Genome-wide analysis of mitochondrial DNA copy number reveals loci implicated in nucleotide metabolism, platelet activation, and megakaryocyte proliferation.

Mitochondrial DNA copy number (mtDNA-CN) measured from blood specimens is a minimally invasive marker of mitochondrial function that exhibits both inter-individual and intercellular variation. To identify genes involved in regulating mitochondrial function, we performed a genome-wide association study (GWAS) in 465,809 White individuals from the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium and the UK Biobank (UKB). We identified 133 SNPs with statistically significant, independent effects associated with mtDNA-CN across 100 loci. A combination of fine-mapping, variant annotation, and co-localization analyses was used to prioritize genes within each of the 133 independent sites. Putative causal genes were enriched for known mitochondrial DNA depletion syndromes (p = 3.09 × 10-15) and the gene ontology (GO) terms for mtDNA metabolism (p = 1.43 × 10-8) and mtDNA replication (p = 1.2 × 10-7). A clustering approach leveraged pleiotropy between mtDNA-CN associated SNPs and 41 mtDNA-CN associated phenotypes to identify functional domains, revealing three distinct groups, including platelet activation, megakaryocyte proliferation, and mtDNA metabolism. Finally, using mitochondrial SNPs, we establish causal relationships between mitochondrial function and a variety of blood cell-related traits, kidney function, liver function and overall (p = 0.044) and non-cancer mortality (p = 6.56 × 10-4).

A Potential Role for the STXBP5-AS1 Gene in Adult ADHD Symptoms.

We aimed to detect Attention-deficit/hyperactivity (ADHD) risk-conferring genes in adults. In children, ADHD is characterized by age-inappropriate levels of inattention and/or hyperactivity-impulsivity and may persists into adulthood. Childhood and adulthood ADHD are heritable, and are thought to represent the clinical extreme of a continuous distribution of ADHD symptoms in the general population. We aimed to leverage the power of studies of quantitative ADHD symptoms in adults who were genotyped. Within the SAGA (Study of ADHD trait genetics in adults) consortium, we estimated the single nucleotide polymorphism (SNP)-based heritability of quantitative self-reported ADHD symptoms and carried out a genome-wide association meta-analysis in nine adult population-based and case-only cohorts of adults. A total of n = 14,689 individuals were included. In two of the SAGA cohorts we found a significant SNP-based heritability for self-rated ADHD symptom scores of respectively 15% (n = 3656) and 30% (n = 1841). The top hit of the genome-wide meta-analysis (SNP rs12661753; p-value = 3.02 × 10-7) was present in the long non-coding RNA gene STXBP5-AS1. This association was also observed in a meta-analysis of childhood ADHD symptom scores in eight population-based pediatric cohorts from the Early Genetics and Lifecourse Epidemiology (EAGLE) ADHD consortium (n = 14,776). Genome-wide meta-analysis of the SAGA and EAGLE data (n = 29,465) increased the strength of the association with the SNP rs12661753. In human HEK293 cells, expression of STXBP5-AS1 enhanced the expression of a reporter construct of STXBP5, a gene known to be involved in "SNAP" (Soluble NSF attachment protein) Receptor" (SNARE) complex formation. In mouse strains featuring different levels of impulsivity, transcript levels in the prefrontal cortex of the mouse ortholog Gm28905 strongly correlated negatively with motor impulsivity as measured in the five choice serial reaction time task (r2 = - 0.61; p = 0.004). Our results are consistent with an effect of the STXBP5-AS1 gene on ADHD symptom scores distribution and point to a possible biological mechanism, other than antisense RNA inhibition, involved in ADHD-related impulsivity levels.

A rare missense variant in RCL1 segregates with depression in extended families.

