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PubMed2024

Identification of a novel intronic variant in COL4A2 gene associated with fetal severe cerebral encephalomalacia and subdural hemorrhage.

BACKGROUND: Genetic variants in COL4A2 are less common than those of COL4A1 and their fetal clinical phenotype has not been well described to date. We present a fetus from China with an intronic variant in COL4A2 associated with a prenatal diagnosis of severe cerebral encephalomalacia and subdural hemorrhage. METHODS: Whole exome sequencing (WES) was applied to screen potential genetic causes. Bioinformatic analysis was performed to predict the pathogenicity of the variant. In in vitro experiment, the minigene assays were performed to assess the variant's effect. RESULTS: In this proband, we observed ventriculomegaly, subdural hemorrhage, and extensive encephalomalacia that initially suggested cerebral hypoxic-ischemic and/or hemorrhagic lesions. WES identified a de novo heterozygous variant c.549 + 5G > A in COL4A2 gene. This novel variant leads to the skipping of exon 8, which induces the loss of 24 native amino acids, resulting in a shortened COL4A2 protein (p.Pro161_Gly184del). CONCLUSION: Our study demonstrated that c.549 + 5G > A in COL4A2 gene is a disease-causing variant by aberrant splicing. This finding enriches the variant spectrum of COL4A2 gene, which not only improves the understanding of the fetal neurological disorders associated with hypoxic-ischemic and hemorrhagic lesions from a clinical perspective but also provides guidance on genetic diagnosis and counseling.

Female↗
PubMed2026

Identification of Two Novel Compound Heterozygous ADAMTS17 Variants Associated With Weill-Marchesani Syndrome 4.

PURPOSE: Weill-Marchesani syndrome 4 (WMS4) is frequently underdiagnosed when standard exome sequencing fails to detect noncoding pathogenic variants. We aimed to identify the genetic cause in a patient with suspected WMS4 and to characterize the splicing-altering mechanism of a deep intronic variant in ADAMTS17. MATERIALS AND METHODS: Whole-exome sequencing combined with whole-genome sequencing was conducted in the proband, who presented with ocular and skeletal manifestations of WMS4. A minigene splicing assay was applied to verify the splicing abnormality caused by the deep intronic variant. RESULTS: The patient showed high myopia, brachydactyly, accelerated growth velocity, and advanced bone age. Two novel compound heterozygous variants in ADAMTS17, c.1655G>A and c.450+38C>A, were identified. The deep intronic variant c.450+38C>A was confirmed by minigene assay to disrupt normal pre-messenger RNA splicing by inducing retention of a 35-base pair intronic segment, leading to a frameshift and premature termination (p.G152Lfs*23). CONCLUSIONS: This study broadens the mutational spectrum of ADAMTS17. Whole-genome sequencing combined with functional splicing validation is essential for resolving molecularly undiagnosed cases of WMS4, particularly when deep intronic variants are suspected, and supports clinical genetic testing and genetic counseling of hereditary connective tissue disorders.

ADAMTS17↗
PubMed2026

A deep intronic IFT172 variant causing pseudoexon inclusion identified by whole-genome sequencing in nephronophthisis.

Nephronophthisis is an autosomal recessive ciliopathy and a major genetic cause of end-stage kidney disease in children and young adults. Although next-generation sequencing panels have improved diagnostic yield, some patients remain genetically unresolved, partly due to deep intronic variants that disrupt pre-mRNA splicing and are not captured by exon-focused approaches. We report a 13-year-old boy who presented with advanced kidney dysfunction, small renal cysts, and kidney histopathology consistent with nephronophthisis. Targeted gene panel sequencing failed to identify causative pathogenic variants beyond a missense variant of uncertain significance. Whole-genome sequencing subsequently revealed compound heterozygous variants in IFT172 (NM_015662.3): a missense variant (c.4696C > T, p.Arg1566Cys) and a deep intronic variant (c.4915-94A > G). In silico analysis predicted activation of cryptic splice sites leading to inclusion of an 86-bp pseudoexon, which was confirmed by a minigene splicing assay. These findings established a molecular diagnosis of IFT172-related nephronophthisis. To our knowledge, this is the first report demonstrating pseudoexon inclusion in IFT172, thereby expanding its mutational spectrum. Our case underscores the importance of evaluating deep intronic regions using whole-genome sequencing and functional validation in genetically unresolved nephronophthisis.

