Identification of a novel de novo NONO variants causing X-linked syndromic intellectual developmental disorder-34 in a fetus.

Congenital diaphragmatic hernia (CDH) is a developmental anomaly associated with high mortality and morbidity, primarily attributed to accompanying pulmonary hypoplasia. Genetic factors are crucial in the etiology and pathogenesis of CDH, with various copy number variations (CNVs) and gene sequence variants implicated in this malformation. Previous studies have underscored the importance of retinoic acid (RA) signaling pathways and related genes. Nonetheless, the complexity of diaphragmatic development involving cell migration, cytoskeleton organization, and myogenesis suggests that candidate CDH genes extend beyond the RA pathway. To explore novel candidate gene variants and their roles in CDH, we performed whole exome sequencing (WES) in CDH-affected fetuses. Following the evaluation of chromosome and array-CGH analyses, 17 CDH cases with normal results in our cohort were subjected to WES. Trio-WES was conducted on eight fetuses, while solo-WES was applied to the remaining nine cases. The identified variants were validated and subjected to segregation analysis via Sanger sequencing. Bioinformatic analysis revealed novel potentially pathogenic variants not only in six genes previously known to be associated with CDH (NR2F2, ZFPM2, ARID1A, CREBBP, PLAT, and RARB) but also in nine additional genes (COL11A1, NEIL2, PCSK5, RBM8A, STAB2, SETD5, TAF4, ZBTB38, and ZNF423) that, based on their functions, database entries, and literature, may be considered candidate genes for CDH. Our findings reinforce that no single gene or variant is responsible for the majority of CDH cases, and also demonstrated the effectiveness of WES in identifying novel candidate genes and variants that contribute to CDH etiology.

Background Hearing loss (HL) is genetically and phenotypically heterogeneous. Variants in GRHL2, which encodes the Grainyhead-like 2 transcription factor, cause autosomal dominant nonsyndromic HL (DFNA28). Only six pathogenic GRHL2 variants have been reported, predominantly associated with high-frequency HL. Objectives To elucidate the molecular etiology of ADNSHL presenting with low-to-mid frequency HL in a four-generation Chinese Han family. Material and methods Whole exome sequencing was performed to identify the candidate variant, followed by Sanger sequencing for co-segregation analysis. Functional assessments included protein subcellular localization, luciferase assays and co-immunoprecipitation. Results A novel nonsense variant, c.648C>G (p.Tyr216Ter), in GRHL2 was identified and co-segregated with the HL phenotype. Contrary to the characteristic high-frequency HL in DFNA28, the 11-year-old proband (IV:4) displayed low-to-mid frequency HL. The variant introduces a premature termination codon, producing a truncated protein missing 409 C-terminal amino acids. Initially classified as likely pathogenic according to the ACMG/AMP guidelines, functional analysis demonstrated cytoplasmic mislocalization and abnormal transcriptional upregulation, potentially via interaction with wild-type GRHL2, prompting reclassification to pathogenic. Conclusion and significance Our findings broaden the GRHL2 mutational spectrum, support a gain-of-function mechanism in DFNA28, and establish GRHL2 as a gene associated with low-to-mid frequency HL, expanding known genotype–phenotype correlations.

