM-protective effect was fully penetrant, mapped to chromosome 8, and identified by genome sequencing as homozygosity for a mis-sense mutation (W81R) in the FERM domain of Janus-associated kinase 3 (Jak3). The causative effect of Jak3W81R was verified by complementation testing in Jak3W81R/− double heterozygotes that were fully protected against CM. Jak3W81R homozygotes showed defects in thymic development with depletion of CD8+ T cell, B cell, and NK cell compartments, and defective T cell-dependent production of IFN-γ. Adoptive transfer of normal splenocytes abrogates CM resistance in Jak3W81R homozygotes, an effect attributed to the CD8+ T cells. Jak3W81R behaves as a dominant negative variant, with significant CM resistance of Jak3W81R/+ heterozygotes, compared to CM-susceptible Jak3+/+ and Jak3+/− controls. CM resistance in Jak3W81R/+ heterozygotes occurs in presence of normal T, B and NK cell numbers. These findings highlight the pathological role of CD8+ T cells and Jak3-dependent IFN-γ-mediated Th1 responses in CM pathogenesis.

The Protein C anticoagulant pathway regulates blood coagulation by preventing the inadequate formation of thrombi. It has two main plasma components: protein C and protein S. Individuals with protein C or protein S deficiency present a dramatically increased incidence of thromboembolic disorders. Here, we present the results of a genome-wide association study (GWAS) for protein C and protein S plasma levels in a set of extended pedigrees from the Genetic Analysis of Idiopathic Thrombophilia (GAIT) Project. A total number of 397 individuals from 21 families were typed for 307,984 SNPs using the Infinium® 317 k Beadchip (Illumina). Protein C and protein S (free, functional and total) plasma levels were determined with biochemical assays for all participants. Association with phenotypes was investigated through variance component analysis. After correcting for multiple testing, two SNPs for protein C plasma levels (rs867186 and rs8119351) and another two for free protein S plasma levels (rs1413885 and rs1570868) remained significant on a genome-wide level, located in and around the PROCR and the DNAJC6 genomic regions respectively. No SNPs were significantly associated with functional or total protein S plasma levels, although rs1413885 from DNAJC6 showed suggestive association with the functional protein S phenotype, possibly indicating that this locus plays an important role in protein S metabolism. Our results provide evidence that PROCR and DNAJC6 might play a role in protein C and free protein S plasma levels in the population studied, warranting further investigation on the role of these loci in the etiology of venous thromboembolism and other thrombotic diseases.

Recurrent rearrangements of chromosome 1q21.1 that occur via non-allelic homologous recombination have been associated with variable phenotypes exhibiting incomplete penetrance, including congenital heart disease (CHD). However, the gene or genes within the ∼1 Mb critical region responsible for each of the associated phenotypes remains unknown. We examined the 1q21.1 locus in 948 patients with tetralogy of Fallot (TOF), 1488 patients with other forms of CHD and 6760 ethnically matched controls using single nucleotide polymorphism genotyping arrays (Illumina 660W and Affymetrix 6.0) and multiplex ligation-dependent probe amplification. We found that duplication of 1q21.1 was more common in cases of TOF than in controls [odds ratio (OR) 30.9, 95% confidence interval (CI) 8.9–107.6); P = 2.2 × 10−7], but deletion was not. In contrast, deletion of 1q21.1 was more common in cases of non-TOF CHD than in controls [OR 5.5 (95% CI 1.4–22.0); P = 0.04] while duplication was not. We also detected rare (n = 3) 100–200 kb duplications within the critical region of 1q21.1 in cases of TOF. These small duplications encompassed a single gene in common, GJA5, and were enriched in cases of TOF in comparison to controls [OR = 10.7 (95% CI 1.8–64.3), P = 0.01]. These findings show that duplication and deletion at chromosome 1q21.1 exhibit a degree of phenotypic specificity in CHD, and implicate GJA5 as the gene responsible for the CHD phenotypes observed with copy number imbalances at this locus.