Humans↗
PubMed2026

Expanding the Genomic Spectrum of NHLRC2-Associated FINCA Disease: Integrated Bioinformatic Characterization of a Novel Deep Intronic Variant Predicted to Activate a Pseudoexon.

NHLRC2-associated FINCA disease is an ultra-rare autosomal recessive multisystem disorder caused by biallelic pathogenic variants in NHLRC2. Its mutational spectrum and genotype-phenotype correlations remain incompletely defined, and the contribution of non-coding variants is poorly understood. Here, we report a male infant with a severe FINCA-like phenotype, including early-onset hemolytic anemia, pulmonary involvement, neurodevelopmental impairment, growth failure, recurrent infections, and fatal progression at 8.5 months. Whole-genome sequencing identified a compound heterozygous NHLRC2 genotype comprising the previously reported pathogenic missense variant c.442G>T (p.Asp148Tyr) and a novel deep intronic variant, c.331+6863A>G. Segregation analysis confirmed inheritance from different parents. Integrated genomic and splicing analysis predicted that c.331+6863A>G creates a strong cryptic donor splice site and supports pseudoexon inclusion. Reconstruction of the predicted aberrant transcript indicated premature termination and potential susceptibility to nonsense-mediated mRNA decay. To our knowledge, this is the first reported deep intronic NHLRC2 variant predicted to activate pseudoexon inclusion. Although experimental validation was unavailable, convergent clinical, segregation, population, and computational evidence supports c.331+6863A>G as the most plausible second disease-associated allele. This case expands the genomic spectrum of NHLRC2-associated FINCA disease and highlights the diagnostic value of phenotype-driven whole-genome sequencing.

Humans↗
PubMed2026

Clinical and functional characterization of a novel homozygous non-canonical splice mutation (c.1910-15_1910-11delinsTTACA) in CEP290 causing Joubert syndrome.

BACKGROUND: Joubert syndrome (JS) is a rare, predominantly autosomal recessive neurodevelopmental disorder characterized by hypotonia, motor delay, intellectual disability, oculomotor apraxia, and the hallmark "molar tooth sign" on axial view of MRI. JS is genetically heterogeneous, with pathogenic variants identified in more than 40 genes involved in primary cilia function. Among these, CEP290 is one of the most frequently mutated genes. RESULTS: In this study, we investigated two children-an 11-year-old boy (the proband) and his 5-year-old sister-both presenting with a similar phenotype consistent with JS. The parents, who self-identified as Chechen, reported distant consanguinity. The family also included a healthy 13-year-old daughter. The proband had previously been evaluated by a neurologist and underwent whole-genome sequencing (WGS); however, no causative variants were identified initially. After phenotype reassessment by a clinical geneticist, we performed a reanalysis of the raw WGS data and identified a novel homozygous intronic variant of uncertain significance (VUS), c.1910-15_1910-11delinsTTACA in CEP290 (NM_025114.4). Sanger sequencing confirmed that both the proband and his affected sister were homozygous for this variant, which they inherited from their heterozygous parents. Their healthy sister did not carry the variant. mRNA-sequencing and targeted cDNA sequencing (read depth ~ 100,000x) demonstrated that this intronic variant causes completely aberrant splicing of CEP290 pre-mRNA. Predominantly this variant causes the skipping of exon 20 in the main CEP290 transcript. Alternatively, the variant results in partial inclusion of intron 19 into the mRNA, elongation of exon 20 by 58 nucleotides, and a homozygous substitution chr12:88114573 (ACTGTGTA> TTACAGTA). No canonical mRNA isoform was detected when the variant was homozygous. Both the predicted severe truncation and the likely degradation of aberrant transcripts through nonsense-mediated decay (NMD) would correspond to complete loss of CEP290 function. Following the reclassification of this VUS to likely pathogenic, the family was able to pursue in vitro fertilization (IVF) with preimplantation genetic testing for monogenic disorders (PGT-M). CONCLUSION: Our study highlights the critical importance of proper phenotyping prior to referral for WES/WGS as well as of combining NGS with functional mRNA studies to achieve a molecular diagnosis for patients with predicted splice-site mutations in JS-associated genes. It also emphasizes the need for functional reassessment of VUS when genomic data are expected to guide reproductive decision-making within affected families.

