Body mass index stratified meta-analysis of genome-wide association studies of polycystic ovary syndrome in women of European ancestry.

BACKGROUND: Polycystic ovary syndrome (PCOS) is a complex multifactorial disorder with a substantial genetic component. However, the clinical manifestations of PCOS are heterogeneous with notable differences between lean and obese women, implying a different pathophysiology manifesting in differential body mass index (BMI). We performed a meta-analysis of genome-wide association study (GWAS) data from six well-characterised cohorts, using a case-control study design stratified by BMI, aiming to identify genetic variants associated with lean and overweight/obese PCOS subtypes. RESULTS: The study comprised 254,588 women (5,937 cases and 248,651 controls) from individual studies performed in Australia, Estonia, Finland, the Netherlands and United States of America, and separated according to three BMI stratifications (lean, overweight and obese). Genome-wide association analyses were performed for each stratification within each cohort, with the data for each BMI group meta-analysed using METAL software. Almost half of the total study population (47%, n = 119,584) were of lean BMI (≤ 25 kg/m2). Two genome-wide significant loci were identified for lean PCOS, led by rs12000707 within DENND1A (P = 1.55 × 10-12) and rs2228260 within XBP1 (P = 3.68 × 10-8). One additional locus, LINC02905, was highlighted as significantly associated with lean PCOS through gene-based analyses (P = 1.76 × 10-6). There were no significant loci observed for the overweight or obese sub-strata when analysed separately, however, when these strata were combined, an association signal led by rs569675099 within DENND1A reached genome-wide significance (P = 3.22 × 10-9) and a gene-based association was identified with ERBB4 (P = 1.59 × 10-6). Nineteen of 28 signals identified in previous GWAS, were replicated with consistent allelic effect in the lean stratum. There were less replicated signals in the overweight and obese groups, and only 4 SNPs were replicated in each of the three BMI strata. CONCLUSIONS: Genetic variation at the XBP1, LINC02905 and ERBB4 loci were associated with PCOS within unique BMI strata, while DENND1A demonstrated associations across multiple strata, providing evidence of both distinct and shared genetic features between lean and overweight/obese PCOS-affected women. This study demonstrated that PCOS-affected women with contrasting body weight are not only phenotypically distinct but also show variation in genetic architecture; lean PCOS women typically display elevated gonadotrophin ratios, lower insulin resistance, higher androgen levels, including adrenal androgens, and more favourable lipid profiles. Overall, these findings add to the growing body of evidence supporting a genetic basis for PCOS as well as differences in genetic patterns relevant to PCOS BMI-subtype.

Epigenome-wide Association Study Shows Differential DNA Methylation of MDC1, KLF9, and CUTA in Autoimmune Thyroid Disease.

CONTEXT: Autoimmune thyroid disease (AITD) includes Graves disease (GD) and Hashimoto disease (HD), which often run in the same family. AITD etiology is incompletely understood: Genetic factors may account for up to 75% of phenotypic variance, whereas epigenetic effects (including DNA methylation [DNAm]) may contribute to the remaining variance (eg, why some individuals develop GD and others HD). OBJECTIVE: This work aimed to identify differentially methylated positions (DMPs) and differentially methylated regions (DMRs) comparing GD to HD. METHODS: Whole-blood DNAm was measured across the genome using the Infinium MethylationEPIC array in 32 Australian patients with GD and 30 with HD (discovery cohort) and 32 Danish patients with GD and 32 with HD (replication cohort). Linear mixed models were used to test for differences in quantile-normalized ß values of DNAm between GD and HD and data were later meta-analyzed. Comb-p software was used to identify DMRs. RESULTS: We identified epigenome-wide significant differences (P < 9E-8) and replicated (P < .05) 2 DMPs between GD and HD (cg06315208 within MDC1 and cg00049440 within KLF9). We identified and replicated a DMR within CUTA (5 CpGs at 6p21.32). We also identified 64 DMPs and 137 DMRs in the meta-analysis. CONCLUSION: Our study reveals differences in DNAm between GD and HD, which may help explain why some people develop GD and others HD and provide a link to environmental risk factors. Additional research is needed to advance understanding of the role of DNAm in AITD and investigate its prognostic and therapeutic potential.

