Deciphering osteoarthritis genetics across 826,690 individuals from 9 populations.

Osteoarthritis affects over 300 million people worldwide. Here, we conduct a genome-wide association study meta-analysis across 826,690 individuals (177,517 with osteoarthritis) and identify 100 independently associated risk variants across 11 osteoarthritis phenotypes, 52 of which have not been associated with the disease before. We report thumb and spine osteoarthritis risk variants and identify differences in genetic effects between weight-bearing and non-weight-bearing joints. We identify sex-specific and early age-at-onset osteoarthritis risk loci. We integrate functional genomics data from primary patient tissues (including articular cartilage, subchondral bone, and osteophytic cartilage) and identify high-confidence effector genes. We provide evidence for genetic correlation with phenotypes related to pain, the main disease symptom, and identify likely causal genes linked to neuronal processes. Our results provide insights into key molecular players in disease processes and highlight attractive drug targets to accelerate translation.

Epigenomic profiling of the infrapatellar fat pad in osteoarthritis.

Osteoarthritis is a prevalent, complex disease of the joints, and affects multiple intra-articular tissues. Here, we have examined genome-wide DNA methylation profiles of primary infrapatellar fat pad and matched blood samples from 70 osteoarthritis patients undergoing total knee replacement surgery. Comparing the DNA methylation profiles between these tissues reveal widespread epigenetic differences. We produce the first genome-wide methylation quantitative trait locus (mQTL) map of fat pad, and make the resource available to the wider community. Using two-sample Mendelian randomization and colocalization analyses, we resolve osteoarthritis GWAS signals and provide insights into the molecular mechanisms underpinning disease aetiopathology. Our findings provide the first view of the epigenetic landscape of infrapatellar fat pad primary tissue in osteoarthritis.

Identifying causal serum protein-cardiometabolic trait relationships using whole genome sequencing.

Cardiometabolic diseases, such as type 2 diabetes and cardiovascular disease, have a high public health burden. Understanding the genetically determined regulation of proteins that are dysregulated in disease can help to dissect the complex biology underpinning them. Here, we perform a protein quantitative trait locus (pQTL) analysis of 248 serum proteins relevant to cardiometabolic processes in 2893 individuals. Meta-analyzing whole-genome sequencing (WGS) data from two Greek cohorts, MANOLIS (n = 1356; 22.5× WGS) and Pomak (n = 1537; 18.4× WGS), we detect 301 independently associated pQTL variants for 170 proteins, including 12 rare variants (minor allele frequency < 1%). We additionally find 15 pQTL variants that are rare in non-Finnish European populations but have drifted up in the frequency in the discovery cohorts here. We identify proteins causally associated with cardiometabolic traits, including Mep1b for high-density lipoprotein (HDL) levels, and describe a knock-out (KO) Mep1b mouse model. Our findings furnish insights into the genetic architecture of the serum proteome, identify new protein-disease relationships and demonstrate the importance of isolated populations in pQTL analysis.

Genetic Landscape of the ACE2 Coronavirus Receptor.