Humans↗
PubMed2025

Whole-genome sequencing, as a powerful diagnostic tool in hearing loss, reveals novel variants in PTPRQ missed by whole-exome sequencing.

BACKGROUND/OBJECTIVES: Hearing loss (HL) is one of the most common congenital disorders, affecting 1-2 in 1,000 newborns. Modern genetic diagnostics using large gene panels and/or whole exome analysis (WES) can identify disease-causing mutations in 25-50 % of patients, with higher solve rates in individuals with earlier onset. RESULTS: Here, we used whole-genome sequencing (WGS) to reanalyze 14 index patients/families who remained without genetic diagnosis by WES. We were able to identify the genetic cause of HL in 6 families (43 %). Two families were diagnosed with DFNB84A caused by compound heterozygous recessive mutations in PTPRQ. Three of the four underlying variants, including a structural variant, a deep intronic variant, and a splice variant, escaped detection by WES. Minigene assays confirmed the pathogenicity of the intronic and the splice variants. In addition, we used protein 3D structure prediction and rigid ligand docking to study the pathogenicity of variants that escape nonsense-mediated decay. CONCLUSION: In our study, we present four novel variants in PTPRQ, three of which were detected only by WGS. To our knowledge, we report here the first pathogenic deep intronic PTPRQ variant causing HL. Our results suggest that the mutational spectrum of PTPRQ is not well covered by standard WES and that PTPRQ-associated hearing loss may be more frequent than previously thought. WGS provides an additional layer of information in the diagnostics of HL.

Humans↗
PubMed2026

X-linked spondyloepiphyseal dysplasia tarda misdiagnosed as growth hormone deficiency: identification of a novel intronic TRAPPC2 variant by whole-genome sequencing.

BACKGROUND: X-linked spondyloepiphyseal dysplasia tarda (SEDT) is a rare skeletal dysplasia caused by pathogenic variants in TRAPPC2 and typically presents in late childhood or adolescence with short-trunk disproportion and vertebral dysplasia. CASE PRESENTATION: We describe a family series centered on an adolescent male initially diagnosed with GHD due to reduced height velocity and subnormal GH stimulation results, who received recombinant human GH (rhGH) therapy for three years with negligible improvement. During puberty, he developed progressive short-trunk disproportion and characteristic radiographic features, including platyspondyly and posterior hump-shaped vertebral endplates, suggestive of SEDT. Whole-exome sequencing (WES) was nondiagnostic, whereas whole-genome sequencing (WGS) identified a novel intronic TRAPPC2 variant, c.239-20_239-12delinsAATGAA, initially classified as a variant of uncertain significance (VUS). Segregation analysis across the family enabled reclassification of the variant to likely pathogenic, confirming X-linked SEDT. The proband's younger brother exhibited earlier radiologic abnormalities and, notably, a favorable response to rhGH, whereas the younger sister-an asymptomatic heterozygous carrier-showed normal spinal morphology, consistent with expected female carrier phenotypes. CONCLUSIONS: This family-based report underscores the generally limited therapeutic effect of rhGH in SEDT while highlighting potential interindividual variability, as evidenced by the younger male sibling's response. It further emphasizes the diagnostic utility of WGS for detecting deep intronic variants missed by WES and the importance of segregation analysis in resolving VUS in rare skeletal dysplasias.