Depression is the most prevalent psychiatric disorder with a complex and elusive etiology that is moderately heritable. Identification of genes would greatly facilitate the elucidation of the biological mechanisms underlying depression, however, its complex etiology has proved to be a major bottleneck in the identification of its genetic risk factors, especially in genome-wide association-like studies. In this study, we exploit the properties of a genetic isolate and its family-based structure to explore whether relatively rare exonic variants influence the burden of depressive symptoms in families. Using a multistep approach involving linkage and haplotype analyses followed by exome sequencing in the Erasmus Rucphen Family (ERF) study, we identified a rare (minor allele frequency (MAF)=1%) missense c.1114C>T mutation (rs115482041) in the RCL1 gene segregating with depression across multiple generations. Rs115482041 showed significant association with depressive symptoms (N=2393, ßT-allele=2.33, P-value=1 × 10-4) and explained 2.9% of the estimated genetic variance of depressive symptoms (22%) in ERF. Despite being twice as rare (MAF<0.5%), c.1114C>T showed similar effect and significant association with depressive symptoms in samples from the independent population-based Rotterdam study (N=1604, ßT-allele=3.60, P-value=3 × 10-2). A comparison of RCL1 expression in human and mouse brain revealed a striking co-localization of RCL1 with the layer 1 interlaminar subclass of astrocytes found exclusively in higher-order primates. Our findings identify RCL1 as a novel candidate gene for depression and offer insights into mechanisms through which RCL1 may be relevant for depression.

The protocadherin 17 gene affects cognition, personality, amygdala structure and function, synapse development and risk of major mood disorders.

Major mood disorders, which primarily include bipolar disorder and major depressive disorder, are the leading cause of disability worldwide and pose a major challenge in identifying robust risk genes. Here, we present data from independent large-scale clinical data sets (including 29 557 cases and 32 056 controls) revealing brain expressed protocadherin 17 (PCDH17) as a susceptibility gene for major mood disorders. Single-nucleotide polymorphisms (SNPs) spanning the PCDH17 region are significantly associated with major mood disorders; subjects carrying the risk allele showed impaired cognitive abilities, increased vulnerable personality features, decreased amygdala volume and altered amygdala function as compared with non-carriers. The risk allele predicted higher transcriptional levels of PCDH17 mRNA in postmortem brain samples, which is consistent with increased gene expression in patients with bipolar disorder compared with healthy subjects. Further, overexpression of PCDH17 in primary cortical neurons revealed significantly decreased spine density and abnormal dendritic morphology compared with control groups, which again is consistent with the clinical observations of reduced numbers of dendritic spines in the brains of patients with major mood disorders. Given that synaptic spines are dynamic structures which regulate neuronal plasticity and have crucial roles in myriad brain functions, this study reveals a potential underlying biological mechanism of a novel risk gene for major mood disorders involved in synaptic function and related intermediate phenotypes.

Exome-sequencing in a large population-based study reveals a rare Asn396Ser variant in the LIPG gene associated with depressive symptoms.

Despite a substantial genetic component, efforts to identify common genetic variation underlying depression have largely been unsuccessful. In the current study we aimed to identify rare genetic variants that might have large effects on depression in the general population. Using high-coverage exome-sequencing, we studied the exonic variants in 1265 individuals from the Rotterdam study (RS), who were assessed for depressive symptoms. We identified a missense Asn396Ser mutation (rs77960347) in the endothelial lipase (LIPG) gene, occurring with an allele frequency of 1% in the general population, which was significantly associated with depressive symptoms (P-value=5.2 × 10-08, ß=7.2). Replication in three independent data sets (N=3612) confirmed the association of Asn396Ser (P-value=7.1 × 10-03, ß=2.55) with depressive symptoms. LIPG is predicted to have enzymatic function in steroid biosynthesis, cholesterol biosynthesis and thyroid hormone metabolic processes. The Asn396Ser variant is predicted to have a damaging effect on the function of LIPG. Within the discovery population, carriers also showed an increased burden of white matter lesions (P-value=3.3 × 10-02) and a higher risk of Alzheimer's disease (odds ratio=2.01; P-value=2.8 × 10-02) compared with the non-carriers. Together, these findings implicate the Asn396Ser variant of LIPG in the pathogenesis of depressive symptoms in the general population.

Rare, low frequency and common coding variants in CHRNA5 and their contribution to nicotine dependence in European and African Americans.

