Human Genetics

Primary open-angle glaucoma (POAG) disproportionately affects individuals of African ancestry, yet rare coding variation in this population remains understudied. To address this gap, we performed a multi-cohort exome-wide meta-analysis across POAAGG, PMBB, All of Us, and UK Biobank, including 4,815 POAG cases and 22,922 controls of genetically inferred African ancestry. Although no gene reached exome-wide significance, we identified several suggestive gene-level associations driven by rare variants (minor allele frequency [≤]0.1% or singletons),including signals in SRF, BLTP3A, METTL2A, and KRT10. Among these, SRF demonstrated the strongest association and was driven by rare missense variants with moderate effect sizes. Given its role in cytoskeletal organization and actin dynamics; processes central to trabecular meshwork function and intraocular pressure regulation SRF represents a biologically plausible candidate gene. Notably, these genes have not been previously highlighted in predominantly European ancestry POAG association studies, suggesting potential ancestry-specific rare variant contributions. Overall, our findings highlight the critical importance of investigating rare coding variation in POAG, in disproportionately affected populations to deepen understanding of POAG etiology and genetic risk.

ImportanceGenome-wide association studies have identified hundreds of common single nucleotide polymorphisms (SNPs) and small insertions/deletions (indels) associated with primary open-angle glaucoma (POAG) risk, though these variants have modest effect sizes and individually may have minor contributions to disease development. As whole-genome sequencing data is becoming more readily available, structural variants and other complex genomic features can be interrogated for contribution to disease risk. ObjectiveTest the association of structural variants in known glaucoma loci with disease risk. DesignCross-sectional study. SettingA multicenter cohort of individuals from the United States who contributed genomic and electronic health record data to the All of Us Research Program. ParticipantsPOAG case/control cohorts were generated in the All of Us Researcher Workbench using age (>40 for cases, >65 for controls) and ICD 9/10 diagnosis codes. Main Outcomes and MeasuresLogistic regression analyses adjusted for age, sex, and the top 10 principal components of ancestry were used to test association of structural variants within 500 kilobases of 309 known open-angle glaucoma risk loci. The significance threshold after Bonferroni correction was set at p<1.6x10-4. Results516 POAG cases and 18,716 controls of European ancestry from the All of Us v8 data release were included in the analysis. Mean age was 77.0 years among cases and 74.7 years among controls. Females comprised 45.7% of cases and 56.5% of controls. An 8,732 base pair deletion upstream of PITX2 (chr4:110680827-110689558) was associated with 7.3-fold higher odds of POAG (95% confidence interval: 2.9-18.5, p= 2.4x10-5, variant carrier frequency= 1.6% in cases and 0.25% in controls). Functional annotation identified multiple enhancers overlapping the deletion, suggesting that this structural variant likely impacts gene regulation and expression. Conclusion and RelevanceWhole genome sequencing data captures rare structural variants with large effect sizes that are missed by conventional SNP and indel genotyping approaches, enabling improved POAG risk stratification. These data also expand the phenotypic spectrum of structural variation in the PITX2 locus from childhood glaucoma to adult-onset disease, where age at diagnosis and clinical severity may be influenced by the extent of disrupted regulatory elements.

Age-related hearing loss (ARHL) is a progressive, bilateral decline in hearing ability that affects one in four individuals over 60 years of age worldwide. While previous genome-wide association studies (GWAS) have identified distinct single-nucleotide variants (SNVs) associated with metabolic and sensory ARHL phenotypes, the contribution of short tandem repeats (STRs) - a neglected yet important class of genetic variants - remains poorly understood. To address this gap, TRTools was used to impute STRs from a high quality, sequencing-derived SNV-STR reference panel to investigate the association between STRs and metabolic and sensory estimates. Heritability analyses revealed that while STRs contribute to estimates of both ARHL components, this class of variation plays a more important role in metabolic hearing loss (6%), which typically increases with age, compared to sensory hearing loss (4%). Further, the inclusion of this class of variant into GWAS analyses uncovered an association between a haplotype consisting of two missense variants (rs7714670 and rs6453022) and an intronic STR (chr5:73778077:A16) in ARHGEF28 (P=3.30x10-9), proving further insight into the variants driving this previously identified signal. Notably, burden analyses revealed that rare and longer repeats were associated with an increased risk of the metabolic phenotype and a reduced risk of the sensory phenotype. Functional annotation of significant and nominally significant STRs revealed potential effects on gene expression and splicing of nearby genes. Our findings provide the first evidence that STRs explain some of the missing heritability of ARHL phenotypes and create an STR resource for researchers to use in future analyses.