Humans↗
PubMed2026

Utility of High-Throughput Genomic Analysis for Genetic Counseling in Large Family with Wilson Disease Carrying a Novel 28-bp ATP7B Splice-Junction Deletion.

Background/Objectives: Wilson disease (WD) is an autosomal recessive disorder of copper metabolism caused by pathogenic variants in the ATP7B gene. Early diagnosis and appropriate treatment are essential for preventing irreversible complications. This study demonstrated the clinical utility of integrated high-throughput genomic analysis for molecular diagnosis and genetic counseling in a large Thai family affected by WD. Methods: A 32-year-old woman with clinical features suggestive of WD underwent clinical, biochemical, and molecular genetic evaluations, including sequencing of the entire ATP7B gene and SNP microarray. Fluorescent PCR followed by capillary electrophoresis was used for segregation analysis in available family members. SNP microarray analysis and whole-exome sequencing were performed on the proband's husband to identify pathogenic variants in the ATP7B gene and other disease-associated genes for reproductive risk assessment. Results: The proband presented with hepatic dysfunction, Kayser-Fleischer rings, low serum ceruloplasmin, and a family history of fatal liver disease. She also developed progressive weakness, with nerve conduction findings consistent with axonal sensorimotor polyneuropathy predominantly affecting the lower limbs. Sequencing identified a novel homozygous 28-bp splice-junction deletion, c.4022-24_4025del, which disrupted the canonical splice acceptor site at the intron 19/exon 20 boundary and was classified as pathogenic variant. Segregation analysis confirmed carrier status in the proband's father and identified heterozygous carrier or homozygous wild-type status among her living siblings. SNP microarray analysis revealed a 46.7 Mb copy-neutral long contiguous stretch of homozygosity (CN-LCSH) encompassing ATP7B, with CN-LCSH regions accounting for 2.046% of the total autosomal genome. These findings potentially reflected segmental uniparental isodisomy or identity by descent, while the overall homozygosity pattern did not support recent consanguinity. Combined genomic analyses of the proband's husband revealed no pathogenic or likely pathogenic ATP7B variants. Based on the available testing, all offspring are expected to be heterozygous carriers, and the risk of an affected child is considered very low. Conclusions: This study highlights the value of integrated genomic analysis for molecular diagnosis, cascade testing, and reproductive risk counseling. Further functional studies should be conducted to validate their pathogenicity.

ATP7B↗
PubMed2025

Novel compound heterozygous DOCK6 variants expand the mutational spectrum in prenatal diagnosis of Adams-Oliver syndrome 2.

BACKGROUND: Adams-Oliver syndrome (AOS) is a rare developmental disorder, and the DOCK6 gene is an identified AOS gene. This report highlights the prenatal diagnosis of AOS-2 by ultrasonography and genetic testing. METHODS: A growth-restricted fetus with bilateral ventriculomegaly, paraventricular calcifications, and ventricular septal defect underwent trio-whole-exome sequencing (trio-WES). Functional validation of the splice-altering variant was performed via minigene assays and protein structural modeling. RESULTS: Trio-WES revealed compound heterozygous DOCK6 variants: a paternal frameshift (c.3190_3191del; p. Leu1064Valfs60) and a maternal splice-site variant (c.3241-1G > T). Minigene assays demonstrated that c.3241-1G > T caused intron 26 retention (486 bp), introducing a premature termination codon (p. Val1081Glufs37). Structural modeling confirmed the loss of critical DHR2 domains in both truncated proteins. CONCLUSIONS: This study expands the mutational spectrum of DOCK6 and underscores the importance of combining prenatal imaging with functional genomics for early diagnosis of AOS2.

Adult↗
PubMed2026

[Genetic and phenotypic analysis of three children with Neurodevelopmental disorders due to variants of DEAF1 gene].

