Human Molecular Genetics

Fragile X-associated tremor/ataxia syndrome (FXTAS) is an adult-onset neurodegenerative disease associated with carriers of premutation (PM) alleles of the fragile X messenger ribonucleoprotein 1 (FMR1) gene. Expanded CGG repeats in the PM alleles are sufficient to cause cellular stress and toxicity in animal and cellular models of FXTAS. Here, we show that in a Drosophila transgenic model of FXTAS, expanded CGG repeats result in the overexpression of heat shock RNA omega-nuclear (Hsr{omega}-n), a stress-related long non-coding RNA (lncRNA) that dominantly enhances expanded CGG-induced neurotoxicity, whereas its reduction suppresses CGG-induced neurotoxicity. Furthermore, overexpression of Hsr{omega}-n lncRNA concomitantly resulted in a significant enhancement in its association with Hrb87F, the fly ortholog of one of the previously identified CGG repeat RNA-binding proteins, hnRNP A2/B1, and with the imitation switch (ISWI) protein, a chromatin remodeling factor. We extended the findings of Hsr{omega}-n to show that the mammalian stress-related lncRNAs SatIII and Neat I are elevated in cells expressing expanded CGG repeats. Together, these findings highlight the critical roles of stress-related lncRNAs in CGG repeat-mediated toxicity and support a model in which the total steady-state levels of stress-related lncRNAs increase in the presence of expanded CGG repeats as part of the normal regulatory response to combat stress. However, the effects of chronic expanded CGG repeat expression in post-mitotic neurons may result in altered distribution and/or sequestration of specific RNA-binding proteins or chromatin remodeling factors associated with stress-related lncRNAs. This could contribute to the altered cellular homeostasis and genomic instability associated with FXTAS. SignificanceIndividuals carrying the FMR1 allele with expanded CGG repeats (55-200) in the 5' untranslated region are predisposed to fragile X-associated tremor/ataxia syndrome (FXTAS), a late-onset neurodegenerative disorder. FXTAS demonstrates incomplete penetrance, suggesting the possible involvement of genetic modifiers in its pathogenesis. We used a transgenic Drosophila FXTAS model to conduct genetic, molecular, and biochemical analyses. Our findings revealed that the expression of expanded CGG repeats aberrantly activates stress-related Hsr{omega}-n lncRNA in Drosophila melanogaster. Overexpression of this lncRNA exacerbates CGG-mediated neurodegeneration, whereas its repression alleviates this effect. Extending these findings to mammalian systems, we observed the upregulation of SATIII and Neat I, functional homologs of Hsr{omega}-n lncRNA, in cells expressing expanded CGG repeats. Collectively, these findings identify stress-related lncRNAs as key mediators of CGG-mediated toxicity and underscore their potential as therapeutic targets for FXTAS.

Children with Down syndrome (DS) experience delays in cognitive, physical, and motor development. Overexpression of Dual-specificity tyrosine phosphorylation-regulated kinase-1A (DYRK1A), a gene on human chromosome 21 (Hsa21) and triplicated in individuals with Trisomy 21, contributes to neurodevelopmental delays associated with DS, and is a candidate for therapies to improve neurodevelopmental phenotypes. Male and female Ts65Dn DS model pups are trisomic for [~]100 Hsa21 orthologs including Dyrk1a, and both sexes show significant DYRK1A overexpression on postnatal day 6 (P6) in the hippocampus, cerebral cortex, and cerebellum. This study tested the hypothesis that normalization of Dyrk1a copy number in Ts65Dn pups prior to P6 would diminish physical and behavioral developmental outcomes in Ts65Dn mice, thus providing a standard of comparison for success of interventions targeting Dyrk1a. At P3-P21, Ts65Dn compared to euploid pups showed sex-specific deficits in physical, motor, and behavioral development. Male Ts,Dyrk1a+/+/Dox-Cre mice showed improved emergence to running on P19, and both sexes of Ts,Dyrk1a+/+/Dox-Cre mice exhibited reduced isolation-induced ultrasonic vocalizations during the second postnatal week. Dyrk1a normalization in Ts65Dn pups did not improve all abnormal phenotypes, perhaps because of developmental dysregulation between Dyrk1a RNA and DYRK1A protein levels, involvement of other trisomic genes, or improvements in only adult mice.