Age-related hearing loss is a widespread sensory impairment affecting a significant proportion of the elderly population, yet the genetic underpinnings of this condition remain incompletely understood. In this study, we investigate a non-synonymous variant (rs2242416) in the CRIP3 gene, which is expressed in auditory hair cells, in the context of hearing loss. Firstly, we find the variant shows strong and consistent association with hearing loss across multiple genome-wide association studies. Secondly, this variant, substitutes the nonpolar isoleucine for the polar threonine at an amino acid site that is otherwise highly conserved across placental mammals. Thirdly, by causing the amino acid change, the variant subtly alters the structure of the CRIP3 protein. Together, these three analyses provide phenotypic, evolutionary, and molecular evidence for the functionality of CRIP3 and its role in hearing loss. Moreover, the population genetics of the CRIP3 locus reveals an increased frequency of the derived threonine allele of rs2242416 in Eurasian populations following the out-of-Africa migration of humans more than 50,000 years ago. Nevertheless, the role of Darwinian selection in this increased frequency remains inconclusive. Overall, our results make a compelling argument to auditory researchers to make a CRIP3 mouse model to pinpoint the precise role of the protein in auditory function. Such a model will pave the way for therapeutic interventions targeting the CRIP3 protein to mitigate age-related hearing loss.

We conducted the first genome-wide association meta-analyses of global and sectoral peripapillary retinal nerve fibre layer (pRNFL) thickness and Bruchs membrane opening-minimum rim width (BMO-MRW), the major optic nerve head structural and neurodegeneration biomarkers, including up to 25,942 and 12,080 participants, respectively, from the International Glaucoma Genetics Consortium. We identified 9 global pRNFL thickness and 9 global BMO-MRW loci, along with 28 and 19 loci for pRNFL and BMO-MRW sectors, respectively, comprising both shared and sector-specific loci. To identify intraocular pressure (IOP)-independent drug targets, global pRNFL thickness and BMO-MRW were conditioned on IOP. IOP-independent loci were then prioritised to identify candidate causal genes using transcriptome-wide association study and colocalization analysis. Several genes, such as NMNAT2 and TRIOBP, had robust associations with both phenotypes, with potential IOP-independent therapeutic translation for glaucoma. Overall, we identified novel loci for pRNFL thickness and BMO-MRW, highlighting potential drug-target genes acting independently from IOP, and elucidating genetic differences among pRNFL sectors.

Microtia is a common congenital craniofacial malformation characterized by the partial or complete absence of the external ear structure. Despite its relatively high incidence, the pathogenesis of microtia remain poorly understood. In this study, we analyzed both single-cell and bulk RNA sequencing data from microtia cases and identified a population of COL1+HES1+ mesenchymal stem cell in perichondrium with significantly higher expression of the CRABP2 gene, a gene that encodes a nuclear transporter of retinoic acid. Gene expression analysis further confirmed that the RA signaling intensity and stemness are both higher in COL1+HES1+ perichondral stem cells from microtia patients, possibly due to elevated CRABP2 levels. Through histological verification we further confirmed the presence of this cell population with high CRABP2 expression in the perichondrium. Mechanistically, the elevated CRABP2 expression in perichondral stem cells seen in microtia patients may cause dysregulated RA signaling and disrupt the regulation of stem cell differentiation during auricular development. Histological analysis further revealed higher KLF2 expression as well as cartilage hypoplasia in microtia samples. Our study identified that the CRABP2-induced RA dysregulation in COL1+HES1+ perichondral stem cells may contribute to microtia. These findings offer new insights into the etiology of microtia and provide potential directions for prenatal prevention and tissue engineering treatments.