Background

Venous Thrombosis (VT) is a common multifactorial disease associated with a major public health burden. Genetics factors are known to contribute to the susceptibility of the disease but how many genes are involved and their contribution to VT risk still remain obscure. We aimed to identify genetic variants associated with VT risk.

Methodology/Principal Findings

We conducted a genome-wide association study (GWAS) based on 551,141 SNPs genotyped in 1,542 cases and 1,110 controls. Twelve SNPs reached the genome-wide significance level of 2.0×10−8 and encompassed four known VT-associated loci, ABO, F5, F11 and FGG. By means of haplotype analyses, we also provided novel arguments in favor of a role of HIVEP1, PROCR and STAB2, three loci recently hypothesized to participate in the susceptibility to VT. However, no novel VT-associated loci came out of our GWAS. Using a recently proposed statistical methodology, we also showed that common variants could explain about 35% of the genetic variance underlying VT susceptibility among which 3% could be attributable to the main identified VT loci. This analysis additionally suggested that the common variants left to be identified are not uniformly distributed across the genome and that chromosome 20, itself, could contribute to ∼7% of the total genetic variance.

Conclusions/Significance

This study might also provide a valuable source of information to expand our understanding of biological mechanisms regulating quantitative biomarkers for VT.

Background

Thirty-two common variants associated with body mass index (BMI) have been identified in genome-wide association studies, explaining ∼1.45% of BMI variation in general population cohorts. We performed a genome-wide association study in a sample of young adults enriched for extremely overweight individuals. We aimed to identify new loci associated with BMI and to ascertain whether using an extreme sampling design would identify the variants known to be associated with BMI in general populations.

Methodology/Principal Findings

From two large Danish cohorts we selected all extremely overweight young men and women (n = 2,633), and equal numbers of population-based controls (n = 2,740, drawn randomly from the same populations as the extremes, representing ∼212,000 individuals). We followed up novel (at the time of the study) association signals (p<0.001) from the discovery cohort in a genome-wide study of 5,846 Europeans, before attempting to replicate the most strongly associated 28 SNPs in an independent sample of Danish individuals (n = 20,917) and a population-based cohort of 15-year-old British adolescents (n = 2,418). Our discovery analysis identified SNPs at three loci known to be associated with BMI with genome-wide confidence (P<5×10−8; FTO, MC4R and FAIM2). We also found strong evidence of association at the known TMEM18, GNPDA2, SEC16B, TFAP2B, SH2B1 and KCTD15 loci (p<0.001), and nominal association (p<0.05) at a further 8 loci known to be associated with BMI. However, meta-analyses of our discovery and replication cohorts identified no novel associations.

Significance

Our results indicate that the detectable genetic variation associated with extreme overweight is very similar to that previously found for general BMI. This suggests that population-based study designs with enriched sampling of individuals with the extreme phenotype may be an efficient method for identifying common variants that influence quantitative traits and a valid alternative to genotyping all individuals in large population-based studies, which may require tens of thousands of subjects to achieve similar power.

RNA polymerase (Pol) III synthesizes the tRNAs, the 5S ribosomal RNA and a small number of untranslated RNAs. In vitro, it also transcribes short interspersed nuclear elements (SINEs). We investigated the distribution of Pol III and its associated transcription factors on the genome of mouse embryonic stem cells using a highly specific tandem ChIP-Seq method. Only a subset of the annotated class III genes was bound and thus transcribed. A few hundred SINEs were associated with the Pol III transcription machinery. We observed that Pol III and its transcription factors were present at 30 unannotated sites on the mouse genome, only one of which was conserved in human. An RNA was associated with >80% of these regions. More than 2200 regions bound by TFIIIC transcription factor were devoid of Pol III. These sites were associated with cohesins and often located close to CTCF-binding sites, suggesting that TFIIIC might cooperate with these factors to organize the chromatin. We also investigated the genome-wide distribution of the ubiquitous TFIIS variant, TCEA1. We found that, as in Saccharomyces cerevisiae, TFIIS is associated with class III genes and also with SINEs suggesting that TFIIS is a Pol III transcription factor in mammals.