The common nonsynonymous variant rs16969968 in the α5 nicotinic receptor subunit gene (CHRNA5) is the strongest genetic risk factor for nicotine dependence in European Americans and contributes to risk in African Americans. To comprehensively examine whether other CHRNA5 coding variation influences nicotine dependence risk, we performed targeted sequencing on 1582 nicotine-dependent cases (Fagerström Test for Nicotine Dependence score⩾4) and 1238 non-dependent controls, with independent replication of common and low frequency variants using 12 studies with exome chip data. Nicotine dependence was examined using logistic regression with individual common variants (minor allele frequency (MAF)⩾0.05), aggregate low frequency variants (0.05>MAF⩾0.005) and aggregate rare variants (MAF<0.005). Meta-analysis of primary results was performed with replication studies containing 12 174 heavy and 11 290 light smokers. Next-generation sequencing with 180 × coverage identified 24 nonsynonymous variants and 2 frameshift deletions in CHRNA5, including 9 novel variants in the 2820 subjects. Meta-analysis confirmed the risk effect of the only common variant (rs16969968, European ancestry: odds ratio (OR)=1.3, P=3.5 × 10(-11); African ancestry: OR=1.3, P=0.01) and demonstrated that three low frequency variants contributed an independent risk (aggregate term, European ancestry: OR=1.3, P=0.005; African ancestry: OR=1.4, P=0.0006). The remaining 22 rare coding variants were associated with increased risk of nicotine dependence in the European American primary sample (OR=12.9, P=0.01) and in the same risk direction in African Americans (OR=1.5, P=0.37). Our results indicate that common, low frequency and rare CHRNA5 coding variants are independently associated with nicotine dependence risk. These newly identified variants likely influence the risk for smoking-related diseases such as lung cancer.

Genome-wide analyses of borderline personality features.

The heritability of borderline personality (BP) features has been established in multiple twin and family studies. Using data from the borderline subscale of the Personality Assessment Inventory Borderline Features Scale (PAI-BOR) collected in two Dutch cohorts (N=7125), the Netherlands Twin Register and The Netherlands Study of Depression and Anxiety, we show that heritability of the PAI-BOR total score using genome-wide single-nucleotide polymorphism (SNPs) is estimated at 23%, and that the genetic variance is substantially higher in affect instability items compared with the other three subscales of the PAI-BOR (42.7% vs non-significant estimates for self-harm, negative relations and identity problems). We present results from a first genome-wide association study of BP features, which shows a promising signal on chromosome 5 corresponding to SERINC5, a protein involved in myelination. Reduced myelination has been suggested as possibly having a role in the development of psychiatric disorders characterized by lack of social interaction. The signal was confirmed in a third independent Dutch cohort drawn from the Erasmus Rucphen Family study (N=1301). Our analyses were complemented by investigating the heterogeneity that was implied by the differences in genetic variance components in the four subscales of the PAI-BOR. These analyses show that the association of SNPs tagging SERINC5 differs substantially across the 24 items of the PAI-BOR. Further, using reverse regression we showed that the effects were present only in subjects with higher scores on the PAI-BOR. Taken together, these results suggest that future genome-wide analyses can benefit substantially by taking into account the phenotypic and genetic heterogeneity of BP features.

ATP5H/KCTD2 locus is associated with Alzheimer's disease risk.

To identify loci associated with Alzheimer disease, we conducted a three-stage analysis using existing genome-wide association studies (GWAS) and genotyping in a new sample. In Stage I, all suggestive single-nucleotide polymorphisms (at P<0.001) in a previously reported GWAS of seven independent studies (8082 Alzheimer's disease (AD) cases; 12 040 controls) were selected, and in Stage II these were examined in an in silico analysis within the Cohorts for Heart and Aging Research in Genomic Epidemiology consortium GWAS (1367 cases and 12904 controls). Six novel signals reaching P<5 × 10(-6) were genotyped in an independent Stage III sample (the Fundació ACE data set) of 2200 sporadic AD patients and 2301 controls. We identified a novel association with AD in the adenosine triphosphate (ATP) synthase, H+ transporting, mitochondrial F0 (ATP5H)/Potassium channel tetramerization domain-containing protein 2 (KCTD2) locus, which reached genome-wide significance in the combined discovery and genotyping sample (rs11870474, odds ratio (OR)=1.58, P=2.6 × 10(-7) in discovery and OR=1.43, P=0.004 in Fundació ACE data set; combined OR=1.53, P=4.7 × 10(-9)). This ATP5H/KCTD2 locus has an important function in mitochondrial energy production and neuronal hyperpolarization during cellular stress conditions, such as hypoxia or glucose deprivation.