Despite decades of development in anti-seizure medications, ~30% of individuals remain refractory to all treatments, and none of the existing therapies are disease modifying. Identifying targets outside the current preclinical paradigm is critically important. This study aimed to characterize the landscape of current epilepsy treatments at the level of gene interaction networks and identify novel genetic modifiers of epilepsy as potential novel therapeutic targets. We performed a functional network analysis to score genes based on their interactions with known epilepsy genes, and we integrated these functional scores with population genetics data and drug tractability information. In parallel, we performed a meta-analysis of genome-wide association studies of epilepsy-related phenotypes in genetically diverse mice using a large compendium of historical phenotyping data. Genes within mapped loci were prioritized based on functional rankings, and genomic evolutionary rate profiling (GERP) was used to identify highly single-nucleotide polymorphisms at evolutionarily constrained positions. Functional network analyses of known epilepsy genes revealed a strong involvement of neurodevelopmental processes in epilepsy pathogenesis, which are not targeted by existing or emerging treatments. Meta-analysis of seizure traits in mice identified 118 non-overlapping loci harboring potential seizure phenotype modifiers. Using functional rankings, we prioritized 168 candidate genes within these loci and used GERP scores to filter down to 75 SNPs as candidate variants within these genes. Among them, five genes-Ephb2, En2, Cadps2, Igsf21, and Cep170-contain regulatory variants in evolutionarily constrained sites. Four of these genes are validated as modifiers of neurological traits, including epilepsy susceptibility. This study prioritized epilepsy modifier genes that are strongly predicted to influence neurodevelopmental processes, which are underrepresented among current therapeutic targets. Furthermore, the identified genes represent novel candidate modifiers with potential clinical relevance. Our systems-level analysis offers a novel view into the potential target landscape, pointing toward promising new directions for disease-modifying treatments.

Abstract Background Mutations in the ZMIZ1 gene have been implicated in neurodevelopmental disorder with dysmorphic facies and distal skeletal anomalies (NEDDFSA). However, the underlying cellular and physiological mechanisms remain poorly understood. Methods Exome sequencing was performed to identify candidate variants. qPCR, Western blot, immunofluorescence, CCK-8, and wound-healing assays were employed to assess gene function in human skeletal muscle cells (HSkMCs). RNA-seq and co-immunoprecipitation coupled with mass spectrometry (Co-IP/MS) were used for transcriptomic and interactome profiling. Results Here, we identified a novel de novo missense variant c.910G>C (p.A304P) in ZMIZ1 in a patient with NEDDFSA. The p.A304P variant significantly increased ZMIZ1 mRNA and protein expression levels and altered its subcellular localization. Functional assays demonstrated enhanced proliferation and migration in HSkMCs expressing the mutant ZMIZ1 . qPCR validation revealed significant dysregulation of key genes in the TGF-β1 signaling pathway. Transcriptome analysis identified the cytokine-cytokine receptor interaction pathway as the most significantly enriched pathway. Additionally, we identified a novel interaction between ZMIZ1 and the transcription factor GTF2I. Conclusion Our study identifies a novel likely pathogenic variant in ZMIZ1 associated with NEDDFSA. These findings provide new insights into the cellular and physiological mechanisms underlying NEDDFSA, highlighting ZMIZ1’s role as a regulatory hub in multiple signaling pathways. Impact Our study identifies a novel likely pathogenic variant in the ZMIZ1 gene associated with NEDDFSA, the second in China, and demonstrates its functional impact on cellular processes and signaling pathways, particularly in muscle cells. This finding expands the known ZMIZ1 mutation spectrum and provides the first functional evidence of its role in muscle cells, highlighting its potential as a regulatory hub in multiple pathways beyond the nervous system. These results offer new insights into the cellular mechanisms underlying NEDDFSA, especially in non-neurological tissues, and may facilitate the development of targeted therapies for related disorders.

Only 15% of young-onset breast cancers have identifiable hereditary germline pathogenic variants (PVs) in an established breast cancer susceptibility gene. However, it is believed that a significant proportion of these breast cancers have additional monogenic or rare risk variants that require identification. To uncover novel cancer susceptibility genes, we performed germline whole-exome/genome sequencing of samples from 564 patients with young-onset breast cancer (aged <40 years), as well as samples from 4032 female controls. The identified candidate variants were further genotyped in 6,967 independent breast cancer cases across all age groups. We identified two PVs that were significantly associated with the risk of hormone receptor-negative young-onset breast cancer: POLH p.K589T (OR = 3.65, 95% confidence interval [CI] = 1.28–10.4, P = 0.0095) and RAD51 p.M1fs (OR = 2.15, 95% CI = 1.15–4.02, P = 0.014). When BRCA1/2 PV carriers were excluded from the analysis, only RAD51 p.M1fs retained a significant association. Whole-genome sequencing of tumor samples carrying these germline risk variants revealed that they harbored mutational signatures indicative of a deficiency of homologous recombination. These findings suggest that hereditary POLH p.K589T and RAD51 p.M1fs are candidate variants associated with an increased risk of hormone receptor-negative breast cancer.