Objective: Antipsychotic medications are a cornerstone in the treatment of many psychiatric disorders, but they are associated with adverse metabolic effects including weight gain and hypercholesterolemia. To investigate the underlying genetic architecture of these effects, we conducted the largest-by-far and most ethnically diverse genome-wide association study (GWAS) of longitudinal changes in total cholesterol (TC) and body mass index (BMI) using an antipsychotic-treated cohort from the Million Veteran Program (MVP). Methods: The study included 59,372 participants for TC and 39,112 for BMI across European (EUR), African American (AFR), and Hispanic (HIS) ancestries. GWAS, trans-ancestry meta-analysis, functional annotation, and summary-data-based Mendelian randomization (SMR) analyses were performed to identify associated loci, enriched biological pathways, and gene expression signals. Results: We identified genome-wide significant and suggestive loci for both traits, with stronger associations for TC and clinically significant weight gain (BMI > 1.5 kg/m2) than for overall BMI change. Significant loci included genes involved in cholesterol metabolism and lipid homeostasis for TC and genes previously implicated in BMI-related traits for BMI > 1.5 kg/m2. Trans-ancestry meta-analysis highlighted suggestive loci shared across ancestries, and functional annotation demonstrated significant enrichment of protein homeostasis and synaptic function gene sets for BMI > 1.5 kg/m2. SMR analyses further identified suggestive gene expression associations in whole blood and liver. Conclusions: These findings implicate lipid metabolism, body weight regulation, and central nervous system mechanisms in antipsychotic-associated metabolic changes. This work advances understanding of genetic susceptibility to metabolic adverse effects of antipsychotic treatment and may inform future precision medicine approaches to risk prediction and treatment selection.

Consanguinity increases the risk of autosomal recessive disorders and may result in the co-segregation of multiple pathogenic variants within the same family. Although most affected families are explained by a single genetic diagnosis, multilocus pathogenic variation can produce complex and overlapping clinical phenotypes. We investigated a consanguineous Pakistani family with three affected siblings, including dizygotic twins presenting with neurodevelopmental disorder and hearing loss, using detailed clinical evaluation, long-read whole-genome sequencing, bulk transcriptomics, protein profiling, and segregation analysis to determine the underlying molecular diagnoses. One sibling presented with isolated non-syndromic hearing loss, whereas the dizygotic twins exhibited severe neurodevelopmental impairment characterized by global developmental delay, spastic quadriplegic cerebral palsy, microcephaly, and white matter abnormalities. Long-read whole-genome sequencing identified a homozygous start-loss variant in HPDL (c.3G>C) in both twins, consistent with HPDL-related neurodevelopmental disorder with progressive spasticity and brain white matter abnormalities (NEDSWMA). In addition, a novel homozygous nonsense variant in EPS8 (c.1294C>T) was identified in one twin and the sibling with isolated hearing loss, explaining the auditory phenotype. Long-read transcriptomic analysis demonstrated absence of detectable EPS8 transcripts in both individuals homozygous for the nonsense variant, providing transcript-level evidence consistent with a loss-of-function mechanism. Genome-wide comprehensive proteomic profiling (SomaScan) identified distinct protein abundance profiles across family members, with the most pronounced alterations observed in the twins affected by HPDL-related neurodevelopmental disease, particularly the individual harboring pathogenic variants in both EPS8 and HPDL. This study expands the mutational spectrum of EPS8 and highlights the independent segregation of two autosomal recessive disorders within the complex consanguineous family, resulting in distinct and blended phenotypes.