OBJECTIVE: To explore the genetic characteristics and clinical phenotypes of three children with novel DEAF1 gene variants. METHODS: Three children who were referred to Capital Children's Medical Center Affiliated to Capital Medical University between January 2018 and December 2025 were selected as study subjects and underwent whole exome sequencing (WES). Candidate variants were verified by Sanger sequencing, and their pathogenicity was evaluated based on the guidelines from American College of Medical Genetics and Genomics (ACMG). A systematic search of databases including PubMed and CNKI was conducted to compile previously reported cases of DEAF1 variants for clinical phenotype comparison. For the non-canonical splice site variant c.870+5G>C located in the intronic region, wild-type and mutant minigene reporter vectors were constructed and transfected into HeLa and 293T cells, respectively, and the splicing patterns were analyzed by RT-PCR and Sanger sequencing. This study was approved by the Medical Ethics Committee of Capital Institute of Pediatrics (Ethics No.: SHERLL 2020001). RESULTS: All three children were found to have carried de novo heterozygous variants of the DEAF1 gene, including two missense variants (c.764G>A, c.641T>C) in the important SAND domain and a splice site variant (c.870+5G>C) in a non-canonical splicing region. The c.764G>A and c.870+5G>C variants were unreported previously. All children had presented with intellectual developmental delay, and two were accompanied by autism spectrum disorder, and two had epilepsy and sleep disorders. In vitro minigene splicing assay showed that the c.870+5G>C variant can lead to abnormal splicing. CONCLUSIONS: This study reported three children with novel DEAF1 variants, two of which have not been previously described, thereby enriched the mutational spectrum of the DEAF1 gene. In vitro functional assay combined with the clinical manifestations of the patients confirmed the pathogenicity of the non-canonical splice site variant in the intronic region.

Humans↗
PubMed2025

Genetic analysis of four cases of Poirier Bienvenu neurodevelopmental syndrome associated with CSNK2B variant.

BACKGROUND: CSNK2B deficiency underlies the pathogenesis of Poirier-Bienvenu neurodevelopmental syndrome (POBINDS). In this study, we present four cases of pediatric seizures caused by de novo variants in CSNK2B, with the aim to reinforce the clinical and variant data pertaining to early genetic factors associated with epilepsy. METHODS: Trio whole exome sequencing were used to detect variants in the proband and her family members, and bioinformatics annotation was performed for the variant. Sanger sequencing and CSNK2B cDNA sequencing were employed to ascertain the carrier status of additional family members and evaluate the potential impact of variants on splicing. RESULTS: All four cases presented with epilepsy as the initial manifestation, accompanied by global developmental delay, particularly in language and motor developmental delay. Cases 1, 3 and 4 exhibited full-scale tonic-clonic seizures, while case 2 displayed myoclonic and typical absence seizures. Furthermore, case 2 demonstrated delayed growth and development compared to age-matched peers. No abnormality was detected in the head magnetic resonance imaging (MRI). Genetic analysis revealed novel heterozygous variants in the CSNK2B gene in all four cases, including c.175 + 1G > A, c.73-2A > G, c.291 + 1G > A and c.481delA. In case 2, reverse transcription analysis of CSNK2B mRNA revealed the retention of the 3' end sequence of Intron 2 and deletion of the 5' end sequence of Exon 3. In treatment, four case received a combination of one to three types of antiseizure medication and rehabilitation training individually. Case 1 continued to experience seizures to varying degrees, while cases 2-4 demonstrated effective seizure control. Overall motor and intellectual development improved in all four cases, however, there was slow recovery in language function. CONCLUSION: This study elucidates the molecular etiology of epilepsy in four cases with POBINDS and expands the mutational spectrum of pathogenic variants in the CSNK2B, highlighting their impact on splicing. The highly genetic heterogeneous phenotype of POBINDS relies on the detection of pathogenic variants in CSNK2B. Conventional antiseizure medication effectively control seizures, while rehabilitation treatment can significantly improve intelligence and motor function to varying degrees; however, language recovery tends to be relatively slow.