Age-dependent genetic regulation of osteoarthritis: independent effects of immune system genes.

OBJECTIVES: Osteoarthritis (OA) is a joint disease with a heritable component. Genetic loci identified via genome-wide association studies (GWAS) account for an estimated 26.3% of the disease trait variance in humans. Currently, there is no method for predicting the onset or progression of OA. We describe the first use of the Collaborative Cross (CC), a powerful genetic resource, to investigate knee OA in mice, with follow-up targeted multi-omics analysis of homologous regions of the human genome. METHODS: We histologically screened 275 mice for knee OA and conducted quantitative trait locus (QTL) mapping in the complete cohort (> 8 months) and the younger onset sub-cohort (8-12 months). Multi-omic analysis of human genetic datasets was conducted to investigate significant loci. RESULTS: We observed a range of OA phenotypes. QTL mapping identified a genome-wide significant locus on mouse chromosome 19 containing Glis3, the human equivalent of which has been identified as associated with OA in recent GWAS. Mapping the younger onset sub-cohort identified a genome-wide significant locus on chromosome 17. Multi-omic analysis of the homologous region of the human genome (6p21.32) indicated the presence of pleiotropic effects on the expression of the HLA - DPB2 gene and knee OA development risk, potentially mediated through the effects on DNA methylation. CONCLUSIONS: The significant associations at the 6p21.32 locus in human datasets highlight the value of the CC model of spontaneous OA that we have developed and lend support for an immune role in the disease. Our results in mice also add to the accumulating evidence of a role for Glis3 in OA.

Leveraging osteoclast genetic regulatory data to identify genes with a role in osteoarthritis.

There has been a growing interest in the role of the subchondral bone and its resident osteoclasts in the progression of osteoarthritis (OA). A recent genome-wide association study (GWAS) identified 100 independent association signals for OA traits. Most of these signals are led by noncoding variants, suggesting that genetic regulatory effects may drive many of the associations. We have generated a unique human osteoclast-like cell-specific expression quantitative trait locus (eQTL) resource for studying the genetics of bone disease. Considering the potential role of osteoclasts in the pathogenesis of OA, we performed an integrative analysis of this dataset with the recently published OA GWAS results. Summary data-based Mendelian randomization (SMR) and colocalization analyses identified 38 genes with a potential role in OA, including some that have been implicated in Mendelian diseases with joint/skeletal abnormalities, such as BICRA, EIF6, CHST3, and FBN2. Several OA GWAS signals demonstrated colocalization with more than one eQTL peak, including at 19q13.32 (hip OA with BCAM, PRKD2, and BICRA eQTL). We also identified a number of eQTL signals colocalizing with more than one OA trait, including FAM53A, GCAT, HMGN1, MGAT4A, RRP7BP, and TRIOBP. An SMR analysis identified 3 loci with evidence of pleiotropic effects on OA-risk and gene expression: LINC01481, CPNE1, and EIF6. Both CPNE1 and EIF6 are located at 20q11.22, a locus harboring 2 other strong OA candidate genes, GDF5 and UQCC1, suggesting the presence of an OA-risk gene cluster. In summary, we have used our osteoclast-specific eQTL dataset to identify genes potentially involved with the pathogenesis of OA.

Bone Trans-omics: Integrating Omics to Unveil Mechanistic Molecular Networks Regulating Bone Biology and Disease.