BackgroundThe transport of transfer RNAs (tRNAs) from the nucleus to the cytoplasm is a crucial step in the regulation of gene expression and protein synthesis. This process is mediated by specialized export molecules, among which XPOT (Exportin-t, XPO3) plays a central role by recognizing and transporting mature tRNAs through the nuclear pore complex. XPOT is not essential in RNA trafficking in the simple organisms, however the potential impact of XPOT deficiency in human health remains unresolved. MethodsWe identified eight patients from five unrelated families with rare biallelic germline variants in XPOT resulting in putative loss-of-function. Functional analyses were carried out in patient-derived fibroblasts, lymphoblastoid cells and zebrafish models. Ex vivo immunohistochemical stainings for Xpot were performed in the mouse cochlea. xpot knockout zebrafish models were generated to assess the morphology and hearing ability. ResultsAll patients presented with a uniform clinical phenotype that included increased susceptibility to infection, bronchiectasis, severe sensorineural hearing loss, developmental delay, and growth retardation. We demonstrated a complete absence of XPOT protein expression in three patient-derived cell lines. XPOT deficiency leads to disruptions in protein synthesis of the cytokine TNF pathway upon cellular stimulation. Additional XPO1 inhibition in XPOT deficient cells had little effect on cellular functions, suggesting alternative tRNA nuclear transporter pathways. Increased XPOT immunoreactivity was observed in type I spiral ganglion neurons and hair cells of the mouse cochlea, with enrichment in stereocilia. xpot knockout zebrafish model showed dysmorphic features, and reduced hearing, recapitulating key patient phenotypes. ConclusionsOur findings establish a direct connection between impaired XPOT-dependent tRNA export and human pathology. It illustrates that perturbations in nuclear export pathways lead to disease. It also raises the possibility that other nuclear transport receptors may play similarly underappreciated roles in human health and disease. The identification of XPOT as a disease-associated gene opens up new research directions and potential targets for therapeutic intervention.

Menieres disease (MD) is a chronic inner ear disorder characterized by recurrent vertigo, fluctuating sensorineural hearing loss, and tinnitus. Despite these distinctive symptoms, its etiology remains poorly understood. We performed a genome-wide meta-analysis of 8,969 cases and 1,962,542 controls across five large biobanks, identifying five independent genome-wide significant loci and estimating an observed-scale SNP heritability of 7% (SE 0.8%), consistent with a modest but significant genetic contribution to MD risk. Fine-mapping and integrative functional analyses implicate two convergent biological processes - developmental regulation of the inner ear, involving EYA4, EYA1, and LMO4 - and retinoic acid metabolism, with loci near CYP26A1/C1 and ALDH1A2 suggesting disrupted RA signaling in sensory and fluid-pressure homeostasis. These developmental regulator genes are robustly expressed in fetal and adult human inner ear cell types, supporting a model in which altered developmental programs predispose to adult vestibular and auditory dysfunction. Phenome-wide and genetic correlation analyses further reveal shared genetic architecture between MD and related traits, including vertigo, tinnitus, hearing loss, migraine, and sleep apnea, situating MD within a broader spectrum of sensory and neurological disorders. Collectively, these findings establish a genetic framework for Menieres disease risk and implicate developmental regulators and retinoic acid signaling as key contributing pathways.

Defects in motile cilia cause a range of disorders, including heterotaxy (HTX), congenital heart disease (CHD), and primary ciliary dyskinesia (PCD). Although these conditions often co-occur, the genetic and mechanistic bases for tissue-specific manifestations remain poorly understood. Here, we identify compound heterozygous variants in DAW1, a dynein arm assembly factor, in a proband with HTX and complex congenital heart disease but no clinical signs of PCD. Whole-genome sequencing revealed a maternally inherited canonical splice-site variant (c.648+1G>A) and a paternally inherited missense variant (c.341G>A; p.Arg114Gln), both classified as variants of uncertain significance under ACMG/AMP guidelines. Using Xenopus tropicalis, we show that Daw1 depletion disrupts left-right patterning, cardiac looping, and mucociliary flow, all of which are rescued by wild-type human DAW1. Functional testing of patient alleles showed notable tissue specificity: p.Arg114Gln fully rescued mucociliary flow but did not restore left-right patterning, while the splice-site variant resulted in a complete loss of function in both contexts. These findings closely match the probands clinical phenotype and provide strong functional evidence to support reclassifying c.648+1G>A as pathogenic and p.Arg114Gln as a context-dependent hypomorphic allele. This study establishes functional criteria for interpreting DAW1 variants, shows how developmental context clarifies genotype-phenotype relationships, and highlights how in vivo models can support ACMG reclassification of unresolved HTX-related variants.