Germline mutations in PTPN11 cause Noonan syndrome (NS) and NS with multiple lentigines (NSML), yet how specific variants drive divergent clinical outcomes through distinct signaling and developmental mechanisms remains unclear. We find that germline and somatic mutations converge on N-SH2 and PTP domains but diverge at residue-level hotspots, reflecting distinct selective pressures. Clinical stratification of 18 pediatric patients reveals four distinct phenotypic classes including (i) the NSML-associated c.1403C>T (T468M) variant, characterized by lentigines, moderate growth impairment, and distinctive facial features; (ii) variants including the VUS c.1282G>A (V428M) and c.1432A>G (I478V), which were associated with cognitive deficits and variable growth impairment; (iii) c.1471C>A (P491T) and c.1472C>T (P491L), predominantly affecting cardiac and growth phenotypes with limited neurocognitive features; and (iv) a severe, multisystem class comprising c.172A>G (N58D), c.178G>A (G60S), c.844A>G (I282V), c.922A>G (N308D), and c.923A>G (N308S), spanning cardiac, growth, cognitive, and craniofacial abnormalities. Biochemical profiling in HEK293T cells revealed that PTPN11 variants stratify beyond simple gain/loss-of-function dichotomies into strong ERK-dependent hyperactivation, moderate ERK activation with variable protein stability and the paradoxical c.1282G>A variant, which did not increase ERK phosphorylation. In vivo, this variant drove excessive neural crest cell migration in chick embryos, suggesting that its effects on NCC migration may involve ERK-independent mechanisms or context-dependent signaling not captured by steady-state assays. ERK activation did not strictly correlate with clinical severity, yet these functional differences were associated with distinct growth, cardiac, pigmentation, and neurodevelopmental outcomes. Our data suggest lineage-specific sensitivity to SHP2 dosage, with dorsal root ganglia neurons appearing more vulnerable to reduced SHP2 stability than melanocyte precursors. Although direct correlations between specific signaling defects and individual clinical features remain complex, our findings provide a refined framework for PTPN11 variant classification, and reveal unexpected SHP2 functions in neural crest development.

Trimethylamine n-oxide (TMAO) is a plasma metabolite linked to adverse cardiometabolic health with complex regulation involving diet, sex, and host genetics. We explored the role of these factors in the genetic regulation of TMAO by performing a primary-level meta-analysis in 1,482 female and male Diversity Outbred (DO) mice from five distinct studies conducted in various regions of the United States. We identified a quantitative trait locus (QTL) associated with TMAO concentration at [~]86 megabase pairs on mouse chromosome 12 with a highly significant LOD score of 67.67. Alleles at the chromosome 12 QTL inherited from the Cast/EiJ (CAST) and PWK/PhJ (PWK) mouse strains primarily drove the association with reduced TMAO concentrations. The chromosome 12 QTL remained significant in sex-stratified analyses and the mode of inheritance appeared additive; furthermore, the QTL was regulated by sex-by-genotype and sex-by-diet interactions. Using a CAST/EiJ X C57BL/6J F2 cross, positional candidates were prioritized by eQTL analysis. Further analysis in a study utilizing the eight DO founding strains identified that Acyp1 was differentially expressed in hepatic tissue from CAST mice, prompting investigation into its genetic regulation. Acyp1 demonstrated relevant cis- and trans-regulation and was significantly correlated with TMAO and hepatic Fmo3. However, no significant relationships between Acyp1 and TMAO were identified in mice inactivated for Acyp1 or with AAV overexpression of Acyp1 in the liver. Genes within the chromosome 12 QTL have synteny with humans and may translate to the genetic regulation of human plasma TMAO concentrations and atherosclerosis. Author SummaryWe explored the roles of diet, sex, and genetics on the regulation of fasting plasma trimethylamine n-oxide (TMAO) concentration by performing a meta-analysis in 1,482 female and male Diversity Outbred (DO) mice from five unique studies. We identified a QTL associated with TMAO concentration on chromosome 12 at [~]86 mega base pair (Mb) with a highly significant LOD score of 67.67. The locus is modified by both sex and diet.