PURPOSE OF REVIEW: Recent advancements in "omics" technologies and bioinformatics have afforded researchers new tools to study bone biology in an unbiased and holistic way. The purpose of this review is to highlight recent studies integrating multi-omics data gathered from multiple molecular layers (i.e.; trans-omics) to reveal new molecular mechanisms that regulate bone biology and underpin skeletal diseases. RECENT FINDINGS: Bone biologists have traditionally relied on single-omics technologies (genomics, transcriptomics, proteomics, and metabolomics) to profile measureable differences (both qualitative and quantitative) of individual molecular layers for biological discovery and to investigate mechanisms of disease. Recently, literature has grown on the implementation of integrative multi-omics to study bone biology, which combines computational and informatics support to connect multiple layers of data derived from individual "omic" platforms. This emerging discipline termed "trans-omics" has enabled bone biologists to identify and construct detailed molecular networks, unveiling new pathways and unexpected interactions that have advanced our mechanistic understanding of bone biology and disease. While the era of trans-omics is poised to revolutionize our capacity to answer more complex and diverse questions pertinent to bone pathobiology, it also brings new challenges that are inherent when trying to connect "Big Data" sets. A concerted effort between bone biologists and interdisciplinary scientists will undoubtedly be needed to extract physiologically and clinically meaningful data from bone trans-omics in order to advance its implementation in the field.

Bone mineral density loci specific to the skull portray potential pleiotropic effects on craniosynostosis.

Skull bone mineral density (SK-BMD) provides a suitable trait for the discovery of key genes in bone biology, particularly to intramembranous ossification, not captured at other skeletal sites. We perform a genome-wide association meta-analysis (n ~ 43,800) of SK-BMD, identifying 59 loci, collectively explaining 12.5% of the trait variance. Association signals cluster within gene-sets involved in skeletal development and osteoporosis. Among the four novel loci (ZIC1, PRKAR1A, AZIN1/ATP6V1C1, GLRX3), there are factors implicated in intramembranous ossification and as we show, inherent to craniosynostosis processes. Functional follow-up in zebrafish confirms the importance of ZIC1 on cranial suture patterning. Likewise, we observe abnormal cranial bone initiation that culminates in ectopic sutures and reduced BMD in mosaic atp6v1c1 knockouts. Mosaic prkar1a knockouts present asymmetric bone growth and, conversely, elevated BMD. In light of this evidence linking SK-BMD loci to craniofacial abnormalities, our study provides new insight into the pathophysiology, diagnosis and treatment of skeletal diseases.

Identification of Differentially Expressed Genes and Molecular Pathways Involved in Osteoclastogenesis Using RNA-seq.

Osteoporosis is a disease that is characterised by reduced bone mineral density (BMD) and can be exacerbated by the excessive bone resorption of osteoclasts (OCs). Bioinformatic methods, including functional enrichment and network analysis, can provide information about the underlying molecular mechanisms that participate in the progression of osteoporosis. In this study, we harvested human OC-like cells differentiated in culture and their precursor peripheral blood mononuclear cells (PBMCs) and characterised the transcriptome of the two cell types using RNA-sequencing in order to identify differentially expressed genes. Differential gene expression analysis was performed in RStudio using the edgeR package. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to identify enriched GO terms and signalling pathways, with inter-connected regions characterised using protein-protein interaction analysis. In this study, we identified 3201 differentially expressed genes using a 5% false discovery rate; 1834 genes were upregulated, whereas 1367 genes were downregulated. We confirmed a significant upregulation of several well-established OC genes including CTSK, DCSTAMP, ACP5, MMP9, ITGB3, and ATP6V0D2. The GO analysis suggested that upregulated genes are involved in cell division, cell migration, and cell adhesion, while the KEGG pathway analysis highlighted oxidative phosphorylation, glycolysis and gluconeogenesis, lysosome, and focal adhesion pathways. This study provides new information about changes in gene expression and highlights key biological pathways involved in osteoclastogenesis.

Sugar transporter Slc37a2 regulates bone metabolism in mice via a tubular lysosomal network in osteoclasts.