Congenital anomalies of the kidney and urinary tract (CAKUT) are the primary cause of pediatric kidney failure, yet the genetic etiologies remain elusive for most affected individuals. Reanalysis of trio exome sequencing data from 80 Chinese CAKUT patients identified 32 rare, predicted deleterious variants. Replication in unrelated families from a national multicenter database prioritized four novel candidate genes--DOCK11, MIB1, TENM2, and TNS1. These candidates are involved in both well-characterized developmental pathways and more under-explored biological processes relevant to renal and ureteric morphogenesis. CRISPR-Cas9-mediated zebrafish knockout studies were employed to validate the potential association of these genes with kidney abnormalities including significant pericardial edema, malformed renal tubules, and impaired glomerular filtration. These findings offer potential genetic diagnoses for 10% of CAKUT probands, and demonstrate that exome reanalysis can substantially improve diagnostic yield and inform personalized clinical management. Overall, this study expands the known genetic landscape of CAKUT.

Gain-of-function (GoF) missense variants in the Two-Pore Domain (K2P) K+ channel TASK-1 (KCNK3) result in Developmental Delay with Sleep Apnea (DDSA), a neurodevelopmental channelopathy, whilst loss-of-function (LoF) variants cause a hypertensive disorder. However, for the related TASK-3 channel (KCNK9), both LoF and GoF variants underlie a distinct neurodevelopmental disorder, KCNK9 Imprinting Syndrome (KIS). The relationship between genotype and phenotype in these disorders is further complicated because TASK-1 and TASK-3 can co-assemble into heteromeric channels with distinct functional properties. Here we report additional probands with missense variants in KCNK3 and KCNK9 and investigate the effect of four novel variants on the functional properties of both homomeric and heteromeric TASK channels. Interestingly, two of these new GoF variants (R131H and L122V) are found in both TASK-1 and TASK-3 and have equivalent functional effects on heteromeric TASK-1/TASK-3 channels, yet result in different clinical phenotypes. We have also determined a cryoEM structure for the pathogenic L122V mutant TASK-3 channel which suggests its dramatic functional effect is likely due to subtle changes in gating and permeation within the inner cavity. Overall, these results highlight the dominant role that homomeric TASK channels play in defining their associated channelopathies as well as the complexity of interpreting K+ channel dysfunction in pathophysiology.

The genetic relationship between asthma and lung function may be dependent on age-at-onset (AAO) of asthma. We investigated whether the shared genetics between asthma AAO and lung function is dependent on AAO. Asthma cases from UK Biobank were subset according to their AAO and genetic correlation was used to obtain genetically homogeneous groups, i.e., [&le;]20 (LT20), 20-40, and >40 (GT40) years. Association analysis and fine-mapping were performed to identify shared genetics between AAO groups and lung function. Mediation and quantitative trait locus (QTL) analyses were performed to identify mechanisms underlying shared genetic associations. Chr5, chr6, chr12, and chr17 each had one region that displayed a cross-phenotype replicated association with at least one AAO group and lung function. Overlapping credible sets obtained from fine-mapping were observed on chr5 and chr6. Mediation analyses demonstrated that for each region the proportion mediated through asthma on lung function was larger for asthma LT20 compared to 20-40 and GT40 suggesting that their effects on lung function were more strongly driven by this association. Tissue-specific QTL analysis revealed shared etiology on chr5 may be acting through SLC22A5 and C5orf56 which might play an important role in decreased lung function among individuals with earlier-onset asthma.

PurposeTo identify genetic variants associated with glucocorticoid (GC)-induced intraocular pressure (IOP) change using genome-wide association study (GWAS) and whole exome sequencing (WES) analyses. Methods530 participants from the Fluocinolone Acetonide in Diabetic Macular Edema (FAME) trials were analyzed, with replication performed in an independent cohort of 588 participants from the Mass Eye and Ear/Retina Health Center (MEE/RHC). All participants were exposed to GC, primarily via intravitreal injection. IOP was measured at baseline and serially within 6 months following GC exposure. GWAS and rare variant gene burden analyses were applied, adjusting for covariates. ResultsGenetic associations for maximal IOP change within 6 months after GC exposure were evaluated. For the primary outcome across all ancestries in FAME, one variant, rs13425173 within the UBE2E3 locus reached genome-wide significance (P=2.88 X 10-8). In the FAME and MEE/RHC meta-analysis, variant rs1040227, also in the UBE2E3 locus, was significantly associated at the genome-wide level (P=2.88 X 10-8) and showed nominal significance in the MEE/RHC cohort (P=0.02). In the colocalization analyses, the significant FAME GWAS UBE2E3 locus was linked to expression regulation of this gene in six tissues including artery aorta. In gene-level analysis, UBE2E3 also demonstrated subthreshold significance (P=6 X 10-6). 532 FAME and 586 MEE/RHC participants were included in the WES gene burden analysis. One gene, MSTO1, passed false discovery rate correction for the primary outcome in FAME. ConclusionWe have identified genome-wide significant common variants associated with GC-IOP change, as well as genes and rare variants that may influence GC-induced IOP change.