BACKGROUND: SARS-CoV-2, the causal agent of COVID-19, enters human cells using the ACE2 (angiotensin-converting enzyme 2) protein as a receptor. ACE2 is thus key to the infection and treatment of the coronavirus. ACE2 is highly expressed in the heart and respiratory and gastrointestinal tracts, playing important regulatory roles in the cardiovascular and other biological systems. However, the genetic basis of the ACE2 protein levels is not well understood. METHODS: We have conducted the largest genome-wide association meta-analysis of plasma ACE2 levels in >28 000 individuals of the SCALLOP Consortium (Systematic and Combined Analysis of Olink Proteins). We summarize the cross-sectional epidemiological correlates of circulating ACE2. Using the summary statistics-based high-definition likelihood method, we estimate relevant genetic correlations with cardiometabolic phenotypes, COVID-19, and other human complex traits and diseases. We perform causal inference of soluble ACE2 on vascular disease outcomes and COVID-19 severity using mendelian randomization. We also perform in silico functional analysis by integrating with other types of omics data. RESULTS: We identified 10 loci, including 8 novel, capturing 30% of the heritability of the protein. We detected that plasma ACE2 was genetically correlated with vascular diseases, severe COVID-19, and a wide range of human complex diseases and medications. An X-chromosome cis-protein quantitative trait loci-based mendelian randomization analysis suggested a causal effect of elevated ACE2 levels on COVID-19 severity (odds ratio, 1.63 [95% CI, 1.10-2.42]; P=0.01), hospitalization (odds ratio, 1.52 [95% CI, 1.05-2.21]; P=0.03), and infection (odds ratio, 1.60 [95% CI, 1.08-2.37]; P=0.02). Tissue- and cell type-specific transcriptomic and epigenomic analysis revealed that the ACE2 regulatory variants were enriched for DNA methylation sites in blood immune cells. CONCLUSIONS: Human plasma ACE2 shares a genetic basis with cardiovascular disease, COVID-19, and other related diseases. The genetic architecture of the ACE2 protein is mapped, providing a useful resource for further biological and clinical studies on this coronavirus receptor.

Proteomic analysis of 92 circulating proteins and their effects in cardiometabolic diseases.

BACKGROUND: Human plasma contains a wide variety of circulating proteins. These proteins can be important clinical biomarkers in disease and also possible drug targets. Large scale genomics studies of circulating proteins can identify genetic variants that lead to relative protein abundance. METHODS: We conducted a meta-analysis on genome-wide association studies of autosomal chromosomes in 22,997 individuals of primarily European ancestry across 12 cohorts to identify protein quantitative trait loci (pQTL) for 92 cardiometabolic associated plasma proteins. RESULTS: We identified 503 (337 cis and 166 trans) conditionally independent pQTLs, including several novel variants not reported in the literature. We conducted a sex-stratified analysis and found that 118 (23.5%) of pQTLs demonstrated heterogeneity between sexes. The direction of effect was preserved but there were differences in effect size and significance. Additionally, we annotate trans-pQTLs with nearest genes and report plausible biological relationships. Using Mendelian randomization, we identified causal associations for 18 proteins across 19 phenotypes, of which 10 have additional genetic colocalization evidence. We highlight proteins associated with a constellation of cardiometabolic traits including angiopoietin-related protein 7 (ANGPTL7) and Semaphorin 3F (SEMA3F). CONCLUSION: Through large-scale analysis of protein quantitative trait loci, we provide a comprehensive overview of common variants associated with plasma proteins. We highlight possible biological relationships which may serve as a basis for further investigation into possible causal roles in cardiometabolic diseases.

Population-wide copy number variation calling using variant call format files from 6,898 individuals.

Copy number variants (CNVs) play an important role in a number of human diseases, but the accurate calling of CNVs remains challenging. Most current approaches to CNV detection use raw read alignments, which are computationally intensive to process. We use a regression tree-based approach to call germline CNVs from whole-genome sequencing (WGS, >18x) variant call sets in 6,898 samples across four European cohorts, and describe a rich large variation landscape comprising 1,320 CNVs. Eighty-one percent of detected events have been previously reported in the Database of Genomic Variants. Twenty-three percent of high-quality deletions affect entire genes, and we recapitulate known events such as the GSTM1 and RHD gene deletions. We test for association between the detected deletions and 275 protein levels in 1,457 individuals to assess the potential clinical impact of the detected CNVs. We describe complex CNV patterns underlying an association with levels of the CCL3 protein (MAF = 0.15, p = 3.6x10-12 ) at the CCL3L3 locus, and a novel cis-association between a low-frequency NOMO1 deletion and NOMO1 protein levels (MAF = 0.02, p = 2.2x10-7 ). This study demonstrates that existing population-wide WGS call sets can be mined for germline CNVs with minimal computational overhead, delivering insight into a less well-studied, yet potentially impactful class of genetic variant.

Very low-depth whole-genome sequencing in complex trait association studies.