Pathogenic variants in MAPK8IP3 (JIP3) cause Neurodevelopmental Disorder with or without variable Brain Abnormalities (NEDBA), characterized by cognitive impairment, global developmental delay, motor dysfunction, abnormal muscle tone, behavioral dysregulation including autism and attention deficit hyperactivity disorder (ADHD), seizures, and structural brain abnormalities. While more than 30 largely de novo MAPK8IP3 variants have been reported, the functional impact of variants across JIP3 protein structural domains is poorly defined. To address this knowledge gap, we compare clinical features of individuals with a truncating (p.E27X) or one of two missense (p.R578C and p.R1146C) variants from distinct JIP3 functional protein domains to corresponding knock-in mouse models. Our findings showed that all individuals, regardless of variant type, exhibited delays in language and gross motor function, but individual variants were associated with distinct motor, cognitive, and psychiatric symptoms. Corresponding homozygous p.E27X and p.R1147C variant mice resulted in embryonic lethality, consistent with essential roles for JIP3 in early neurodevelopment. Behavioral characterization of viable heterozygous mice revealed variant-specific locomotor, motor coordination, and hindlimb clasping defects that closely recapitulate clinical observations. Heterozygous p.R579C mice exhibited reduced locomotion, impaired motor performance, and hypertonia-like clasping, mirroring human motor deficits. In contrast, p.R1147C mice displayed hyperactivity, hindlimb clasping, and decreased brain weight, paralleling human clinical features. Together, our findings demonstrate that while distinct MAPK8IP3 variants lead to some shared phenotypes, they are also associated with distinct phenotypes that could reveal domain-specific aspects of JIP3 function. This work establishes the first domain-resolved in vivo rodent models of NEDBA and provides a validated translational platform for mechanistic investigation and preclinical therapeutic testing.

Bryant-Li-Bhoj Syndrome (BLBS; OMIM: 619720, 619721) is a Mendelian neurogenetic condition, first described in 2020, with a mixed neurodevelopmental/neurodegenerative phenotype and variable systemic features. To date, 100 affected individuals with 74 unique causative variants have been published. Clinical data and prior functional work in multiple model systems have emphasized the utility of interrogating the pathogenesis of multiple causal variants to identify a convergent, therapeutically targetable mechanism. Additionally, the ability to evaluate the efficacy of future therapeutics relies on the availability of a robustly validated preclinical model. Here, we characterize the developmental and neurobehavioral phenotypes of a novel BLBS mouse model harboring one of the most recurrent causative variants (h3-3a p.T45I). H3.3T45I mice recapitulate the BLBS natural history: perinatal growth restriction, delayed developmental milestones, and progressive motor and gait impairments. Adult mice additionally display craniofacial differences, impaired nest building, hyperactivity in a social context, and male-specific elevated aggression. The non-invasive, clinically translatable endpoints established here provide a validated preclinical platform for evaluating therapeutics for a community whose current standard of care is symptom management. Summary StatementA new mouse model mirrors the developmental delays, motor decline, and behavioral changes seen in individuals with this rare, progressive genetic brain disorder, providing a foundation for testing future therapies.

Defects in lysosomal lipid handling provoke fatal disorders presenting neurovisceral symptoms with variable onset and life spans. A prime example is Niemann-Pick type C disease (NPCD), where export of cholesterol and other lipids from the endosomal-lysosomal system is impaired due to variants of either NPC intracellular cholesterol transporter 1 (NPC1) or NPC intracellular cholesterol transporter 2 (NPC2). Therapeutic options for NPCD are limited to palliative care and disease-modifying drugs, and there is an unmet need for new treatments. Based on positive effects in patient-derived fibroblasts in vitro, we explored how inhibition of bromodomain and extra-terminal domain (BET) proteins affects a well-established mouse model bearing the frequent I1061T variant of NPC1. Treatment with JQ1, a hydrophobic prototype BET protein inhibitor, induced beneficial but sex-dependent molecular and behavioral changes in mice. Our results indicate bromodomain proteins as therapeutic drug target for NPCD and reveal sex-dependent BET protein signaling in mice.