PurposeTo identify disease-causing gene variants in 114 probands with congenital cataract (CC), either isolated or accompanied by additional ocular abnormalities and multisystem disorders.MethodsA total of 114 probands with CC who accept cataract surgery between 2021 and 2022 were enrolled. All probands and available family members underwent a comprehensive ophthalmologic examinations and received clinical diagnoses. Medical history and examination data were collected. Whole-exome sequencing (WES) was performed for all probands, and candidate variants were further validated by co-segregation analysis.ResultsAmong 114 probands, 49 presented with isolated CC, and 11 exhibited CC combined with systemic abnormalities such as congenital heart disease (CHD), laryngeal cartilage hypoplasia and developmental delay, the remaining were associated with congenital ocular anomalies including iris hypoplasia, posterior capsular defect, microcornea, and microphthalmia. In total, 49 variants were identified across 19 genes, 23 of which were novel. The detection rate of variants by WES was substantially higher in inherited families (68.75%, 11/16) than in sporadic cases (38.78%, 38/98), with an overall detection rate of 42.98% (49/114).ConclusionsThis study provided a comprehensive characterization of the diverse clinical phenotypes of CC in Chinese cohort and updates the mutational landscape of CC-associated genes. By employing WES, we expand the genetic spectrum and highlight additional genotype-phenotype correlations in CCs.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12920-025-02255-9.

Genetic tests for behavioral and personality traits in dogs are now being marketed to pet owners, but their predictive accuracy has not been validated. To evaluate the reliability of such tests, we analyzed data from Darwin's Ark, a community science initiative that includes over 3,000 dogs with both genetic data and individual-level behavioral phenotypes. None of the candidate variants had significant associations or predictive power for behavioral traits as previously reported. However, we found strong associations with aesthetic traits that differentiate breeds, such as height, leg length, and ear shape. Our results suggest that earlier studies using breed-average phenotypes, rather than individually measured phenotypes, were confounded by population structure. Behavior in dogs is polygenic and complex, and thus cannot be accurately predicted using tests that consider only a few genetic variants. Furthermore, behavior in dogs is only moderately heritable, and environmental influences inherently limit the potential accuracy of genomic predictions. Developing meaningful, accurate genetic predictions for complex traits that can improve dog health and welfare will require very large cohorts of individually phenotyped dogs.

To identify novel pathogenic genes and variants responsible for oocyte degeneration and death. Whole-exome sequencing was conducted in 78 individuals with primary infertility characterized with oocyte degeneration and death, followed by Sanger sequencing of candidate variants. Pathogenicity of identified COX15 variants was characterized through morphological assessment, AlphaFold3-based structural modeling and functional validation including Western blotting and immunofluorescence. We identified a compound-heterozygous COX15 variants and a homozygous COX15 variant in two affected individuals with oocyte degeneration. The variants c.C664T [p.R222C] was a previously reported recurrent pathogenic variant. The clinical records showed that majority of oocytes retrieved from affected individuals were immature and degenerated. Structural modeling assay showed that all the COX15 variants affected the binding pocket with heme o. The compound-heterozygous COX15 variants c.C905T [p.P302L] and c.C664T [p.R222C] in family 1 significantly decreased COX15 protein level in HeLa cells. While the homozygous variant c.C647T [p.P216L] decreased the COX1 protein level in oocytes from family 2 (II-1), implying the disruption of respiratory complex IV (COXIV) assembly. Our study identified two novel variants in COX15, and functional analysis confirmed the pathogenicity of the variants. Our findings expand the mutational spectrum of COX15 and making it a potential genetic diagnostic marker for those suffering from female infertility causing by oocyte degeneration.