The sequence and timing of permanent tooth eruption is thought to be highly heritable and can have important implications for the risk of malocclusion, crowding, and periodontal disease. We conducted a genome-wide association study of number of permanent teeth erupted between age 6 and 14 years, analyzed as age-adjusted standard deviation score averaged over multiple time points, based on childhood records for 5,104 women from the Danish National Birth Cohort. Four loci showed association at P<5×10−8 and were replicated in four independent study groups from the United States and Denmark with a total of 3,762 individuals; all combined P-values were below 10−11. Two loci agreed with previous findings in primary tooth eruption and were also known to influence height and breast cancer, respectively. The two other loci pointed to genomic regions without any previous significant genome-wide association study results. The intronic SNP rs7924176 in ADK could be linked to gene expression in monocytes. The combined effect of the four genetic variants was most pronounced between age 10 and 12 years, where children with 6 to 8 delayed tooth eruption alleles had on average 3.5 (95% confidence interval: 2.9–4.1) fewer permanent teeth than children with 0 or 1 of these alleles.

Author Summary

While genome-wide association studies (GWAS) initially focused on the disease under investigation, additional findings in secondary traits have shown further benefits of having extensive phenotype data at hand. Using records from the nationwide dental registry for children and genotype data from two GWAS, we were able to identify four genomic loci associated with permanent tooth eruption in children. Two of the identified genomic regions had no previous GWAS findings, whereas two loci were reported in the context of primary dentition. A follow-up in an on-going GWAS showed that rs7924176 also plays a substantial role in primary dentition. During the age period of permanent tooth eruption many important developmental processes take place. Thus, we suggest following up the four reported SNPs in other growth-related traits to further elucidate the genetic background of maturation.

Background

Elevated levels of factor VIII (FVIII) and von Willebrand Factor (vWF) are well-established risk factors for cardiovascular diseases, in particular venous thrombosis. Although high, the heritability of these traits is poorly explained by the genetic factors known so far. The aim of this work was to identify novel single nucleotide polymorphisms (SNPs) that could influence the variability of these traits.

Methods

Three independent genome-wide association studies for vWF plasma levels and FVIII activity were conducted and their results were combined into a meta-analysis totalling 1,624 subjects.

Results

No single nucleotide polymorphism (SNP) reached the study-wide significance level of 1.12 × 10-7 that corresponds to the Bonferroni correction for the number of tested SNPs. Nevertheless, the recently discovered association of STXBP5, STX2, TC2N and CLEC4M genes with vWF levels and that of SCARA5 and STAB2 genes with FVIII levels were confirmed in this meta-analysis. Besides, among the fifteen novel SNPs showing promising association at p < 10-5 with either vWF or FVIII levels in the meta-analysis, one located in ACCN1 gene also showed weak association (P = 0.0056) with venous thrombosis in a sample of 1,946 cases and 1,228 controls.

Conclusions

This study has generated new knowledge on genomic regions deserving further investigations in the search for genetic factors influencing vWF and FVIII plasma levels, some potentially implicated in VT, as well as providing some supporting evidence of previously identified genes.

Aims

To evaluate the associations of emergent genome-wide-association study-derived coronary heart disease (CHD)-associated single nucleotide polymorphisms (SNPs) with established and emerging risk factors, and the association of genome-wide-association study-derived lipid-associated SNPs with other risk factors and CHD events.

Methods and results

Using two case–control studies, three cross-sectional, and seven prospective studies with up to 25 000 individuals and 5794 CHD events we evaluated associations of 34 genome-wide-association study-identified SNPs with CHD risk and 16 CHD-associated risk factors or biomarkers. The Ch9p21 SNPs rs1333049 (OR 1.17; 95% confidence limits 1.11–1.24) and rs10757274 (OR 1.17; 1.09–1.26), MIA3 rs17465637 (OR 1.10; 1.04–1.15), Ch2q36 rs2943634 (OR 1.08; 1.03–1.14), APC rs383830 (OR 1.10; 1.02, 1.18), MTHFD1L rs6922269 (OR 1.10; 1.03, 1.16), CXCL12 rs501120 (OR 1.12; 1.04, 1.20), and SMAD3 rs17228212 (OR 1.11; 1.05, 1.17) were all associated with CHD risk, but not with the CHD biomarkers and risk factors measured. Among the 20 blood lipid-related SNPs, LPL rs17411031 was associated with a lower risk of CHD (OR 0.91; 0.84–0.97), an increase in Apolipoprotein AI and HDL-cholesterol, and reduced triglycerides. SORT1 rs599839 was associated with CHD risk (OR 1.20; 1.15–1.26) as well as total- and LDL-cholesterol, and apolipoprotein B. ANGPTL3 rs12042319 was associated with CHD risk (OR 1.11; 1.03, 1.19), total- and LDL-cholesterol, triglycerides, and interleukin-6.