Orofacial clefts (OFCs), including cleft lip (CL), cleft palate (CP), and cleft lip with cleft palate (CLP), are among the most common craniofacial malformations in humans, with a birth prevalence of approximately 1 in 1,000 live births globally. Non-syndromic forms of OFC are predominantly genetic, with significant variability in prevalence across populations. Understanding the genetic underpinnings of OFCs remains a key public health priority, given the substantial medical and societal burden of these conditions. Recent genome-wide association studies (GWAS) have implicated numerous genetic loci, but challenges remain due to genetic heterogeneity and complex gene-environment interactions. This study aimed to identify sex-specific genetic risk factors for cleft lip with or without cleft palate (CL/P) through a meta-analysis of whole genome sequencing (WGS) data from 1,922 case-parent trios across eight diverse cohorts. Our approach revealed four SNPs in three distinct regions that showed genome-wide significant sex-specific effects. However, despite each of these SNPs passing standard quality control filters, follow-up analyses showed that these signals most likely were technical artifacts caused by sequencing errors, in particular mis-mapped reads due to sequence similarities with the sex chromosomes. These findings highlight the necessity for careful scrutiny when studying differences between the sexes in genetic association studies.

Spinocerebellar ataxia type 3 (SCA3) is an inherited, fatal neurodegenerative disease caused by a pathological CAG repeat expansion in the ATXN3 gene, resulting in the selective degeneration of vulnerable neuronal populations. Recent work has identified impairments in oligodendrocyte maturation as a novel and robust feature of SCA3 pathogenesis. Oligodendrocytes synthesize myelin structural components through the endoplasmic reticulum (ER), rendering this organelle essential for white matter integrity. Despite this, the role of ER function in SCA3 remains unclear. In this study, we show that loss of FICD-mediated AMPylation, a post-translational modification regulating the ER-resident HSP70 chaperone, BiP, rescues motor impairments in a transgenic SCA3 mouse model. Ficd-/- SCA3 mice exhibit significantly reduced levels of nuclear ATXN3 in vulnerable brain regions, while Ficd+/+ littermates show an increased burden of AMPylated BiP in the spinal cord, identifying aberrant AMPylation as a novel contributor of SCA3 pathology. Using unbiased proteomics, we demonstrate that Ficd deletion mitigates the pathological decrease in myelin structural proteins and oligodendrocyte maturation factors, restoring levels of mature, myelinating oligodendrocytes. In parallel, we show that Ficd activates SREBP2-dependent cholesterol biosynthesis to support myelination. Taken as a whole, these findings posit ER homeostasis as a critical driver of oligodendrocyte pathology and identify FICD as a novel target for alleviating non-neuronal toxicity in SCA3.

Pathogenic mutations in the ABCA4 gene cause several inherited retinal diseases, particularly Stargardt disease (STGD1). However, many missense variants remain classified as variants of uncertain significance (VUS) due to inconclusive evidence regarding their pathogenic impact. The missense VUS span across all the domains of ABCA4, with the majority found in the larger extracellular domains (ECDs). The largest uncharacterized region of ABCA4 is located in ECD1, where limited structural information and inconsistent computational predictions hinder clinical interpretation of missense VUS in this region. Here, we integrated in silico analysis with in vitro functional assays to evaluate the pathogenicity of VUS in this region and improve their diagnostic classification. Missense VUS in the ECD1 uncharacterized region were curated from ClinVar. Six multiallelic sites were identified in the uncharacterized region and 13 missense VUS on these multiallelic sites were characterized using the integrated analysis. In the in silico platform, the pathogenicity of the VUS were predicted using multiple algorithms, and the structural effects of the variants were analyzed compared to the wild type. Recombinant variants were expressed in virus-like particles (VLPs), and protein expression, membrane localization, and ATPase activity were quantified relative to wild type to identify potential disease-causing variants. From the integrated analysis, variants with pronounced structural destabilization, impaired membrane trafficking, and reduced or absent N-retinylidene-phosphatidylethanolamine (NRPE) substrate stimulated ATPase activities were identified as potentially deleterious. Notably, VUS at p.H193P and p.I214N showed loss of function, with p.I214N reflecting selectively impaired membrane targeting and p.H193P reflecting combined expression and trafficking defects. Additionally, NRPE-stimulated ATPase activities were impaired in VUS, p.V195L, p.V195I, p.D197H, p.I214F and p.N269S. Overall structural destabilization interfered with the NRPE-stimulated ATPase activities of p.N269S, while the lack of NRPE-stimulated ATPase activities of p.D197H, p.V195L, p.V195I and p.I214F are thought to be due to impaired NRPE interactions with ABCA4. All the VUS at p.R140, p.H193Y, p.D197N and p.N269H showed both the basal and NRPE-stimulated ATPase activities but less than that of the wild type, displaying a mild functional deficit. Together, these findings demonstrated that certain VUS within the unresolved ECD1 region disrupt ABCA4 stability and function, supporting their contribution to disease pathogenesis. This integrative approach highlights key residues likely to be pathogenic and advances the interpretation of VUS in inherited retinal disorders.