MOTIVATION: Very low-depth sequencing has been proposed as a cost-effective approach to capture low-frequency and rare variation in complex trait association studies. However, a full characterization of the genotype quality and association power for very low-depth sequencing designs is still lacking. RESULTS: We perform cohort-wide whole-genome sequencing (WGS) at low depth in 1239 individuals (990 at 1× depth and 249 at 4× depth) from an isolated population, and establish a robust pipeline for calling and imputing very low-depth WGS genotypes from standard bioinformatics tools. Using genotyping chip, whole-exome sequencing (75× depth) and high-depth (22×) WGS data in the same samples, we examine in detail the sensitivity of this approach, and show that imputed 1× WGS recapitulates 95.2% of variants found by imputed GWAS with an average minor allele concordance of 97% for common and low-frequency variants. In our study, 1× further allowed the discovery of 140 844 true low-frequency variants with 73% genotype concordance when compared to high-depth WGS data. Finally, using association results for 57 quantitative traits, we show that very low-depth WGS is an efficient alternative to imputed GWAS chip designs, allowing the discovery of up to twice as many true association signals than the classical imputed GWAS design. AVAILABILITY AND IMPLEMENTATION: The HELIC genotype and WGS datasets have been deposited to the European Genome-phenome Archive (https://www.ebi.ac.uk/ega/home): EGAD00010000518; EGAD00010000522; EGAD00010000610; EGAD00001001636, EGAD00001001637. The peakplotter software is available at https://github.com/wtsi-team144/peakplotter, the transformPhenotype app can be downloaded at https://github.com/wtsi-team144/transformPhenotype. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Evaluation of shared genetic aetiology between osteoarthritis and bone mineral density identifies SMAD3 as a novel osteoarthritis risk locus.

Osteoarthritis (OA) is a common complex disease with high public health burden and no curative therapy. High bone mineral density (BMD) is associated with an increased risk of developing OA, suggesting a shared underlying biology. Here, we performed the first systematic overlap analysis of OA and BMD on a genome wide scale. We used summary statistics from the GEFOS consortium for lumbar spine (n = 31,800) and femoral neck (n = 32,961) BMD, and from the arcOGEN consortium for three OA phenotypes (hip, ncases=3,498; knee, ncases=3,266; hip and/or knee, ncases=7,410; ncontrols=11,009). Performing LD score regression we found a significant genetic correlation between the combined OA phenotype (hip and/or knee) and lumbar spine BMD (rg=0.18, P = 2.23 × 10-2), which may be driven by the presence of spinal osteophytes. We identified 143 variants with evidence for cross-phenotype association which we took forward for replication in independent large-scale OA datasets, and subsequent meta-analysis with arcOGEN for a total sample size of up to 23,425 cases and 236,814 controls. We found robustly replicating evidence for association with OA at rs12901071 (OR 1.08 95% CI 1.05-1.11, Pmeta=3.12 × 10-10), an intronic variant in the SMAD3 gene, which is known to play a role in bone remodeling and cartilage maintenance. We were able to confirm expression of SMAD3 in intact and degraded cartilage of the knee and hip. Our findings provide the first systematic evaluation of pleiotropy between OA and BMD, highlight genes with biological relevance to both traits, and establish a robust new OA genetic risk locus at SMAD3.

Very low-depth sequencing in a founder population identifies a cardioprotective APOC3 signal missed by genome-wide imputation.

Cohort-wide very low-depth whole-genome sequencing (WGS) can comprehensively capture low-frequency sequence variation for the cost of a dense genome-wide genotyping array. Here, we analyse 1x sequence data across the APOC3 gene in a founder population from the island of Crete in Greece (n = 1239) and find significant evidence for association with blood triglyceride levels with the previously reported R19X cardioprotective null mutation (ß = -1.09,σ = 0.163, P = 8.2 × 10-11) and a second loss of function mutation, rs138326449 (ß = -1.17,σ = 0.188, P = 1.14 × 10-9). The signal cannot be recapitulated by imputing genome-wide genotype data on a large reference panel of 5122 individuals including 249 with 4x WGS data from the same population. Gene-level meta-analysis with other studies reporting burden signals at APOC3 provides robust evidence for a replicable cardioprotective rare variant aggregation (P = 3.2 × 10-31, n = 13 480).