IntroductionRubinstein-Taybi syndrome type 2 (RSTS2; OMIM #613684) is a rare autosomal dominant disorder caused by loss-of-function variants in the EP300 gene (OMIM #602700), characterized by intellectual disability, distinctive craniofacial features, and skeletal anomalies.MethodsWhole-exome sequencing (WES) was performed on five pediatric patients presenting with neurodevelopmental delay. Candidate variants were filtered using the TGex platform and validated by Sanger sequencing for familial segregation analysis. The functional impact of variants was assessed using diverse bioinformatic tools, and pathogenicity classifications were assigned according to ACMG/AMP guidelines.ResultsFive novel EP300 variants were identified in this study: c.4774A>G (p.Lys1592Glu), c.4452 + 5G>C, c.3764A>G (p.His1255Arg), c.3591–2A>G, and c.6439C>T (p.Gln2147*). These alterations impair gene function through mechanisms including amino acid substitution, disruption of mRNA splicing, or premature protein truncation. All variants were classified as pathogenic or likely pathogenic per ACMG/AMP criteria. Literature analysis reveals that the predominant clinical manifestations in the Chinese patients encompassed neurodevelopmental impairment, accompanied by motor delay, growth retardation, and microcephaly. Strikingly, archetypal craniofacial dysmorphisms, such as arched eyebrows, long eyelashes, downslanting palpebral fissures, beaked nose, as well as significant skeletal abnormalities were absent, suggesting EP300 variants may present with a broader and more variable phenotypic spectrum than previously recognized.ConclusionThis study reports five novel pathogenic EP300 variants, expanding the variant repertoire of RSTS2 and providing an important basis for clinical diagnosis and genetic counseling.

Mapping genome-wide selective sweeps is of high relevance in cattle population genomics, having successfully identified thousands of genomic regions and candidate genes, with potential to reveal links to agriculturally important traits such as those related to production and adaptation to extreme environments. However, pinpointing the underlying causal variants remains a key priority in understanding molecular mechanisms controlling these traits. The lack of an integrative resource for selective sweeps has impeded meta-analysis and candidate variant prioritization. In the current update of the Bovine Genome Database (BGD; https://bovinegenome.elsiklab.missouri.edu), we address this gap by incorporating a curated dataset consolidating 92 519 selective sweeps identified through 340 genome-wide analyses across 213 cattle populations worldwide. Incorporating this new dataset into BovineMine enables meta-analysis across studies and populations to identify consensus signals, and the exploration of selective sweeps in the context of genes, gene functions, genomic variations, and quantitative trait loci. Furthermore, the BGD JBrowse genome browser enables visualization of sweep regions alongside other genomic features and functional annotations such as histone marks, open chromatin regions, and chromatin states. This BGD update facilitates the prioritization of candidate causal variants and helps identify unanswered questions in disentangling the molecular basis of adaptive and economically important traits in cattle.

ObjectiveAs a critical immune checkpoint, cytotoxic T-lymphocyte-associated protein 4(CTLA-4)deficiency is a well-established cause of inborn errors of immunity. This study characterizes a novel CTLA-4 deletion variant identified in a pediatric case of refractory autoimmune hemolytic anemia (AIHA), with the aim of delineating the clinical profile and elucidating the underlying pathogenic mechanism.MethodsTrio-based whole-exome sequencing (WES) was performed on peripheral blood samples from a 6-year-old female with refractory AIHA and her parents. Candidate variants were validated by Sanger sequencing. Structural modeling of mutant CTLA-4 was conducted, followed by in vitro functional assays in 293T cells to assess mRNA transcription (qPCR) and protein expression (Western blot).ResultsA CTLA-4 (c.362_391del) variant was identified within the immunoglobulin V-set domain of the CTLA-4 protein. In vitro experiments demonstrated significant reductions in both mRNA and protein expression levels caused by this variant.ConclusionThe CTLA-4 (c.362_391del) variant may contribute to refractory AIHA in children. This case highlights the potential necessity of including CTLA-4 variants in the differential diagnosis of pediatric AIHA, particularly when conventional therapies prove ineffective, and warrants further validation in larger cohorts.