Osteoclasts are giant bone-digesting cells that harbor specialized lysosome-related organelles termed secretory lysosomes (SLs). SLs store cathepsin K and serve as a membrane precursor to the ruffled border, the osteoclast's 'resorptive apparatus'. Yet, the molecular composition and spatiotemporal organization of SLs remains incompletely understood. Here, using organelle-resolution proteomics, we identify member a2 of the solute carrier 37 family (Slc37a2) as a SL sugar transporter. We demonstrate in mice that Slc37a2 localizes to the SL limiting membrane and that these organelles adopt a hitherto unnoticed but dynamic tubular network in living osteoclasts that is required for bone digestion. Accordingly, mice lacking Slc37a2 accrue high bone mass owing to uncoupled bone metabolism and disturbances in SL export of monosaccharide sugars, a prerequisite for SL delivery to the bone-lining osteoclast plasma membrane. Thus, Slc37a2 is a physiological component of the osteoclast's unique secretory organelle and a potential therapeutic target for metabolic bone diseases.

Functional Assessment of Calcium-Sensing Receptor Variants Confirms Familial Hypocalciuric Hypercalcemia.

Context: In the clinic it is important to differentiate primary hyperparathyroidism (PHPT) from the more benign, inherited disorder, familial hypocalciuric hypercalcemia (FHH). Since the conditions may sometimes overlap biochemically, identification of calcium-sensing receptor (CASR) gene variants causative of FHH (but not PHPT) is the most decisive diagnostic aid. When novel variants are identified, bioinformatics and functional assessment are required to establish pathogenicity. Objective: We identified 3 novel CASR transmembrane domain missense variants, Thr699Asn, Arg701Gly, and Thr808Pro, in 3 probands provisionally diagnosed with FHH and examined the variants using bioinformatics and functional analysis. Methods: Bioinformatics assessment utilized wANNOVAR software. For functional characterization, each variant was cloned into a mammalian expression vector; wild-type and variant receptors were transfected into HEK293 cells, and their expression and cellular localization were assessed by Western blotting and confocal immunofluorescence, respectively. Receptor activation in HEK293 cells was determined using an IP-One ELISA assay following stimulation with Ca++ ions. Results: Bioinformatics analysis of the variants was unable to definitively assign pathogenicity. Compared with wild-type receptor, all variants demonstrated impaired expression of mature receptor reaching the cell surface and diminished activation at physiologically relevant Ca++ concentrations. Conclusion: Three CASR missense variants identified in probands provisionally diagnosed with FHH result in receptor inactivation and are therefore likely causative of FHH. Inactivation may be due to inadequate processing/trafficking of mature receptor and/or conformational changes induced by the variants affecting receptor signaling. This study demonstrates the value of functional studies in assessing genetic variants identified in hypercalcemic patients.

Molecular structure, expression, and the emerging role of Siglec-15 in skeletal biology and cancer.

Siglec-15, a Siglec family member and type-1 transmembrane protein, is expressed mainly in human macrophages and dendritic cells. It is comprised of a lysine-containing transmembrane domain, two extracellular immunoglobulin (Ig)-like domains and a short cytoplasmic domain. Siglec-15 is highly conserved in vertebrates and acts as an immunoreceptor. It exerts diverse functions on osteoclast physiology as well as the tumor microenvironment. Siglec-15 interacts with adapter protein DAP12 - Syk signaling pathway to regulate the RANKL/RANK-mediated PI3K, AKT, and ERK signaling pathways during osteoclast formation in vitro. Consistently, the lack of the Siglec-15 gene in mice leads to impaired osteoclast activity and osteopetrosis in vivo. In addition, Siglec-15 is expressed by tumor-associated macrophages (TAMs) and regulates the tumor microenvironment by activating the SYK/MAPK signaling pathway. Interestingly, Siglec-15 shares sequence homology to programmed death-ligand 1 (PD-L1) and has a potential immune-regulatory role in cancer immunology. Thus, Siglec-15 might also represent an alternative target for the treatment of cancers that do not respond to anti-PD-L1/PD-1 immunotherapy. Understanding the role of Siglec-15 in osteoclastogenesis and the tumor microenvironment will help us to develop new treatments for bone disorders and cancer.