The mountainous Cretan dietary patterns and their relationship with cardiovascular risk factors: the Hellenic Isolated Cohorts MANOLIS study.

OBJECTIVE: We carried out de novo recruitment of a population-based cohort (MANOLIS study) and describe the specific population, which displays interesting characteristics in terms of diet and health in old age, through deep phenotyping. DESIGN: Cross-sectional study where anthropometric, biochemical and clinical measurements were taken in addition to interview-based completion of an extensive questionnaire on health and lifestyle parameters. Dietary patterns were derived through principal component analysis based on a validated FFQ. SETTING: Geographically isolated Mylopotamos villages on Mount Idi, Crete, Greece. SUBJECTS: Adults (n 1553). RESULTS: Mean age of the participants was 61·6 years and 55·8 % were women. Of the population, 82·7 % were overweight or obese with a significantly different prevalence between overweight men and women (43·4 v. 34·7 %, P=0·002). The majority (70·6 %) of participants were married, while a larger proportion of women were widowed than men (27·8 v. 3·5 %, P<0·001). Smoking was more prevalent in men (38·7 v. 8·2 %, P<0·001), as 88·8% of women had never smoked. Four dietary patterns emerged as characteristic of the population; these were termed 'local', 'high fat and sugar, 'Greek café/tavern' and 'olive oil, fruits and vegetables'. Individuals more adherent to the local dietary pattern presented higher blood glucose (ß=4·026, P<0·001). Similarly, individuals with higher compliance with the Greek café/tavern pattern had higher waist-to-hip ratio (ß=0·012, P<0·001), blood pressure (ß=1·015, P=0·005) and cholesterol (ß=5·398, P<0·001). CONCLUSIONS: Profiling of the MANOLIS elderly population identifies unique unhealthy dietary patterns that are associated with cardiometabolic indices.

TE-Tracker: systematic identification of transposition events through whole-genome resequencing.

BACKGROUND: Transposable elements (TEs) are DNA sequences that are able to move from their location in the genome by cutting or copying themselves to another locus. As such, they are increasingly recognized as impacting all aspects of genome function. With the dramatic reduction in cost of DNA sequencing, it is now possible to resequence whole genomes in order to systematically characterize novel TE mobilization in a particular individual. However, this task is made difficult by the inherently repetitive nature of TE sequences, which in some eukaryotes compose over half of the genome sequence. Currently, only a few software tools dedicated to the detection of TE mobilization using next-generation-sequencing are described in the literature. They often target specific TEs for which annotation is available, and are only able to identify families of closely related TEs, rather than individual elements. RESULTS: We present TE-Tracker, a general and accurate computational method for the de-novo detection of germ line TE mobilization from re-sequenced genomes, as well as the identification of both their source and destination sequences. We compare our method with the two classes of existing software: specialized TE-detection tools and generic structural variant (SV) detection tools. We show that TE-Tracker, while working independently of any prior annotation, bridges the gap between these two approaches in terms of detection power. Indeed, its positive predictive value (PPV) is comparable to that of dedicated TE software while its sensitivity is typical of a generic SV detection tool. TE-Tracker demonstrates the benefit of adopting an annotation-independent, de novo approach for the detection of TE mobilization events. We use TE-Tracker to provide a comprehensive view of transposition events induced by loss of DNA methylation in Arabidopsis. TE-Tracker is freely available at http://www.genoscope.cns.fr/TE-Tracker . CONCLUSIONS: We show that TE-Tracker accurately detects both the source and destination of novel transposition events in re-sequenced genomes. Moreover, TE-Tracker is able to detect all potential donor sequences for a given insertion, and can identify the correct one among them. Furthermore, TE-Tracker produces significantly fewer false positives than common SV detection programs, thus greatly facilitating the detection and analysis of TE mobilization events.