A genome-wide association meta-analysis of plasma Aß peptides concentrations in the elderly.

Amyloid beta (Aß) peptides are the major components of senile plaques, one of the main pathological hallmarks of Alzheimer disease (AD). However, Aß peptides' functions are not fully understood and seem to be highly pleiotropic. We hypothesized that plasma Aß peptides concentrations could be a suitable endophenotype for a genome-wide association study (GWAS) designed to (i) identify novel genetic factors involved in amyloid precursor protein metabolism and (ii) highlight relevant Aß-related physiological and pathophysiological processes. Hence, we performed a genome-wide association meta-analysis of four studies totaling 3 528 healthy individuals of European descent and for whom plasma Aß1-40 and Aß1-42 peptides levels had been quantified. Although we did not observe any genome-wide significant locus, we identified 18 suggestive loci (P<1 × 10(-)(5)). Enrichment-pathway analyses revealed canonical pathways mainly involved in neuronal functions, for example, axonal guidance signaling. We also assessed the biological impact of the gene most strongly associated with plasma Aß1-42 levels (cortexin 3, CTXN3) on APP metabolism in vitro and found that the gene protein was able to modulate Aß1-42 secretion. In conclusion, our study results suggest that plasma Aß peptides levels are valid endophenotypes in GWASs and can be used to characterize the metabolism and functions of APP and its metabolites.

Candidate-gene association study searching for genetic factors involved in migraine chronification.

INTRODUCTION: Chronic migraine (CM) is at the severe end of the clinical migraine spectrum, but its genetic background is unknown. Our study searched for evidence that genetic factors are involved in the chronification process. METHODS: We initially selected 144 single-nucleotide polymorphisms (SNPs) from 48 candidate genes, which we tested for association in two stages: The first stage encompassed 262 CM patients, the second investigated 226 patients with high-frequency migraine (HFM). Subsequently, SNPs with p values < 0.05 were forwarded to the replication stage containing 531 patients with CM or HFM. RESULTS: Eight SNPs were significantly associated with CM and HFM in the two-stage phase. None survived replication in the third stage. DISCUSSION: We present the first comprehensive genetic association study for migraine chronification. There were no significant findings. Future studies may benefit from larger, genome-wide data sets or should use other genetic approaches to identify genetic factors involved in migraine chronification.

Integrating metabolomics profiling measurements across multiple biobanks.

To optimize the quality of large scale mass-spectrometry based metabolomics data obtained from semiquantitative profiling measurements, it is important to use a strategy in which dedicated measurement designs are combined with a strict statistical quality control regime. This assures consistently high-quality results across measurements from individual studies, but semiquantitative data have been so far only comparable for samples measured within the same study. To enable comparability and integration of semiquantitative profiling data from different large scale studies over the time course of years, the measurement and quality control strategy has to be extended. We introduce a strategy to allow the integration of semiquantitative profiling data from different studies. We demonstrate that lipidomics data generated in samples from three different large biobanks acquired in the time course of 3 years can be effectively combined when using an appropriate measurement design and transfer model. This strategy paves the way toward an integrative usage of semiquantitative metabolomics data sets of multiple studies to validate biological findings in another study and/or to increase the statistical power for discovery of biomarkers or pathways by combining studies.

A genetic variant in SLC6A20 is associated with Type 2 diabetes in white-European and Chinese populations.