Genome-wide analysis of thyroid function in Australian adolescents highlights SERPINA7 and NCOA3.

OBJECTIVE: Genetic factors underpin the narrow intraindividual variability of thyroid function, although precise contributions of environmental vs genetic factors remain uncertain. We sought to clarify the heritability of thyroid function traits and thyroid peroxidase antibody (TPOAb) positivity and identify single nucleotide polymorphisms (SNPs) contributing to the trait variance. METHODS: Heritability of thyroid-stimulating hormone (TSH), free T4 (fT4), free T3 (fT3) and TPOAb in a cohort of 2854 euthyroid, dizygous and monozygous twins (age range 11.9-16.9 years) from the Brisbane Longitudinal Twin Study (BLTS) was assessed using structural equation modelling. A genome-wide analysis was conducted on 2832 of these individuals across 7 522 526 SNPs as well as gene-based association analyses. Replication analysis of the association results was performed in the Raine Study (n = 1115) followed by meta-analysis to maximise power for discovery. RESULTS: Heritability of thyroid function parameters in the BLTS was 70.8% (95% CI: 66.7-74.9%) for TSH, 67.5% (59.8-75.3%) for fT4, 59.7% (54.4-65.0%) for fT3 and 48.8% (40.6-56.9%) for TPOAb. The genome-wide association study (GWAS) in the discovery cohort identified a novel association between rs2026401 upstream of NCOA3 and TPOAb. GWAS meta-analysis found associations between TPOAb and rs445219, also near NCOA3, and fT3 and rs12687280 near SERPINA7. Gene-based association analysis highlighted SERPINA7 for fT3 and NPAS3 for fT4. CONCLUSION: Our findings resolve former contention regarding heritability estimates of thyroid function traits and TPOAb positivity. GWAS and gene-based association analysis identified variants accounting for a component of this heritability.

The bone marrow microenvironment of pre-B acute lymphoblastic leukemia at single-cell resolution.

The bone marrow microenvironment (BMM) plays a key role in leukemia progression, but its molecular complexity in pre-B cell acute lymphoblastic leukemia (B-ALL), the most common cancer in children, remains poorly understood. To gain further insight, we used single-cell RNA sequencing to characterize the kinetics of the murine BMM during B-ALL progression. Normal pro- and pre-B cells were found to be the most affected at the earliest stages of disease and this was associated with changes in expression of genes regulated by the AP1-transcription factor complex and regulatory factors NELFE, MYC and BCL11A. Granulocyte-macrophage progenitors show reduced expression of the tumor suppressor long non-coding RNA Neat1 and disruptions in the rate of transcription. Intercellular communication networks revealed monocyte-dendritic precursors to be consistently active during B-ALL progression, with enriched processes including cytokine-mediated signaling pathway, neutrophil-mediated immunity and regulation of cell migration and proliferation. In addition, we confirmed that the hematopoietic stem and progenitor cell compartment was perturbed during leukemogenesis. These findings extend our understanding of the complexity of changes and molecular interactions among the normal cells of the BMM during B-ALL progression.

Characterisation of genetic regulatory effects for osteoporosis risk variants in human osteoclasts.