GWAS of lipids in Greenlanders finds association signals shared with Europeans and reveals an independent PCSK9 association signal.

Perturbation of lipid homoeostasis is a major risk factor for cardiovascular disease (CVD), the leading cause of death worldwide. We aimed to identify genetic variants affecting lipid levels, and thereby risk of CVD, in Greenlanders. Genome-wide association studies (GWAS) of six blood lipids, triglycerides, LDL-cholesterol, HDL-cholesterol, total cholesterol, as well as apolipoproteins A1 and B, were performed in up to 4473 Greenlanders. For genome-wide significant variants, we also tested for associations with additional traits, including CVD events. We identified 11 genome-wide significant loci associated with lipid traits. Most of these loci were already known in Europeans, however, we found a potential causal variant near PCSK9 (rs12117661), which was independent of the known PCSK9 loss-of-function variant (rs11491147). rs12117661 was associated with lower LDL-cholesterol (ßSD(SE) = -0.22 (0.03), p = 6.5 × 10-12) and total cholesterol (-0.17 (0.03), p = 1.1 × 10-8) in the Greenlandic study population. Similar associations were observed in Europeans from the UK Biobank, where the variant was also associated with a lower risk of CVD outcomes. Moreover, rs12117661 was a top eQTL for PCSK9 across tissues in European data from the GTEx portal, and was located in a predicted regulatory element, supporting a possible causal impact on PCSK9 expression. Combined, the 11 GWAS signals explained up to 16.3% of the variance of the lipid traits. This suggests that the genetic architecture of lipid levels in Greenlanders is different from Europeans, with fewer variants explaining the variance.

Bridging the diversity gap: Analytical and study design considerations for improving the accuracy of trans-ancestry genetic prediction.

Genetic prediction of common complex disease risk is an essential component of precision medicine. Currently, genome-wide association studies (GWASs) are mostly composed of European-ancestry samples and resulting polygenic scores (PGSs) have been shown to poorly transfer to other ancestries partly due to heterogeneity of allelic effects between populations. Fixed-effects (FETA) and random-effects (RETA) trans-ancestry meta-analyses do not model such ancestry-related heterogeneity, while ancestry-specific (AS) scores may suffer from low power due to low sample sizes. In contrast, trans-ancestry meta-regression (TAMR) builds ancestry-aware PGS that account for more complex trans-ancestry architectures. Here, we examine the predictive performance of these four PGSs under multiple genetic architectures and ancestry configurations. We show that the predictive performance of FETA and RETA is strongly affected by cross-ancestry genetic heterogeneity, while AS PGS performance decreases in under-represented target populations. TAMR PGS is also impacted by heterogeneity but maintains good prediction performance in most situations, especially in ancestry-diverse scenarios. In simulations of human complex traits, TAMR scores currently explain 25% more phenotypic variance than AS in triglyceride levels and 33% more phenotypic variance than FETA in type 2 diabetes in most non-European populations. Importantly, a high proportion of non-European-ancestry individuals is needed to reach prediction levels that are comparable in those populations to the one observed in European-ancestry studies. Our results highlight the need to rebalance the ancestral composition of GWAS to enable accurate prediction in non-European-ancestry groups, and demonstrate the relevance of meta-regression approaches for compensating some of the current population biases in GWAS.

Genome-wide meta-analysis of 92 cardiometabolic protein serum levels.

OBJECTIVES: Global cardiometabolic disease prevalence has grown rapidly over the years, making it the leading cause of death worldwide. Proteins are crucial components in biological pathways dysregulated in disease states. Identifying genetic components that influence circulating protein levels may lead to the discovery of biomarkers for early stages of disease or offer opportunities as therapeutic targets. METHODS: Here, we carry out a genome-wide association study (GWAS) utilising whole genome sequencing data in 3,005 individuals from the HELIC founder populations cohort, across 92 proteins of cardiometabolic relevance. RESULTS: We report 322 protein quantitative trait loci (pQTL) signals across 92 proteins, of which 76 are located in or near the coding gene (cis-pQTL). We link those association signals with changes in protein expression and cardiometabolic disease risk using colocalisation and Mendelian randomisation (MR) analyses. CONCLUSIONS: The majority of previously unknown signals we describe point to proteins or protein interactions involved in inflammation and immune response, providing genetic evidence for the contributing role of inflammation in cardiometabolic disease processes.