IntroductionGenetic variants within the APOE-TOMM40 locus are associated with Alzheimers disease (AD). A specific role for TOMM40 is indicated by the finding that 523 poly-T variants are associated with AD risk, but the mechanism for this effect has not been established. Our studies have shown that suppression of Tomm40 in mice increased brain cholesterol content, an AD risk factor, and thus the present study sought to assess whether major 523 poly-T variants (Short [S] and Very Long [VL]) are associated with altered lipid content of brain and other tissues. MethodsWe utilized a mouse model containing the entire human APOE3-TOMM40 locus to quantify cholesterol and triglyceride levels in brain, liver, and white adipose tissue (WAT), as well as brain content of the AD biomarkers A{beta} 42 and tau, in mice carrying two homozygous TOMM40 523 poly-T genotypes (S/S and VL/VL). ResultsMale mice carrying the 523-S/S genotype, but not females, showed higher brain cholesterol and triglyceride levels than VL/VL carriers, together with greater brain A{beta} 42 content. WAT showed similar lipid differences as in the brain, while hepatic lipid content was broadly similar between 523-S/S and -VL/VL genotypes, though there was a trend for higher triglycerides in VL/VL mice in a sex- and age-dependent manner. DiscussionThese results demonstrate that TOMM40 523 poly-T variants drive tissue-specific, sex-, and age-dependent lipid differences in humanized APOE3-TOMM40 mice, with the S/S genotype linked to elevated brain cholesterol and A{beta} 42 levels, effects that link this locus to AD pathogenesis.

Differences in physical features and disease prevalence between men and women are examples of sexual dimorphisms. However, sex differences can manifest not only in trait means but also in how strongly risk factors are linked to diseases (e. g. BMI to cardiovascular disease), a question heavily under-researched. To fill this gap, we set out to identify sex differences in phenotype correlations (rP) and decompose them into genetic (rG) and environmental (rE) contributions. Our analysis revealed 250 trait pairs with significant sex-different phenotypic correlations in the UK Biobank. Overall, we observed a predominance of environmental contributions to sex-different effects: 182 trait pairs (73%) exhibited exclusively sex-different rE, while 68 (27%) showed sex differences in both rE and rG, and no trait pair was affected solely by sex-specific rG. For example, we detected sex-different environmental correlation between C-reactive protein and BMI (rE(men) = 0.07 vs rE(women) = 0.25), but no sex-difference in genetic correlation. On the contrary, glycated haemoglobin and LDL cholesterol showed genetic correlation only in women (rG(women) = 0.17; 95% CI = [0.1, 0.23]), but environmental correlation only in men (rE(men) = -0.18; 95% CI = [-0.19, -0.16]). Some of the observed sex differences - including those involving testosterone, SHBG, urate, waist-hip ratio, and triglycerides - may reflect underlying sex-specific genetic architectures, as evidenced by low between-sex genetic correlations. In conclusion, environmental factors are the predominant contributors to sex differences in phenotypic correlations between complex traits, with modest detectable contributions from sex-specific genetic architectures. Recognising these patterns can inform the development of more effective, sex-informed interventions.