Humans↗
PubMed2026

Individualized antisense oligonucleotides for SCN2A-related developmental epileptic encephalopathy.

SCN2A variants are among the most common genetic causes of developmental and epileptic encephalopathies (DEEs), which can present with uncontrolled seizures at birth and account for 1-2% of all epileptic encephalopathies. A substantial fraction of causal variants are gain-of-function or mixed-function variants associated with increased channel open probability or greater sodium current flux. Here two parallel n = 1 clinical studies were conducted in two patients (9-year-old and 14-year-old boys) with SCN2A-related DEE. Individualized allele-selective antisense oligonucleotides (ASOs) were designed to target heterozygous intronic single-nucleotide polymorphisms (SNPs) for decreased expression of mutant SCN2A transcript while preserving the wild-type copy. Primary endpoints included quantitative change from baseline in seizure frequency and neurodevelopment, including motor scores. Efficacy measures were also individualized to each patient's phenotype, including refractory seizures, developmental delay, autism spectrum disorder, choreoathetosis and gastrointestinal dysfunction. Patients experienced a reduction in seizure frequency (26% and 90% in the two patients, respectively), decreased use of concomitant medications and improvement in neurodevelopmental skills. Both ASOs were well tolerated, with no ASO-related serious adverse events. Continued long-term follow-up of these preliminary positive safety and efficacy findings is needed to confirm the disease-modifying potential of these ASOs. Haplotype phasing in a separate cohort of infants with SCN2A-related disorder (SCN2A-RD), diagnosed by rapid whole-genome sequencing, identified 16% of patients with compatible SNPs. These data provide a pathway from n = 1 to n of more patients with SCN2A-RD and other monogenic disorders. ClinicalTrials.gov registration: NCT06314490 .

Adolescent↗
PubMed2024

SLC12A1 variant c.1684+1 G>A causes Bartter syndrome type 1 by promoting exon 13 skipping.

BACKGROUND: Bartter syndrome type 1, an autosomal recessive genetic disorder, is caused by pathogenic loss-of-function variants in the SLC12A1 gene. It is characterized by metabolic alkalosis and prenatal-onset polyuria leading to polyhydramnios. METHODS: We identified pathogenic gene in a 12-day-old newborn boy with Bartter syndrome type 1 using whole-exome sequencing. Sanger sequencing validated the identified variants. A minigene assay was performed to investigate the effect of a novel splice site variant on pre-mRNA splicing. RESULTS: We found a compound heterozygous variants in the SLC12A1 gene, consisting of a known pathogenic missense mutation (NM_000338: c.769 G>A; p.Gly257Ser) and a novel splice site variant (c.1684+1 G>A). In silico predictions and an in vitro minigene splicing assay demonstrated that the splicing variant c.1684+1 G>A abolished a consensus splice donor site of SLC12A1 intron 13, resulting in complete exon 13 skipping, translational frameshift, and premature termination codon, ultimately leading to loss of SLC12A1 function. CONCLUSION: Using a cell-based in vitro assay, we revealed the aberrant effect of the pathogenic splicing variant SLC12A1 c.1684+1 G>A on pre-mRNA splicing. Our findings expand the gene mutation spectrum of Bartter syndrome type 1, providing a basis for genetic diagnosis and the development of genetic medicines.

Humans↗
PubMed2026

Seventeen-year follow-up of hypophosphatasia diagnosed in middle-aged siblings harboring a novel intronic and a rare missense ALPL gene mutation.