BACKGROUND: Osteoporosis is a complex disease with a strong genetic contribution. A recently published genome-wide association study (GWAS) for estimated bone mineral density (eBMD) identified 1103 independent genome-wide significant association signals. Most of these variants are non-coding, suggesting that regulatory effects may drive many of the associations. To identify genes with a role in osteoporosis, we integrate the eBMD GWAS association results with those from our previous osteoclast expression quantitative trait locus (eQTL) dataset. RESULTS: We identify sixty-nine significant cis-eQTL effects for eBMD GWAS variants after correction for multiple testing. We detect co-localisation of eBMD GWAS and osteoclast eQTL association signals for 21 of the 69 loci, implicating a number of genes including CCR5, ZBTB38, CPE, GNA12, RIPK3, IQGAP1 and FLCN. Summary-data-based Mendelian Randomisation analysis of the eBMD GWAS and osteoclast eQTL datasets identifies significant associations for 53 genes, with TULP4 presenting as a strong candidate for pleiotropic effects on eBMD and gene expression in osteoclasts. By performing analysis using the GARFIELD software, we demonstrate significant enrichment of osteoporosis risk variants among high-confidence osteoclast eQTL across multiple GWAS P value thresholds. Mice lacking one of the genes of interest, the apoptosis/necroptosis gene RIPK3, show disturbed bone micro-architecture and increased osteoclast number, highlighting a new biological pathway relevant to osteoporosis. CONCLUSION: We utilise a unique osteoclast eQTL dataset to identify a number of potential effector genes for osteoporosis risk variants, which will help focus functional studies in this area.

Correction: Large-scale genome-wide meta-analysis of polycystic ovary syndrome suggests shared genetic architecture for different diagnosis criteria.

[This corrects the article DOI: 10.1371/journal.pgen.1007813.].

Advanced Genetic Approaches in Discovery and Characterization of Genes Involved With Osteoporosis in Mouse and Human.

Osteoporosis is a complex condition with contributions from, and interactions between, multiple genetic loci and environmental factors. This review summarizes key advances in the application of genetic approaches for the identification of osteoporosis susceptibility genes. Genome-wide linkage analysis (GWLA) is the classical approach for identification of genes that cause monogenic diseases; however, it has shown limited success for complex diseases like osteoporosis. In contrast, genome-wide association studies (GWAS) have successfully identified over 200 osteoporosis susceptibility loci with genome-wide significance, and have provided most of the candidate genes identified to date. Phenome-wide association studies (PheWAS) apply a phenotype-to-genotype approach which can be used to complement GWAS. PheWAS is capable of characterizing the association between osteoporosis and uncommon and rare genetic variants. Another alternative approach, whole genome sequencing (WGS), will enable the discovery of uncommon and rare genetic variants in osteoporosis. Meta-analysis with increasing statistical power can offer greater confidence in gene searching through the analysis of combined results across genetic studies. Recently, new approaches to gene discovery include animal phenotype based models such as the Collaborative Cross and ENU mutagenesis. Site-directed mutagenesis and genome editing tools such as CRISPR/Cas9, TALENs and ZNFs have been used in functional analysis of candidate genes in vitro and in vivo. These resources are revolutionizing the identification of osteoporosis susceptibility genes through the use of genetically defined inbred mouse libraries, which are screened for bone phenotypes that are then correlated with known genetic variation. Identification of osteoporosis-related susceptibility genes by genetic approaches enables further characterization of gene function in animal models, with the ultimate aim being the identification of novel therapeutic targets for osteoporosis.

Genetic regulatory mechanisms in human osteoclasts suggest a role for the STMP1 and DCSTAMP genes in Paget's disease of bone.

Paget's disease of bone (PDB) is characterised by focal abnormalities of bone remodelling, with increased osteoclastic resorption the primary feature of the disease. Genetic factors have been shown to play an important role in PDB, and genome-wide association studies (GWAS) have identified 7 genetic loci as associated with PDB at the genome-wide level. Expression quantitative trait locus (eQTL) studies using cell types that are directly relevant to the disease of interest are increasingly being used to identify putative effector genes for GWAS loci. We have recently constructed a unique osteoclast-specific eQTL resource using cells differentiated in vitro from 158 subjects for study of the genetics of bone disease. Considering the major role osteoclasts have in PDB, we used this resource to investigate potential genetic regulatory effects for the 7 PDB genome-wide significant loci on genes located within 500 kb of each locus. After correction for multiple testing, we observed statistically significant associations for rs4294134 with expression of the gene STMP1, and rs2458413 with expression of the genes DPYS and DCSTAMP. The eQTL associations observed for rs4294134 with STMP1, and rs2458413 with DCSTAMP were further supported by eQTL data from other tissue types. The product of the STMP1 gene has not been extensively studied, however the DCSTAMP gene has an established role in osteoclast differentiation and the associations seen between rs2458413 and PDB are likely mediated through regulatory effects on this gene. This study highlights the value of eQTL data in determining which genes are relevant to GWAS loci.