Pathogenic variants in genes involved in transcriptional regulation and RNA processing have emerged as points of functional convergence in neurodevelopmental disorders (NDDs), but their specific disease mechanisms remain unknown. By screening 1,562 Finnish extended families from the Northern Finland Intellectual Disability cohort affected by cognitive impairment, we discovered a family with six affected members carrying a heterozygous loss- of-function variant in INTS6. INTS6 is a conserved member of the phosphatase module of the Integrator complex, which regulates RNA polymerase II activity, with a reported role in the pathogenesis of NDDs. To determine the variants transcriptomic effects, we performed RNA-sequencing of induced pluripotent stem cells (iPSCs) and iPSC-derived neuronal cells from cases and controls, revealing transcriptome-wide splicing defects, with increased intron retention observed in genes involved in translation, cell cycle and RNA processing in variant carriers. CRISPR-Cas9 knock-in iPSCs confirmed that the variant was associated with downregulation of transcription factors and developmental processes in early neuron differentiation. In addition, downregulated genes in variant carrier neurons were enriched for synaptic genes, suggesting effects on neuronal development. These findings highlight the critical role of INTS6 in transcriptional regulation of human neurodevelopment and reinforce its association with NDDs.

CDKL5 Deficiency Disorder (CDD) is a rare developmental epileptic encephalopathy typically caused by loss of function variants in the gene encoding the X-linked serine-threonine kinase CDKL5. CDKL5 is highly expressed in the brain during development, and key neuronal functions of the kinase include cytoskeletal organisation and synaptic stability. However, at present, little is known about the function of astroglia in CDD. Given the importance of these cells in synaptic development and homeostasis, as well as dysfunction in other epileptic diseases, it was hypothesised that astrocytes may contribute to CDD pathology. Induced pluripotent stem cells harbouring a CDKL5 loss-of-function mutation (and isogenic controls) were derived from CDD patient fibroblasts and differentiated into astrocytes (iAstros). Analysis of iAstros revealed transcriptomic, proteomic and functional dysregulation in CDKL5-mutant iAstros relating to water transport and immunological function, including a diminished response to TNFa stimulation. Moreover, iAstros showed increased branching and reduced phosphorylation of the known CDKL5 target end-binding protein 2 (EB2) - indicative of disrupted cytoskeletal regulation in a manner similar to CDKL5-null neurons. Finally, we report the generation of novel in vitro models of CDD. CDKL5 was knocked down in adult and foetal human organotypic brain slices through transduction with an AAV encoding a novel CDKL5 shRNA. Slices transduced with the CDKL5 shRNA displayed increased spontaneous network activity, demonstrating the functionality of this model. Importantly, interrogation of these models revealed dysregulation of key astrocytic proteins congruous with the human glial stem cell model. Consequently, this study describes the generation of novel human models of CDD and their associated astrocytic dysfunction - paving the way for novel discovery and therapeutic intervention.

Genome-wide association studies (GWASs) have disproportionately focused on European (EUR) populations, limiting the characterization of genetic architecture in other ancestries. To address this imbalance, we integrated large-scale biobanks from Japan, Korea, Taiwan, and China to perform the largest phenome-wide meta-analysis to date in East Asian (EAS) populations, encompassing over one million individuals across 127 complex traits. We identified 8,010 previously unreported associations and observed substantial genetic sharing across EAS subpopulations, while also detecting cohort-specific heterogeneity within the broader EAS context. Transethnic analyses revealed moderate genetic correlations between EAS and EUR populations, indicating both shared and ancestry-specific components of disease risk. Pleiotropy analyses highlighted prominent signals within the HLA region, supported by protein-protein interaction connectivity and immune-related pathway enrichment. Decomposition of genome-wide association matrices further uncovered structured cross-trait architectures, revealing a predominantly shared polygenic backbone driven by metabolic, biochemical, and anthropometric traits, together with two discrete latent components enriched for immune-related processes. Together, our findings refine the genetic architecture of complex traits in East Asian populations at unprecedented scale and clarify the balance between shared and population-specific determinants of human diseases.