To investigate the pathogenicity of splice-site variants in DNAH17, and analyze their impact on sperm morphology and motility, we employed whole-exome sequencing (WES) and Sanger sequencing to identify and validate candidate variants. Computational predictions of splicing defects were performed using varSEAK and MobiDetails. Functional validation was conducted using minigene splicing assays in HEK293T cells. Structural modeling of mutant proteins was performed with AlphaFold3 and visualized by PyMOL. Two novel splice-site variants (DNAH17: c.11677 + 5G > T/c.11677 + 5G > A) were identified in a proband with asthenozoospermia and multiple morphological abnormalities of the sperm flagella (MMAF). Bioinformatics tools predicted disruption of the canonical donor splice site (MaxEntScan score reduction: 54.2% for G > A, 39.5% for G > T; SpliceAI donor loss scores > 0.8). Minigene assays confirmed exon 72 skipping, leading to a frameshift mutation (p.Asn3844Lysfs*13) that truncates the AAA6 domain. This study expands the mutational spectrum of DNAH17-related male infertility by demonstrating that splice-site variants disrupting the AAA6 domain represent a novel pathogenic mechanism underlying asthenozoospermia and MMAF. These findings underscore the necessity of integrating splice-site analysis into genetic diagnostics for male infertility.

Experimental insights into MMACHC variants using a novel minigene system.

Exome and genome sequencing have greatly improved the diagnosis of rare genetic disorders but remain limited in their ability to identify and classify non-coding variants, including intronic variants, cryptic splice-site alterations, and disruptions in regulatory regions. RNA sequencing (RNA-seq) has emerged as a powerful tool to bridge this gap by providing functional insights into genomic variants that disrupt splicing or gene expression, thereby aiding in variant interpretation and classification. We retrospectively reviewed 30 cases from the Utah Penelope Program and the Undiagnosed Diseases Network over a three-year period, in which RNA-seq was performed on whole blood and/or fibroblasts following either negative DNA sequencing or the identification of candidate variants requiring functional assessment. In these cases, RNA-seq identified exon skipping, cryptic splice-site activation, and intron retention, leading to transcript disruption. Additionally, positional enrichment analysis clarified X-inactivation patterns and dosage effects, confirming the pathogenicity of copy number variants. By detecting these transcript-level alterations, RNA-seq provided functional evidence supporting the reclassification of multiple variants of uncertain significance, contributing to diagnostic resolution in selected cases. This study underscores the clinical utility of RNA-seq in detecting splicing and regulatory defects that DNA sequencing and predictive tools alone cannot resolve. Integrating RNA-seq into clinical workflows can support variant classification, aid in diagnostic resolution for selected cases, and provide mechanistic insights into genetic disorders, contributing to patient care and genetic counseling.

Tropheryma whipplei is the bacterial agent responsible for Whipple’s disease (WD). However, WD occurs only very rarely in Tw-infected individuals. We investigated the cause of disease in two relatives with WD. We studied a son and his mother, both presenting an articular form of WD, at the ages of 38 and 60 years, respectively, these episodes occurring five years apart. Both were otherwise healthy. We performed whole-exome sequencing, characterized a candidate variant biochemically, and performed an immunological analysis on the patients’ leukocytes. Both patients were heterozygous for a rare missense variant of the gene encoding the transcription factor IRF4 (p.R25S), for which haploinsufficiency was previously reported to underlie WD in a large multiplex family. This variant was hypomorphic for DNA binding and transcription induction. It did not exert negative dominance. Immunity was otherwise normal in these two patients. Haploinsufficiency for IRF4 can thus underlie WD in T. whipplei-infected individuals from at least two unrelated families.