Hypophosphatasia (HPP) is the rare inborn-error-of-metabolism that features impaired mineralization of the skeleton and teeth due to a deactivating mutation or mutations of the gene ALPL which encodes the tissue-nonspecific isoenzyme of alkaline phosphatase (TNSALP). We report 17-year follow-up of twin sisters and a brother referred in middle-age for painful proximal femoral "stress fractures" and then diagnosed with HPP. They reported generalized muscle and bone pain, metatarsal fractures, arthropathy and, since childhood, tooth loss. Their concordant findings were explained by compound heterozygosity in ALPL for a rare maternal missense mutation (c.1403C > T, p.Ala468Val) in exon 12, together with a novel presumably paternal change (c.863-14G > A) predicting a cryptic mRNA splice site in intron 8. Fractures continued during follow-up until one sister received a three-and-one-half-year course of hydroxyapatite-targeted TNSALP supplementation therapy (asfotase alfa) during which substantial improvement occurred in her clinical, biochemical, and functional parameters as well as quality of life. Following subsequent unplanned treatment cessation she suffered significant clinical deterioration, including new fractures and loss of mobility. Her bone histopathology documented osteomalacia. Treatment resumption restored its benefits. Among ten asymptomatic family members evaluated in this four-generation kindred, eight were carriers heterozygous for either ALPL mutation. Those harboring the maternal missense defect manifested mild hypophosphatasemia, suggesting a dominant-negative mutation effect. This experience underscores the importance of in-depth phenotyping and then clinical follow-up to characterize ALPL variant combinations, and for maintaining effective asfotase alfa treatment.

Humans↗
PubMed2026

Dual Aberrant Splicing Caused by an Apparently Missense CHD7 Variant, c.5273A>G (p.Asp1758Gly), in CHARGE Syndrome.

CHARGE syndrome is a rare congenital disorder primarily attributed to heterozygous pathogenic variants of the CHD7 gene. Most pathogenic CHD7 variants are loss-of-function (LoF) variants, whereas the interpretation of missense variants remains challenging in the absence of functional evidence for their pathogenicity. We report a female infant presenting with clinical features characteristic of CHARGE syndrome. Targeted sequencing identified a heterozygous CHD7 variant (NM_017780.4:c.5273A>G), initially annotated as a missense substitution p.Asp1758Gly. This variant has been previously reported and registered with conflicting pathogenicity classifications; however, its transcript-level consequences remain unclear. Long-PCR-based RNA sequencing of total RNA from peripheral blood mononuclear cells revealed two aberrant splicing patterns associated with the variant: a predominant transcript carrying a 28-bp deletion due to cryptic donor splice-site activation, and a minor transcript with partial intron 24 retention. Both transcripts were predicted to result in premature termination codons. These findings demonstrate that c.5273A>G functions as a LoF variant through dual aberrant splicing rather than a simple missense substitution. This case underscores the importance of RNA-level splicing analysis for the accurate interpretation and classification of CHD7 missense variants.

CHD7↗
PubMed2025

KDM3B Regulates Postradiation Fibrotic Responses in Prostate Stroma via N6-methyladenosine Modification of LOX.

PURPOSE: Genome-wide association studies have uncovered single-nucleotide polymorphisms (SNPs) linked to radiation therapy (RT)-induced toxicities in patients with prostate cancer. SNP rs17599026, located in intron 21 of the KDM3B gene, has been associated with late-onset urinary toxicity, with an increased frequency of urination observed 2 years post-RT compared with pretreatment conditions. This study aimed to explore the underlying mechanisms driving this association. METHODS AND MATERIALS: A clustered regularly interspaced short palindromic repeats-dead Cas9 prime editing system was used to mimic KDM3B genetic variants in prostate stromal cell lines. Murine models with wild-type and heterozygous Kdm3b genotypes were used to assess fibrosis following radiation. RNA immunoprecipitation, transcript stability assays, and protein analysis elucidated the role of N6-methyladenosine (m6A) modification in regulating lysyl oxidase (LOX) expression. α-ketoglutarate (α-KG) supplementation was tested for its effects on KDM3B protein stability, LOX expression, and fibrosis mitigation. RESULTS: The rs17599026 SNP reduced KDM3B protein expression via circular RNA and microRNA-mediated mechanisms, leading to decreased m6A modification and increased stability of LOX messenger RNA. Elevated LOX expression promoted collagen cross-linking and fibrosis in prostate stroma. α-KG supplementation restored KDM3B protein levels, reduced LOX expression, and mitigated fibrosis in vitro and in vivo. CONCLUSIONS: KDM3B genetic variations influence radiation-induced fibrosis through posttranscriptional regulation of LOX. Dietary α-KG supplementation may serve as a mechanism-based strategy to alleviate radiation toxicity in patients with prostate cancer, offering a potential therapeutic pathway to improve treatment outcomes.

