VARista: a free web platform for streamlined whole-genome variant analysis across T2T, hg38, and hg19.

With the increasing importance of genomic data in understanding genetic diseases, there is an essential need for efficient and user-friendly tools that simplify variant analysis. Although multiple tools exist, many present barriers such as steep learning curves, limited reference genome compatibility, or costs. We developed VARista, a free web-based tool, to address these challenges and provide a streamlined solution for researchers, particularly those focusing on rare monogenic diseases. VARista offers a user-centric interface that eliminates much of the technical complexity typically associated with variant analysis. The tool directly supports VCF files generated using reference genomes hg19, hg38, and the emerging T2T, with seamless remapping capabilities between them. Features such as gene summaries and links, tissue and cell-specific gene expression data for both adults and fetuses, as well as automated PCR design and integration with tools such as SpliceAI and AlphaMissense, enable users to focus on the biology and the case itself. As we demonstrate, VARista proved effective in narrowing down potential disease-causing variants, prioritizing them effectively, and providing meaningful biological context, facilitating rapid decision-making. VARista stands out as a freely available and comprehensive tool that consolidates various aspects of variant analysis into a single platform that embraces the forefront of genomic advancements. Its design inherently supports a shift in focus from technicalities to critical thinking, thereby promoting better-informed decisions in genetic disease research. Given its unique capabilities and user-centric design, VARista has the potential to become an essential asset for the genomic research community. https://VARista.link.

Familial Occurrence of Isolated Late-onset Nasolacrimal Duct Obstruction in Two Unrelated Families.

Late-onset nasolacrimal duct obstruction (NLDO) as a result of inflammatory processes causing dacryo-stenosis is a common entity affecting mostly women. While a few mechanisms have been suggested as contributors to the expression of NLDO, the trigger for the inflammation remains mostly unknown. Familial predilection for this condition has not been previously reported. We present two families with multiple individuals affected with congenital or late-onset NLDO, describe the signs and symptoms of the affected individuals, and explore their medical history for any contributing factors. Family A, spanning four generations, included 7 female patients affected by late-onset NLDO. Family B, spanning two generations, included 8 individuals affected by either congenital or late-onset NLDO. This case series suggests a familial predisposition to NLDO, apparently with an autosomal dominant inheritance pattern. Further studies are needed to elucidate the molecular basis of this genetic predisposition.

Compound Heterozygosity for Late-Onset Cardiomyopathy-Causative ALPK3 Coding Variant and Novel Intronic Variant Cause Infantile Hypertrophic Cardiomyopathy.

Hypertrophic and dilated cardiomyopathy (HCM, DCM) are leading causes of cardiovascular morbidity and mortality in children. The pseudokinase alpha-protein kinase 3 (ALPK3) plays an essential role in sarcomere organization and cardiomyocyte differentiation. ALPK3 coding mutations are causative of recessively inherited pediatric-onset DCM and HCM with variable expression of facial dysmorphism and skeletal abnormalities and implicated in dominantly inherited adult-onset cardiomyopathy. We now report two variants in ALPK3-a coding variant and a novel intronic variant affecting splicing. We demonstrate that compound heterozygosity for both variants is highly suggestive to be causative of infantile-onset HCM with webbed neck, and heterozygosity for the coding variant presents with adult-onset HCM. Our data validate partial penetrance of heterozygous loss-of-function ALPK3 mutations in late-onset hypertrophic cardiomyopathy and expand the genotypic spectrum of autosomal recessive ALPK3-related cardiac disease with Noonan-like features.

IHH enhancer variant within neighboring NHEJ1 intron causes microphthalmia anophthalmia and coloboma.