Rhabdomyolysis is the acute breakdown of skeletal muscle resulting from failure of cellular homeostasis in response to metabolic stress. Recurrent forms are frequently linked to inherited defects affecting energy metabolism or calcium handling. Ryanodine receptor type 3 (RyR3) is an intracellular calcium release channel, expressed in skeletal muscle, that contributes to the fine-tuning of calcium signaling. Although variants in other calcium-handling proteins have been implicated in rhabdomyolysis, the role of RyR3 has not been established. In this study, we report rare compound heterozygous missense variants in RYR3 identified in two unrelated individuals with severe, fever-triggered recurrent rhabdomyolysis. Muscle biopsies revealed mild structural changes with triadic disorganization, mitochondrial alterations, lipid accumulation, and autophagic material, while overall muscle architecture was largely preserved. Structural modeling supports the pathogenicity of the variants, and calcium flux analysis demonstrated significantly reduced ryanodine receptor-mediated calcium release in patient-derived myoblasts. Functional analyses showed that RyR3 deficiency impaired starvation-induced autophagy, characterized by defective autophagosome formation and reduced autophagic flux, and increased susceptibility to metabolic stress. Mitochondrial bioenergetic profiling revealed reduced oxidative phosphorylation capacity and decreased membrane potential under stress conditions, consistent with compromised mitochondrial adaptation. In zebrafish, ryr3 knockdown resulted in structural and functional muscle abnormalities, including reduced myotome area and decreased locomotor activity, associated with impaired autophagic flux. This study establishes a novel association between recessive RYR3 variants and recurrent rhabdomyolysis and identifies RyR3 as a critical regulator of skeletal muscle stress adaptation through calcium-dependent control of autophagy and mitochondrial homeostasis. More broadly, our findings further highlight autophagy as a central determinant of muscle resilience in the context of rhabdomyolysis.

Individuals with congenital heart disease (CHD) have an increased risk of neurodevelopmental disorders (NDDs), a relationship likely driven by multiple interacting factors, including shared genetic influences, the underlying cardiac malformation, and risks associated with medical and surgical care. To investigate the genetic component of this association, we analyzed genome-wide association study (GWAS) summary statistics for CHD from the UK Biobank and FinnGen, focusing on two phenotypes present in both cohorts: congenital malformations of cardiac septa (Q21) and congenital malformations of great arteries (Q25). A fixed-effect meta-analysis was performed using METAL to increase statistical power. Neurodevelopmental phenotypes were represented by Austism Spectrum Disorder (ASD) and Attention-deficit/hyperactivity disorder (ADHD) GWAS results from the Psychiatric Genomics Consortium. Genome-wide genetic correlations were estimated using the High-Definition Likelihood. Local genetic correlations were assessed using LAVA. Multiple testing was controlled using Bonferroni correction. No significant genome-wide genetic correlations were observed between CHD phenotypes and either ASD or ADHD. However, LAVA identified a single significant local genetic correlation between cardiac septal defects and ASD on chromosome 6. Within this block, two loci near GMDS showed significant association with cardiac septal malformations. Bayesian colocalization did not reveal a variant with strong evidence for a shared causal signal, though three GMDS variants demonstrated moderate support for colocalization. These findings suggest that while global genetic overlap between CHD and NDDs is limited, specific loci, such as the GMDS region on chromosome 6, may contribute to shared developmental pathways underlying both phenotypes.