AIMS: To investigate whether polymorphisms in SLC6A20 are associated with susceptibility to Type 2 diabetes. METHODS: In the Rotterdam Study, a prospective, population-based cohort (n = 5974), 22 tagging polymorphisms with minor allele frequencies>0.05 across SLC6A20 were studied. Replication studies were performed in an independent Dutch case-control study (DiaGene-Rotterdam Study 2 n = 3133), and in a Chinese Han case-control population (n = 2279). A meta-analysis of the results was performed. RESULTS: In the Rotterdam study, the minor alleles of rs13062383, rs10461016 and rs2286489 increased the risk of Type 2 diabetes (hazard ratio 1.37, 95% CI 1.15-1.63, hazard ratio 1.30 95% CI 1.09-1.54 and hazard ratio 1.20, 95% CI 1.07-1.35, respectively). In the DiaGene/Rotterdam Study 2, the A allele of rs13062383 increased the risk of Type 2 diabetes (odds ratio 1.45, 95% CI 1.19-1.76). In the Chinese Han study, the rs13062383 A allele also increased the risk of Type 2 diabetes (odds ratio 1.21, 95% CI 1.03-1.42). Meta-analysis showed a highly significant association of rs13062383 with Type 2 diabetes (odds ratio 1.35, 95% CI 1.21-1.47; P = 3.3 × 10⁻8). CONCLUSIONS: In conclusion, rs13062383 in SLC6A20 increased the susceptibility to Type 2 diabetes in populations with different genetic backgrounds.

A K(ATP) channel gene effect on sleep duration: from genome-wide association studies to function in Drosophila.

Humans sleep approximately a third of their lifetime. The observation that individuals with either long or short sleep duration show associations with metabolic syndrome and psychiatric disorders suggests that the length of sleep is adaptive. Although sleep duration can be influenced by photoperiod (season) and phase of entrainment (chronotype), human familial sleep disorders indicate that there is a strong genetic modulation of sleep. Therefore, we conducted high-density genome-wide association studies for sleep duration in seven European populations (N=4251). We identified an intronic variant (rs11046205; P=3.99 × 10(-8)) in the ABCC9 gene that explains ≈5% of the variation in sleep duration. An influence of season and chronotype on sleep duration was solely observed in the replication sample (N=5949). Meta-analysis of the associations found in a subgroup of the replication sample, chosen for season of entry and chronotype, together with the discovery results showed genome-wide significance. RNA interference knockdown experiments of the conserved ABCC9 homologue in Drosophila neurons renders flies sleepless during the first 3 h of the night. ABCC9 encodes an ATP-sensitive potassium channel subunit (SUR2), serving as a sensor of intracellular energy metabolism.

Lipoprotein receptor-related protein 1 variants and dietary fatty acids: meta-analysis of European origin and African American studies.

OBJECTIVE: Low-density lipoprotein-related receptor protein 1 (LRP1) is a multi-functional endocytic receptor and signaling molecule that is expressed in adipose and the hypothalamus. Evidence for a role of LRP1 in adiposity is accumulating from animal and in vitro models, but data from human studies are limited. The study objectives were to evaluate (i) relationships between LRP1 genotype and anthropometric traits, and (ii) whether these relationships were modified by dietary fatty acids. DESIGN AND METHODS: We conducted race/ethnic-specific meta-analyses using data from 14 studies of US and European whites and 4 of African Americans to evaluate associations of dietary fatty acids and LRP1 genotypes with body mass index (BMI), waist circumference and hip circumference, as well as interactions between dietary fatty acids and LRP1 genotypes. Seven single-nucleotide polymorphisms (SNPs) of LRP1 were evaluated in whites (N up to 42 000) and twelve SNPs in African Americans (N up to 5800). RESULTS: After adjustment for age, sex and population substructure if relevant, for each one unit greater intake of percentage of energy from saturated fat (SFA), BMI was 0.104 kg m(-2) greater, waist was 0.305 cm larger and hip was 0.168 cm larger (all P<0.0001). Other fatty acids were not associated with outcomes. The association of SFA with outcomes varied by genotype at rs2306692 (genotyped in four studies of whites), where the magnitude of the association of SFA intake with each outcome was greater per additional copy of the T allele: 0.107 kg m(-2) greater for BMI (interaction P=0.0001), 0.267 cm for waist (interaction P=0.001) and 0.21 cm for hip (interaction P=0.001). No other significant interactions were observed. CONCLUSION: Dietary SFA and LRP1 genotype may interactively influence anthropometric traits. Further exploration of this, and other diet x genotype interactions, may improve understanding of interindividual variability in the relationships of dietary factors with anthropometric traits.