Identification of Novel Loci Associated With Hip Shape: A Meta-Analysis of Genomewide Association Studies.

We aimed to report the first genomewide association study (GWAS) meta-analysis of dual-energy X-ray absorptiometry (DXA)-derived hip shape, which is thought to be related to the risk of both hip osteoarthritis and hip fracture. Ten hip shape modes (HSMs) were derived by statistical shape modeling using SHAPE software, from hip DXA scans in the Avon Longitudinal Study of Parents and Children (ALSPAC; adult females), TwinsUK (mixed sex), Framingham Osteoporosis Study (FOS; mixed), Osteoporotic Fractures in Men study (MrOS), and Study of Osteoporotic Fractures (SOF; females) (total N = 15,934). Associations were adjusted for age, sex, and ancestry. Five genomewide significant (p < 5 × 10-9 , adjusted for 10 independent outcomes) single-nucleotide polymorphisms (SNPs) were associated with HSM1, and three SNPs with HSM2. One SNP, in high linkage disequilibrium with rs2158915 associated with HSM1, was associated with HSM5 at genomewide significance. In a look-up of previous GWASs, three of the identified SNPs were associated with hip osteoarthritis, one with hip fracture, and five with height. Seven SNPs were within 200 kb of genes involved in endochondral bone formation, namely SOX9, PTHrP, RUNX1, NKX3-2, FGFR4, DICER1, and HHIP. The SNP adjacent to DICER1 also showed osteoblast cis-regulatory activity of GSC, in which mutations have previously been reported to cause hip dysplasia. For three of the lead SNPs, SNPs in high LD (r2 > 0.5) were identified, which intersected with open chromatin sites as detected by ATAC-seq performed on embryonic mouse proximal femora. In conclusion, we identified eight SNPs independently associated with hip shape, most of which were associated with height and/or mapped close to endochondral bone formation genes, consistent with a contribution of processes involved in limb growth to hip shape and pathological sequelae. These findings raise the possibility that genetic studies of hip shape might help in understanding potential pathways involved in hip osteoarthritis and hip fracture. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals, Inc.

Large-scale genome-wide meta-analysis of polycystic ovary syndrome suggests shared genetic architecture for different diagnosis criteria.

Polycystic ovary syndrome (PCOS) is a disorder characterized by hyperandrogenism, ovulatory dysfunction and polycystic ovarian morphology. Affected women frequently have metabolic disturbances including insulin resistance and dysregulation of glucose homeostasis. PCOS is diagnosed with two different sets of diagnostic criteria, resulting in a phenotypic spectrum of PCOS cases. The genetic similarities between cases diagnosed based on the two criteria have been largely unknown. Previous studies in Chinese and European subjects have identified 16 loci associated with risk of PCOS. We report a fixed-effect, inverse-weighted-variance meta-analysis from 10,074 PCOS cases and 103,164 controls of European ancestry and characterisation of PCOS related traits. We identified 3 novel loci (near PLGRKT, ZBTB16 and MAPRE1), and provide replication of 11 previously reported loci. Only one locus differed significantly in its association by diagnostic criteria; otherwise the genetic architecture was similar between PCOS diagnosed by self-report and PCOS diagnosed by NIH or non-NIH Rotterdam criteria across common variants at 13 loci. Identified variants were associated with hyperandrogenism, gonadotropin regulation and testosterone levels in affected women. Linkage disequilibrium score regression analysis revealed genetic correlations with obesity, fasting insulin, type 2 diabetes, lipid levels and coronary artery disease, indicating shared genetic architecture between metabolic traits and PCOS. Mendelian randomization analyses suggested variants associated with body mass index, fasting insulin, menopause timing, depression and male-pattern balding play a causal role in PCOS. The data thus demonstrate 3 novel loci associated with PCOS and similar genetic architecture for all diagnostic criteria. The data also provide the first genetic evidence for a male phenotype for PCOS and a causal link to depression, a previously hypothesized comorbid disease. Thus, the genetics provide a comprehensive view of PCOS that encompasses multiple diagnostic criteria, gender, reproductive potential and mental health.