Genomic sequences residing within introns of few genes have been shown to act as enhancers affecting expression of neighboring genes. We studied an autosomal recessive phenotypic continuum of microphthalmia, anophthalmia and ocular coloboma, with no apparent coding-region disease-causing mutation. Homozygosity mapping of several affected Jewish Iranian families, combined with whole genome sequence analysis, identified a 0.5 Mb disease-associated chromosome 2q35 locus (maximal LOD score 6.8) harboring an intronic founder variant in NHEJ1, not predicted to affect NHEJ1. The human NHEJ1 intronic variant lies within a known specifically limb-development enhancer of a neighboring gene, Indian hedgehog (Ihh), known to be involved in eye development in mice and chickens. Through mouse and chicken molecular development studies, we demonstrated that this variant is within an Ihh enhancer that drives gene expression in the developing eye and that the identified variant affects this eye-specific enhancer activity. We thus delineate an Ihh enhancer active in mammalian eye development whose variant causes human microphthalmia, anophthalmia and ocular coloboma. The findings highlight disease causation by an intronic variant affecting the expression of a neighboring gene, delineating molecular pathways of eye development.

Partial penetrance and phenotypic variability of aplasia of lacrimal and salivary glands caused by a novel FGF10 donor splice-site mutation.

Thirteen affected individuals of six generations of a single kindred presented with epiphora evident from infancy. Physical exam and Schirmer test revealed variable expression of tear deficiency, congenital punctal atresia, and dry mouth with multiple caries, without concomitant abnormalities of the ears or digits, commensurate with a diagnosis of aplasia of the lacrimal and salivary glands (ALSG). Reconstruction of the upper lacrimal drainage system was performed in some of the affected individuals. Genetic analysis, testing six affected individuals and three non-affected family members, identified a single novel heterozygous splice-site variant, c.429 + 1, G > T in fibroblast growth factor 10 (FGF10) (NM_004465.1), segregating throughout the family as expected for dominant heredity. RT-PCR assays of HEK-293 cells transfected with wild type or mutant FGF10 demonstrated that the variant causes skipping of Exon 2. Notably, individuals sharing the same variant exhibited phenotypic variability, with unilateral or bilateral epiphora, as well as variable expression of dry mouth and caries. Moreover, one of the variant carriers had no ALSG-related clinical findings, demonstrating incomplete penetrance. While coding mutations in FGF10 are known to cause malformations in the nasolacrimal system, this is the second FGF10 splice-site variant and the first donor-site variant reported to cause ALSG. Thus, our study of a unique large kindred with multiple affected individuals heterozygous for the same FGF10 variant highlights intronic splice-site mutations and phenotypic variability/partial penetrance in ALSG.

Hyperinsulinism/hyperammonemia syndrome caused by biallelic SLC25A36 mutation.

Hyperinsulinism/hyperammonemia (HI/HA) syndrome has been known to be caused by dominant gain-of-function mutations in GLUD1, encoding the mitochondrial enzyme glutamate dehydrogenase. Pathogenic GLUD1 mutations enhance enzymatic activity by reducing its sensitivity to allosteric inhibition by GTP. Two recent independent studies showed that a similar HI/HA phenotype can be caused by biallelic mutations in SLC25A36, encoding pyrimidine nucleotide carrier 2 (PNC2), a mitochondrial nucleotide carrier that transports pyrimidine and guanine nucleotides across the inner mitochondrial membrane: one study reported a single case caused by a homozygous truncating mutation in SLC25A36 resulting in lack of expression of SLC25A36 in patients' fibroblasts. A second study described two siblings with a splice site mutation in SLC25A36, causing reduction of mitochondrial GTP content, putatively leading to hyperactivation of glutamate dehydrogenase. In an independent study, through combined linkage analysis and exome sequencing, we demonstrate in four individuals of two Bedouin Israeli related families the same disease-causing SLC25A36 (NM_018155.3) c.284 + 3A > T homozygous splice-site mutation found in the two siblings. We demonstrate that the mutation, while causing skipping of exon 3, does not abrogate expression of mRNA and protein of the mutant SLC25A36 in patients' blood and fibroblasts. Affected individuals had hyperinsulinism, hyperammonemia, borderline low birth weight, tonic-clonic seizures commencing around 6 months of age, yet normal intellect and no significant other morbidities. Chronic constipation, hypothyroidism, and developmental delay previously described in a single patient were not found. We thus verify that biallelic SLC25A36 mutations indeed cause HI/HA syndrome and clearly delineate the disease phenotype.