A genome-wide linkage study of individuals with high scores on NEO personality traits.

The NEO-Five-Factor Inventory divides human personality traits into five dimensions: neuroticism, extraversion, openness, conscientiousness and agreeableness. In this study, we sought to identify regions harboring genes with large effects on the five NEO personality traits by performing genome-wide linkage analysis of individuals scoring in the extremes of these traits (>90th percentile). Affected-only linkage analysis was performed using an Illumina 6K linkage array in a family-based study, the Erasmus Rucphen Family study. We subsequently determined whether distinct, segregating haplotypes found with linkage analysis were associated with the trait of interest in the population. Finally, a dense single-nucleotide polymorphism genotyping array (Illumina 318K) was used to search for copy number variations (CNVs) in the associated regions. In the families with extreme phenotype scores, we found significant evidence of linkage for conscientiousness to 20p13 (rs1434789, log of odds (LOD)=5.86) and suggestive evidence of linkage (LOD >2.8) for neuroticism to 19q, 21q and 22q, extraversion to 1p, 1q, 9p and12q, openness to 12q and 19q, and agreeableness to 2p, 6q, 17q and 21q. Further analysis determined haplotypes in 21q22 for neuroticism (P-values = 0.009, 0.007), in 17q24 for agreeableness (marginal P-value = 0.018) and in 20p13 for conscientiousness (marginal P-values = 0.058, 0.038) segregating in families with large contributions to the LOD scores. No evidence for CNVs in any of the associated regions was found. Our findings imply that there may be genes with relatively large effects involved in personality traits, which may be identified with next-generation sequencing techniques.

Delirium in the acute phase after stroke and the role of the apolipoprotein E gene.

OBJECTIVE: To study the association between the epsilon 4 allele of apolipoprotein E (APOEε4) and delirium in a stroke population. METHODS: 527 consecutive stroke patients were screened for delirium during the first week of admission with the confusion assessment method. In three hundred fifty-three patients genomic DNA isolation was available. RESULTS: The incidence of delirium after stroke in the 353 patients was 11.3%. There was no association between APOEε4 and delirium. Even after adjustment for IQCODE, stroke localization, stroke subtype, stroke severity, infection, and brain atrophy no association was found (odds ratio: 0.9; 95% confidence interval: 0.4-2.1). Delirium did not last longer in patients with an APOEε4 allele compared to patients without an APOEε4 allele (median: 5.6 days [range: 1-21] versus median: 4.6 days [range: 1-15], p = 0.5). CONCLUSION: There was no association between the presence of an APOEε4 allele and the occurrence of delirium in the acute phase after stroke.

Apolipoprotein E4 allele in a population-based study of early-onset Alzheimer's disease.

Several studies have reported an association of the apolipoprotein E allele epsilon 4 (APOE*4) to familial and sporadic late-onset Alzheimer's disease (LOAD). Here we report on the relationship between APOE*4 and early-onset Alzheimer's disease (EOAD) in a Dutch population-based study. The frequency of the APOE*4 allele was 2.3 times higher among EOAD cases compared to controls. Among patients, the allele frequency was 1.6 times higher in those with a positive family history than in those without. A significant increase in risk of EOAD was found for subjects homozygous for APOE*4 regardless of family history of dementia, but an increase in EOAD risk for APOE*4 heterozygotes could only be shown in subjects with a positive family history. Our study demonstrates a significant association between APOE*4 and EOAD which is modified by family history of dementia.

Presenile dementia and cerebral haemorrhage linked to a mutation at codon 692 of the beta-amyloid precursor protein gene.

Several families with an early-onset form of familial Alzheimer's disease have been found to harbour mutations at a specific codon (717) of the gene for the beta-amyloid precursor protein (APP) on chromosome 21. We now report, a novel base mutation in the same exon of the APP gene which co-segregates in one family with presenile dementia and cerebral haemorrhage due to cerebral amyloid angiopathy. The mutation results in the substitution of alanine into glycine at codon 692. These results suggest that the clinically distinct entities, presenile dementia and cerebral amyloid angiopathy, can be caused by the same mutation in the APP gene.