Background/Objectives: Spinocerebellar ataxia (SCA), or hereditary ataxia, is a progressive neurodegenerative disorder primarily affecting motor control and voluntary muscle coordination due to cerebellar or spinocerebellar dysfunction. While numerous genetic variants have been linked to SCA in both humans and dogs, some cases remain genetically unexplained. This study aimed to describe the clinical and pathological phenotype, and to identify the genetic basis, of an atypical form of SCA observed in a mixed-breed dog presenting with additional clinical signs beyond classic SCA. Methods: Clinical and postmortem examinations were performed to document neurological and systemic pathology. Whole-genome sequencing (WGS) was conducted on the affected dog, and variant filtering was carried out using a control cohort of over 700 unaffected dog genomes to identify candidate variants. Results: In addition to classical SCA features, the affected dog exhibited retinal and optic nerve degeneration and severe, non-regenerative anemia. WGS did not reveal any known SCA-associated variants. Variant filtering identified a novel homozygous 4-base-pair frameshift deletion in CLPX (caseinolytic mitochondrial matrix peptidase chaperone subunit X) [XM_038580726.1:c.1723_1726del; chr30:g.29943285_29943288del]. This variant is predicted to cause a frameshift and premature stop codon within 17 amino acids, truncating approximately 6.64% of the protein. Conclusions: This is the first report associating a CLPX variant with SCA in any species. Given the gene’s high evolutionary conservation and known role in mitochondrial protein homeostasis, this finding may have implications for understanding CLPX-related neurodegeneration and anemia in both veterinary and human medicine.

Background/Objectives: Chronic thromboembolic pulmonary hypertension (CTEPH) is a rare disease, in which multiple genetic and environmental factors may contribute. This study aimed to identify potential genetic determinants in patients with CTEPH and to compare their occurrence to a control group, which included patients with pulmonary embolism who had not developed CTEPH. Methods: Tier 1 and 2 genes related to coagulation, fibrinolysis and platelet disorders—as recommended by the International Society on Thrombosis and Haemostasis—and genes associated with vascular conditions were analyzed in n = 15 patients with CTEPH and n = 17 controls using next-generation sequencing. Non-synonymous, rare variants were collected and interpreted. Results: As expected, no single gene or variant was consistently present among CTEPH patients. Instead, individuals carried different mutations and combinations of variants. We identified several variants that were not found in the control group. Candidate variants were detected in F12, F13A1, F13B, F5, KNG1, SERPIND1, THBD, ADAMTS13, VWF, STIM1, ETV6, THPO, MPL, SERPINA1, ENG, RASA1, ACVRL1, GDF2, NFE2, SOX17 and RNF213. We did not detect exclusive variants in FGA, CPB2, and BMPR2 although they were suggested as candidates in previous studies. Elevated factor VIII and von Willebrand factor in CTEPH could not be explained by mutations in VWF and F8. Conclusions: Our study supports the hypothesis of heterogeneous genetic background in CTEPH, involving multiple pathways such as coagulation, altered fibrinolysis and impaired angiogenesis. These results provide a basis for more detailed investigations into specific genes and variants potentially associated with CTEPH in larger cohorts.

Genetic diagnostic yield by MRI pattern in children with cerebral palsy: a population-based study.

ABSTRACTMarine biological invasions, increasingly facilitated by maritime transport, represent a major dimension of global change, threatening biodiversity, ecosystem services, and human well‐being worldwide. Although the factors shaping invasion success have been widely studied, the evolutionary processes occurring during the transport stage remain poorly understood. Using high‐salinity selection experiments with the model invasive ascidian Ciona robusta, we tested whether transport‐related stress imposed genotype‐dependent filtering. We quantified survival dynamics and employed whole‐genome resequencing together with transcriptomic profiling to characterize genome‐wide responses to environmental filtering. Survival analyses revealed significant mortality differences among genotypes under hypersaline conditions. Whole‐genome resequencing of survivors identified genomic regions with marked genetic differentiation and allele frequency shifts, particularly in osmoregulatory genes such as solute carriers and ion channels. Transcriptomic profiling further demonstrated genotype‐specific expression patterns consistent with stress responses, highlighting the functional relevance of candidate variants. Collectively, our findings show that transport stress drives genotype‐dependent survival and functional genomic signatures consistent with selection. Acknowledging transport as an evolutionary filter and integrating such processes into invasion risk frameworks are essential for developing effective management and prevention measures in an era of accelerating global trade and climate change.