A novel splice-affecting HNF1A variant with large population impact on diabetes in Greenland.

Background: The genetic disease architecture of Inuit includes a large number of common high-impact variants. Identification of such variants contributes to our understanding of the genetic aetiology of diseases and improves global equity in genomic personalised medicine. We aimed to identify and characterise novel variants in genes associated with Maturity Onset Diabetes of the Young (MODY) in the Greenlandic population. Methods: Using combined data from Greenlandic population cohorts of 4497 individuals, including 448 whole genome sequenced individuals, we screened 14 known MODY genes for previously identified and novel variants. We functionally characterised an identified novel variant and assessed its association with diabetes prevalence and cardiometabolic traits and population impact. Findings: We identified a novel variant in the known MODY gene HNF1A with an allele frequency of 1.9% in the Greenlandic Inuit and absent elsewhere. Functional assays indicate that it prevents normal splicing of the gene. The variant caused lower 30-min insulin (ß = -232 pmol/L, ßSD = -0.695, P = 4.43 × 10-4) and higher 30-min glucose (ß = 1.20 mmol/L, ßSD = 0.441, P = 0.0271) during an oral glucose tolerance test. Furthermore, the variant was associated with type 2 diabetes (OR 4.35, P = 7.24 × 10-6) and HbA1c (ß = 0.113 HbA1c%, ßSD = 0.205, P = 7.84 × 10-3). The variant explained 2.5% of diabetes variance in Greenland. Interpretation: The reported variant has the largest population impact of any previously reported variant within a MODY gene. Together with the recessive TBC1D4 variant, we show that close to 1 in 5 cases of diabetes (18%) in Greenland are associated with high-impact genetic variants compared to 1-3% in large populations. Funding: Novo Nordisk Foundation, Independent Research Fund Denmark, and Karen Elise Jensen's Foundation.

Genetic mechanisms of 184 neuro-related proteins in human plasma.

Understanding the genetic basis of neuro-related proteins is essential for dissecting the disease etiology of neuropsychiatric disorders and other complex traits and diseases. Here, the SCALLOP Consortium conducted a genome-wide association meta-analysis of over 12,500 individuals for 184 neuro-reiated proteins in human plasma. The analysis identified 117 cis-regulatory protein quantitative trait loci (cis-pQTL) and 166 trans-pQTL. The mapped pQTL capture on average 50% of each protein's heritability. Mendelian randomization analyses revealed multiple proteins showing potential causal effects on neuro-reiated traits as well as complex diseases such as hypertension, high cholesterol, immune-related disorders, and psychiatric disorders. Integrating with established drug information, we validated 13 combinations of protein targets and diseases or side effects with available drugs, while suggesting hundreds of re-purposing and new therapeutic targets for diseases and comorbidities. This consortium effort provides a large-scale proteogenomic resource for biomedical research.

Unraveling Neuro-Proteogenomic Landscape and Therapeutic Implications for Human Behaviors and Psychiatric Disorders.

Understanding the genetic basis of neuro-related proteins is essential for dissecting the molecular basis of human behavioral traits and the disease etiology of neuropsychiatric disorders. Here, the SCALLOP Consortium conducted a genome-wide association meta-analysis of over 12,500 individuals for 184 neuro-related proteins in human plasma. The analysis identified 117 cis-regulatory protein quantitative trait loci (cis-pQTL) and 166 trans-pQTL. The mapped pQTL capture on average 50% of each protein's heritability. Mendelian randomization analyses revealed multiple proteins showing potential causal effects on neuro-related traits such as sleeping, smoking, feelings, alcohol intake, mental health, and psychiatric disorders. Integrating with established drug information, we validated 13 out of 13 matched combinations of protein targets and diseases or side effects with available drugs, while suggesting hundreds of re-purposing and new therapeutic targets. This consortium effort provides a large-scale proteogenomic resource for biomedical research on human behaviors and other neuro-related phenotypes.