Glaucoma is the leading cause of irreversible blindness; vision loss is preventable with timely treatment, but early detection is challenging, leaving [~]50% undiagnosed, highlighting the need for improved risk assessment tools. We developed a polygenic risk score (PRS) using data from >6 million individuals. PRS performance was exceptional in European ancestries; top 10% PRS individuals had 10-fold increased risk (OR=10.0) relative to the remainder. Performance remained good across all major ancestry groups; high-PRS individuals were at high absolute risk, especially Africans. High-risk individuals (top 10% PRS) developed glaucoma up to 25 years earlier than those in the lowest 10% and were at 100 times the risk. The PRS also predicted need for treatment escalation in early glaucoma and both prevalent and incident surgery. Risk profiling with this PRS which is clinically available, enables earlier identification and more timely treatment of high-risk individuals for preventable vision loss, with a reduced screening and monitoring burden for those at low-risk. Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSGlaucoma is the leading cause of irreversible blindness worldwide. Early detection is essential because current treatments cannot restore lost vision. Population-wide screening using conventional risk factors for glaucoma (such as elevated intraocular pressure) is not currently cost-effective, though a better stratification method could improve early diagnosis rates and reduce over-monitoring of lower risk cases. Glaucoma is one of the most heritable common complex diseases, suggesting that genetics-based approaches could revolutionize risk stratification. Previous polygenic risk scores (PRS) indexing glaucoma genetic risk were limited by restricted applicability outside European ancestry populations, small discovery sample sizes, modest predictive power, poor clinical availability. The inability to cover a broad range of ancestries limited the practical clinical application of earlier versions of glaucoma PRS for risk stratification and patient management. Added value of this studyA novel PRS, trained on >6 million individuals, has significantly improved glaucoma risk prediction across major ancestry groups, identifying high-risk individuals (top 10% of risk versus remainder) with odds ratios up to 10 in Europeans and 3.4-7.5 in non-Europeans (larger than for any other common complex disease). The excellent PRS risk stratification remained after adjusting for intraocular pressure; adding PRS to a prediction model with age, sex and intraocular pressure increased the Area Under the Curve for predicting glaucoma status from 0.63 to 0.82. High-risk individuals (top 10% PRS) developed glaucoma up to 25 years earlier than those in the lowest 10% and were at 100 times greater risk. The PRS also predicted structural progression, the need for treatment initiation and escalation in early glaucoma, both prevalent and incident incisional surgery for glaucoma, and the likelihood of glaucoma being present in first-degree relatives. These improvements impart strong clinical utility for population risk stratification, and the personalised management of individuals with clinical features suggestive of early stage glaucoma. Implications of all the available evidenceThis clinically available, enhanced PRS enables accurate identification of both high- and low-risk individuals across diverse populations, supporting earlier diagnosis, targeted monitoring, and timely treatment. It provides a clinical tool which can be applied for precision prevention and management of glaucoma, enabling cost-effective, equitable, genetically-informed glaucoma risk stratification across the entire population.

Several lines of evidence suggest that normal-range facial features and nonsyndromic orofacial clefts (OFCs) exhibit a shared genetic basis. Approaches designed to leverage this relationship hold the possibility of revealing new OFC risk loci by boosting discovery power. To test this idea, we applied a pleiotropy-informed GWAS method (cFDR-GWAS) with summary statistics from large, independent European GWASs of normal facial shape (n=4,680; n=3,566) and nonsyndromic cleft lip with or without cleft palate (nsCL/P, n=3,969). The cFDR approach identified 21 independent genomic loci significantly associated with nsCL/P, providing further evidence of the interconnected genetic architecture between these traits. The five original nsCL/P GWAS signals were detected and joined by nine additional loci previously implicated in other OFC association studies. The remaining seven loci represent new nsCL/P genomic regions, and three of these replicated (P < 0.05) in an independent nsCL/P cohort: ASPSCR1, MSX2, and RALYL. A relaxed 10% cFDR-GWAS threshold identified 15 more independent loci with comparable effect sizes to those detected at the strict 5% threshold, two of which replicated: FHOD3 and SMARCA2. Gene expression patterns in major cell types and spatial transcriptomics data highlighted our gene candidates roles in craniofacial development. In conclusion, the application of an empirical Bayesian strategy to draw on association signals from genetically related traits can boost the power to identify and prioritize OFC risk loci missed by agnostic gene mapping approaches. These results hold promise that the cFDR-GWAS approach may be able to enhance our understanding of the genetic architecture of other structural birth defects.