Male↗
PubMed2026

A Novel Splice Variant in the COL1A1 Gene Leads to Exon 46 Skipping and Osteogenesis Imperfecta.

BACKGROUND: Osteogenesis imperfecta (OI) is a clinical and genetic disorder characterised by bone fragility, growth deficiency and skeletal deformity. Ninety per cent of OI cases are attributable to autosomal dominant variants in the COL1A1 and COL1A2 genes. METHODS: Candidate variants were identified and verified through trio whole-exome sequencing (trio-WES), copy number variation sequencing (CNV-seq) and Sanger sequencing. Minigene splicing assays were performed in HeLa and HEK293T cells with pcDNA3.1 and pcMINI-C vectors to investigate the function of the candidate variants. A systematic review of COL1A1 splicing variants and the corresponding genotype-phenotype spectrum was performed. RESULTS: Trio-WES revealed a novel heterozygous variant in the C-terminal region of the COL1A1 gene: NM_000088.4:c.3423+5G>A. Sanger sequencing confirmed the variant in both the proband (II-2) and her foetus (III-1) who were clinically suspected of having OI. The c.3423+5G>A variant causes complete skipping of Exon 46, as demonstrated by a minigene splicing assay. We retrieved 419 COL1A1 splicing variants from PubMed, excluded 15 without phenotypic data and 2 linked to Ehlers-Danlos syndrome and stratified the remaining 402 variants into three types on the basis of splice site location: (1) Variants at canonical splicing sites (77.8%, 313/402) mostly cause mild phenotypes, whereas a minority may be severe. (2) Intron variants in other locations, such as splice region variants (17.9%, 72/402), usually cause mild clinical phenotypes, and deep intronic splice variants (0.4%, 2/402) that may result in severe phenotypes. (3) Other variants (3.7%, 15/402), such as exon variants or fragment loss, are extremely rare. We also preliminarily discuss the mechanisms underlying phenotypic variability and the characteristics of C-terminal variants. CONCLUSIONS: This intron variant in COL1A1 was classified as likely pathogenic and was confirmed to disrupt COL1A1 expression. The summary analysis results also revealed a correlation among splicing variants, C-terminal region variants and disease, suggesting that variant location provides a useful framework for prognosis prediction.

Female↗
PubMed2024

Identification of four TTN variants in three families with fetal akinesia deformation sequence.

BACKGROUND: TTN is a complex gene with large genomic size and highly repetitive structure. Pathogenic variants in TTN have been reported to cause a range of skeletal muscle and cardiac disorders. Homozygous or compound heterozygous mutations tend to cause a wide spectrum of phenotypes with congenital or childhood onset. The onset and severity of the features were considered to be correlated with the types and location of the TTN variants. METHODS: Whole-exome sequencing was performed on three unrelated families presenting with fetal akinesia deformation sequence (FADS), mainly characterized by reduced fetal movements and limb contractures. Sanger sequencing was performed to confirm the variants. RT-PCR analysis was performed. RESULTS: TTN c.38,876-2 A > C, a meta transcript-only variant, with a second pathogenic or likely pathogenic variant in trans, was observed in five affected fetuses from the three families. Sanger sequencing showed that all the fetal variants were inherited from the parents. RT-PCR analysis showed two kinds of abnormal splicing, including intron 199 extension and skipping of 8 bases. CONCLUSIONS: Here we report on three unrelated families presenting with FADS caused by four TTN variants. In addition, our study demonstrates that pathogenic meta transcript-only TTN variant can lead to defects which is recognizable prenatally in a recessive manner.

Humans↗