Conclusion

Several SNPs predicting CHD events appear to involve pathways not currently indexed by the established or emerging risk factors; others involved changes in blood lipids including triglycerides or HDL-cholesterol as well as LDL-cholesterol. The overlapping association of SNPs with multiple risk factors and biomarkers supports the existence of shared points of regulation for these phenotypes.

Rheumatoid arthritis (RA) is a major cause of adult chronic inflammatory arthritis and a typical complex trait. Although several genetic determinants have been identified, they account for only a part of the genetic susceptibility. We conducted a genome-wide association study of RA in Japanese using 225,079 SNPs genotyped in 990 cases and 1,236 controls from two independent collections (658 cases and 934 controls in collection1; 332 cases and 302 controls in collection2), followed by replication studies in two additional collections (874 cases and 855 controls in collection3; 1,264 cases and 948 controls in collection4). SNPs showing p<0.005 in the first two collections and p<10−4 by meta-analysis were further genotyped in the latter two collections. A novel risk variant, rs2000811, in intron2 of the myelin basic protein (MBP) at chromosome 18q23 showed strong association with RA (p = 2.7×10−8, OR 1.23, 95% CI: 1.14–1.32). The transcription of MBP was significantly elevated with the risk allele compared to the alternative allele (p<0.001). We also established by immunohistochemistry that MBP was expressed in the synovial lining layer of RA patients, the main target of inflammation in the disease. Circulating autoantibody against MBP derived from human brain was quantified by ELISA between patients with RA, other connective tissue diseases and healthy controls. As a result, the titer of anti-MBP antibody was markedly higher in plasma of RA patients compared to healthy controls (p<0.001) and patients with other connective tissue disorders (p<0.001). ELISA experiment using citrullinated recombinant MBP revealed that a large fraction of anti-MBP antibody in RA patients recognized citrullinated MBP. This is the first report of a genetic study in RA implicating MBP as a potential autoantigen and its involvement in pathogenesis of the disease.

Background

Alcohol has been shown to critically modulate cyclic adenosine-3′,5′ monophosphate (cAMP) signaling. A number of downstream effectors that respond to the cAMP signals (e.g., protein kinase A, cAMP response element binding protein) have, in turn, been examined in relation to alcohol consumption. These studies did not, however, delineate the point at which the actions of alcohol on the cAMP cascade might translate into differences in drinking behavior. To further understand the role of cAMP synthesis in alcohol drinking and dependence, we investigated a specific adenylyl cyclase isoform, adenylyl cyclase (AC) Type 7, whose activity is selectively enhanced by ethanol.

Methods

We measured alcohol consumption and preference in mice in which one copy of the Adcy7 gene was disrupted (Adcy7+/−). To demonstrate relevance of this gene for alcohol dependence in humans, we tested the association of polymorphisms in the ADCY7 gene with alcohol dependence in a sample of 1703 alcohol-dependent individuals and 1347 control subjects.

Results

We show that Adcy7+/− female mice have higher preference for alcohol than wild-type mice, whereas there is little difference in alcohol consumption or preference between Adcy7+/− male mice and wild-type control subjects. In the human sample, we found that single nucleotide polymorphisms in ADCY7 associate with alcohol dependence in women, and these markers are also associated with ADCY7 expression (messenger RNA) levels.

Conclusions

These findings implicate adenylyl cyclase Type 7 as a critical component of the molecular pathways contributing to alcohol drinking and the development of alcohol dependence.