Expression Quantitative Trait Locus Study of Bone Mineral Density GWAS Variants in Human Osteoclasts.

Osteoporosis is a complex disease with a strong genetic component. Genomewide association studies (GWAS) have been very successful at identifying common genetic variants associated with bone parameters. A recently published study documented the results of the largest GWAS for bone mineral density (BMD) performed to date (n = 142,487), identifying 307 conditionally independent single-nucleotide polymorphisms (SNPs) as associated with estimated BMD (eBMD) at the genomewide significance level. The vast majority of these variants are non-coding SNPs. Expression quantitative trait locus (eQTL) studies using disease-specific cell types have increasingly been integrated with the results from GWAS to identify genes through which the observed GWAS associations are likely mediated. We generated a unique human osteoclast-specific eQTL data set using cells differentiated in vitro from 158 participants. We then used this resource to characterize the 307 recently identified BMD GWAS SNPs for association with nearby genes (±500 kb). After correction for multiple testing, 24 variants were found to be significantly associated with the expression of 32 genes in the osteoclast-like cells. Bioinformatics analysis suggested that these variants and those in strong linkage disequilibrium with them are enriched in regulatory regions. Several of the eQTL associations identified are relevant to genes that present strongly as having a role in bone, particularly IQGAP1, CYP19A1, CTNNB1, and COL6A3. Supporting evidence for many of the associations was obtained from publicly available eQTL data sets. We have also generated strong evidence for the presence of a regulatory region on chromosome 15q21.2 relevant to both the GLDN and CYP19A1 genes. In conclusion, we have generated a unique osteoclast-specific eQTL resource and have used this to identify 32 eQTL associations for recently identified BMD GWAS loci, which should inform functional studies of osteoclast biology. © 2018 American Society for Bone and Mineral Research.

Life-Course Genome-wide Association Study Meta-analysis of Total Body BMD and Assessment of Age-Specific Effects.

Bone mineral density (BMD) assessed by DXA is used to evaluate bone health. In children, total body (TB) measurements are commonly used; in older individuals, BMD at the lumbar spine (LS) and femoral neck (FN) is used to diagnose osteoporosis. To date, genetic variants in more than 60 loci have been identified as associated with BMD. To investigate the genetic determinants of TB-BMD variation along the life course and test for age-specific effects, we performed a meta-analysis of 30 genome-wide association studies (GWASs) of TB-BMD including 66,628 individuals overall and divided across five age strata, each spanning 15 years. We identified variants associated with TB-BMD at 80 loci, of which 36 have not been previously identified; overall, they explain approximately 10% of the TB-BMD variance when combining all age groups and influence the risk of fracture. Pathway and enrichment analysis of the association signals showed clustering within gene sets implicated in the regulation of cell growth and SMAD proteins, overexpressed in the musculoskeletal system, and enriched in enhancer and promoter regions. These findings reveal TB-BMD as a relevant trait for genetic studies of osteoporosis, enabling the identification of variants and pathways influencing different bone compartments. Only variants in ESR1 and close proximity to RANKL showed a clear effect dependency on age. This most likely indicates that the majority of genetic variants identified influence BMD early in life and that their effect can be captured throughout the life course.