Leber congenital amaurosis (LCA) and Early-onset severe retinal dystrophy (EOSRD) manifest within the first months and the first years of life, respectively. They are the leading cause of severe vision impairment in childhood. Using next generation sequencing, we identified eight families of patients with LCA/EOSRD carrying biallelic combination of six germline variants in DDX41, encoding a DEAD-box ATPase RNA helicase involved in RNA splicing, innate immunity and hematopoiesis. In fibroblasts from a patient carrying the homozygous missense variant c.1187T>C (p. Ile396Thr) and in the retina of Ddx41I396T/I396T mice, DDX41 protein expression was decreased. Electroretinogram recordings in these animals also revealed significant visual dysfunction since the first month of age, supporting a pathogenic role of DDX41 in retinal physiology. Immunohistochemical staining showed that the protein localized to nuclei in all major retinal cell types and to photoreceptor synapses, while biochemical assays showed that LCA/EOSRD variants disrupt DDX41 interactions with RNA through misfolding or the formation of non-productive aggregates, resulting in loss-of-function. Transcriptomic profiling of mutant mouse retinas revealed dysregulation of gene networks associated with Muller cells (MCs), glial cells essential for maintaining retinal structure, metabolic balance, and immune surveillance. The dysregulated pathways chiefly involved cell morphogenesis and junction formation, consistent with immunohistological analyses of widespread architectural disruption and nuclear disorganization, identifying MCs as a site of dysfunction. Together, these findings establish for the first time the involvement of DDX41 in LCA/EOSRD and provide new insights into the role of helicases in retinal homeostasis.

Low-frequency variants (LFVs), defined by minor allele frequencies (MAF) of 1-5%, occupy the gap between common and rare variants in both frequency and effect size. The conventional genome-wide association study (GWAS) significance threshold (5x10-) is overly conservative for LFVs, which account for more than 25% of variants in GWAS. This limitation may obscure meaningful associations in highly heritable yet genetically complex disorders such as autism spectrum disorder (ASD). We hypothesize that the scarcity of significant LFVs in ASD GWAS reflects statistical constraints rather than a true lack of association. To address this, we derived a MAF-specific genome-wide significance threshold using linkage disequilibrium-informed simulations applied to ASD GWAS summary statistics, identifying 2.03x10- as optimal. Applying this threshold revealed three novel LFVs mapping to zinc finger proteins (ZNF420, ZNF781) and known ASD-related genes (KMT2E, PRKDC, MCM4). Enrichment analyses suggested their function in nervous system development and gene regulation. Our findings highlight the contribution of LFVs to ASD risk and underscore the importance of frequency-aware association strategies.

Background/ObjectivesCoronary artery disease (CAD) remains the leading cause of mortality worldwide, with South Asia bearing a disproportionately high and rising burden, particularly at younger ages. This pilot study aimed to investigate genetic variants associated with premature coronary artery disease (PCAD) using whole genome sequencing (WGS). MethodsWGS was conducted on 12 people (5 PCAD cases, 7 matched controls) to assess feasibility and methodology for future large-scale research. High-quality genomic DNA was sequenced at a minimum read depth of 10x with a quality threshold of Q30. Variant calling with stringent quality control identified single nucleotide polymorphisms (SNPs), followed by annotation against gnomAD for allele frequencies and ClinVar for pathogenicity. Protein-coding variants were filtered, and candidate genes were prioritized for comparative analysis between cases and controls. ResultsAn average of over 8.8 million SNPs per individual was identified, with comparable overall variant distributions between cases and controls. Initial analyses revealed 120 SNPs exclusively present in PCAD cases. All protein-coding variants were rare (allele frequency <0.0001), and none were previously classified as pathogenic in ClinVar. After filtration, 87 candidate genes were prioritized. Enriched or unique variants in PCAD cases are mapped to genes involved in lipid metabolism, endothelial dysfunction, inflammatory signaling, immune regulation, thrombosis, vascular remodeling, and metabolic processes. Additional variants were identified in genes related to smooth muscle proliferation, oxidative stress, and other biological pathways. ConclusionsThis WGS pilot study provides an initial overview of the genomic landscape of PCAD in a South Asian cohort, highlighting potential rare variants across multiple biological pathways implicated in atherosclerosis that needs validation in a large-scale study. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/25342163v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1429c57org.highwire.dtl.DTLVardef@264dd8org.highwire.dtl.DTLVardef@c0c6f6org.highwire.dtl.DTLVardef@46752d_HPS_FORMAT_FIGEXP M_FIG C_FIG