Neuroblastoma amplified sequence gene (NBAS) variants are associated with short stature, optic atrophy, and Pelger-Huet anomaly (SOPH) syndrome. We previously identified compound heterozygous variants in NBAS to cause atypical Osteogenesis Imperfecta (OI), with these patients presenting with short stature, developmental delay and recurrent long-bone fractures. However, skeletal disease progression due to these variants and the disease mechanisms underlying NBAS-associated OI remain poorly understood. Here, we provide a clinical update on previously identified patients and investigate the role of NBAS during skeletal development using zebrafish knockout and patient-specific missense variant zebrafish models. Homozygous knockout larvae exhibited delayed operculum development, reduced bone ossification, and defects in Meckels cartilage morphology and its underlying cellular structure. Homozygous missense larvae displayed milder cartilage defects without any major defects to early skeletal structures. Seemingly opposing phenotypes were observed in compound heterozygous zebrafish carrying the knockout and missense alleles in trans, with no obvious phenotypes seen in the Meckels cartilage and accelerated operculum development observed. Together, our results demonstrate that different nbas variants differentially affect skeletal development, suggesting complex spectrums of phenotypic and pathogenic mechanisms in NBAS-associated atypical OI.

While substantial research has examined the role of infant nutrition in early brain and cognitive development, the links between later childhood nutrition, brain development, and cognitive and academic skills remain less explored. In this work, we investigated for the first time the direct and indirect associations between nutrition intake, white matter microstructure and structural connectivity, and cognitive and academic outcomes. Using longitudinal data from typically developing children aged 2 to 14 years, we combined neuroimaging, dietary, and cognitive measures to test direct associations between nutrition and brain structure and connectivity, as well as cognitive and learning outcomes, and to assess whether brain development mediated these relationships. We found that specific nutrients, including DHA, sphingomyelin, iron, niacin, choline, and palmitoleic acid (omega-7), were associated with improved brain structure and connectivity, as well as better cognitive and learning outcomes. We also found that brain development partly mediated the association between childhood nutrition and learning outcomes. To our knowledge, this is the first study to show such a pathway in school-age children. These results add to the growing literature demonstrating the ongoing importance of nutrition beyond infancy in supporting childhood brain and cognitive development.

Background and Aims: Residual pulmonary vascular obstruction (RPVO) defined as the persistence of thrombotic material within the pulmonary arteries several months after an acute pulmonary embolism (PE) is associated with an increased risk of severe complications, including recurrent events and chronic pulmonary hypertension. However, the genomic architecture underlying RPVO in unprovoked PE remains poorly understood, and this study aims to address this gap. Method: By leveraging genetic and imaging RPVO data from three independent cohorts totaling 586 unprovoked PE patients, we conducted a meta-analysis of genome wide association study (GWAS) of RPVO using a dedicated statistical method to handle the semi-continuous distribution of RPVO. The meta-GWAS was complemented by haplotype association analyses and transcriptome wide association studies as well as Mendelian Randomization (MR) approaches based on plasma metabolites and proteins. Results: Through meta-GWAS, we identified one locus, OSTN, associated with RPVO (lead variant rs59109356 associated with a ~2-fold increase of RPVO, p=3.92x10-8). A second locus, CCN4, previously reported to associate with pulmonary fibrosis, was also identified, with evidence of association approaching genome-wide significance (p=6.7x10-8). We also identified a common haplotype spanning over AHSG/HRG/KNG1 associated with a ~3-fold increase of RPVO (p=2.96x10-8). Using plasma protein-based MR, we demonstrated that one unit increase in genetically determined plasma levels of IL-1 R AcP encoding IL1RAP was associated with a 28% (p=1.32x10-6) reduction in RPVO. We also observed statistical evidence that the CCN4 (p=0.06) and IL1RAP (p=0.02) loci associate with the risk of PE recurrence in a sample of 1,617 unprovoked PE patients. Conclusions: By identifying novel molecular determinants of RPVO that map to loci involved in inflammatory pathways and vascular remodeling, our study provides evidence that inflammation is the predominant, and likely the key mechanism underlying RPVO, whereas impaired fibrinolysis appears to play a more limited role.