Gene-based whole genome sequencing meta-analysis of 250 circulating proteins in three isolated European populations.

OBJECTIVE: Deep sequencing offers unparalleled access to rare variants in human populations. Understanding their role in disease is a priority, yet prohibitive sequencing costs mean that many cohorts lack the sample size to discover these effects on their own. Meta-analysis of individual variant scores allows the combination of rare variants across cohorts and study of their aggregated effect at the gene level, boosting discovery power. However, the methods involved have largely not been field-tested. In this study, we aim to perform the first meta-analysis of gene-based rare variant aggregation optimal tests, applied to the human cardiometabolic proteome. METHODS: Here, we carry out this analysis across MANOLIS, Pomak and ORCADES, three isolated European cohorts with whole-genome sequencing (total N = 4,422). We examine the genetic architecture of 250 proteomic traits of cardiometabolic relevance. We use a containerised pipeline to harmonise variant lists across cohorts and define four sets of qualifying variants. For every gene, we interrogate protein-damaging variants, exonic variants, exonic and regulatory variants, and regulatory only variants, using the CADD and Eigen scores to weigh variants according to their predicted functional consequence. We perform single-cohort rare variant analysis and meta-analyse variant scores using the SMMAT package. RESULTS: We describe 5 rare variant pQTLs (RV-pQTL) which pass our stringent significance threshold (7.45 × 10-11) and quality control procedure. These were split between four cis signals for MARCO, TEK, MMP2 and MPO, and one trans association for GDF2 in the SERPINA11 gene. We show that the cis-MPO association, which was not detectable using the single-point data alone, is driven by 5 missense and frameshift variants. These include rs140636390 and rs119468010, which are specific to MANOLIS and ORCADES, respectively. We show how this kind of signal could improve the predictive accuracy of genetic factors in common complex disease such as stroke and cardiovascular disease. CONCLUSIONS: Our proof-of-concept study demonstrates the power of gene-based meta-analyses for discovering disease-relevant associations complementing common-variant signals by incorporating population-specific rare variation.

Insights into the genetic architecture of haematological traits from deep phenotyping and whole-genome sequencing for two Mediterranean isolated populations.

Haematological traits are linked to cardiovascular, metabolic, infectious and immune disorders, as well as cancer. Here, we examine the role of genetic variation in shaping haematological traits in two isolated Mediterranean populations. Using whole-genome sequencing data at 22× depth for 1457 individuals from Crete (MANOLIS) and 1617 from the Pomak villages in Greece, we carry out a genome-wide association scan for haematological traits using linear mixed models. We discover novel associations (p < 5 × 10-9) of five rare non-coding variants with alleles conferring effects of 1.44-2.63 units of standard deviation on red and white blood cell count, platelet and red cell distribution width. Moreover, 10.0% of individuals in the Pomak population and 6.8% in MANOLIS carry a pathogenic mutation in the Haemoglobin Subunit Beta (HBB) gene. The mutational spectrum is highly diverse (10 different mutations). The most frequent mutation in MANOLIS is the common Mediterranean variant IVS-I-110 (G>A) (rs35004220). In the Pomak population, c.364C>A ("HbO-Arab", rs33946267) is most frequent (4.4% allele frequency). We demonstrate effects on haematological and other traits, including bilirubin, cholesterol, and, in MANOLIS, height and gestation age. We find less severe effects on red blood cell traits for HbS, HbO, and IVS-I-6 (T>C) compared to other b+ mutations. Overall, we uncover allelic diversity of HBB in Greek isolated populations and find an important role for additional rare variants outside of HBB.