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The post-transcriptional modification of tRNAs plays a crucial role in tRNA structure and function. Pathogenic variants in tRNA-modification enzymes have been implicated in a wide range of human neurodevelopmental and neurological disorders. However, the molecular basis for many of these disorders remains unknown. Here, we describe a comprehensive cohort of 43 individuals from 31 unrelated families with bi-allelic variants in tRNA methyltransferase 1 (TRMT1). These individuals present with a neurodevelopmental disorder universally characterized by developmental delay and intellectual disability, accompanied by variable behavioral abnormalities, epilepsy, and facial dysmorphism. The identified variants include ultra-rare TRMT1 variants, comprising missense and predicted loss-of-function variants, which segregate with the observed clinical pathology. Our findings reveal that several variants lead to mis-splicing and a consequent loss of TRMT1 protein accumulation. Moreover, cells derived from individuals harboring TRMT1 variants exhibit a deficiency in tRNA modifications catalyzed by TRMT1. Molecular analysis reveals distinct regions of TRMT1 required for tRNA-modification activity and binding. Notably, depletion of Trmt1 protein in zebrafish is sufficient to induce developmental and behavioral phenotypes along with gene-expression changes associated with disrupted cell cycle, immune response, and neurodegenerative disorders. Altogether, these findings demonstrate that loss of TRMT1-catalyzed tRNA modifications leads to intellectual disability and provides insight into the molecular underpinnings of tRNA-modification deficiency caused by pathogenic TRMT1 variants.

Background: The heterogeneous features of enamel renal syndrome (ERS) make diagnosis and treatment challenging. The main symptoms are disturbed amelogenesis and nephrocalcinosis. Bi-allelic likely pathogenic (LP) or pathogenic (P) variants in FAM20A have been associated with the syndrome since 2012. Affected patients often receive extensive dental treatment because of deviant orofacial morphology. However, knowledge about long-term prognosis and treatment guidelines are still lacking. The complex nature of ERS might endanger both dental and general health. The purpose of this article is to highlight the risks of overlooking the symptoms of the syndrome, and to discuss management strategies, surveillance and prognosis.

Case presentation: We report the management of a case with suspected ERS after initial dental treatment elsewhere with no adjustment for the syndrome. Dental treatment was revised and followed for 8 years. Complementary medical examinations were conducted, and ERS was genetically confirmed, revealing homozygosity for a LP c.755_757del, p.(Phe252del) variant in FAM20A. The nephrological investigation revealed medullary calcium deposits, normal renal function and hypophosphatemia. Urine analysis revealed hypocitraturia and hypocalciuria. Accordingly, the patient now medicates with potassium citrate to decrease the risk of progressive renal stone formation.

Conclusion: We herein describe a patient with confirmed ERS with an 8-year follow-up. Diagnostic delay until adulthood led to complicated dental treatment. The results of nephrological investigations are presented. The importance of dental and medical multidisciplinary management in syndromic disorders affecting the formation of the enamel is also exemplified. The dental prognosis after rehabilitation is likely affected by anatomical variations and patient cooperation. The prognosis for renal function seems to be good. However, lifelong surveillance of renal function is recommended. Registration: The ethics committee in Uppsala, Sweden, determined that ethical approval was not necessary in this case (2019-04835). Informed consent was obtained from the participant in writing and is documented in the medical records.

RNA binding motif protein X‐linked (RBMX) encodes the heterogeneous nuclear ribonucleoprotein G (hnRNP G) that regulates splicing, sister chromatid cohesion and genome stability. RBMX knock down experiments in various model organisms highlight the gene’s importance for brain development. Deletion of the RGG/RG motif in hnRNP G has previously been associated with Shashi syndrome, however involvement of other hnRNP G domains in intellectual disability remain unknown. In the current study, we present the underlying genetic and molecular cause of Gustavson syndrome. Gustavson syndrome was first reported in 1993 in a large Swedish five-generation family presented with profound X-linked intellectual disability and an early death. Extensive genomic analyses of the family revealed hemizygosity for a novel in-frame deletion in RBMX in affected individuals (NM_002139.4; c.484_486del, p.(Pro162del)). Carrier females were asymptomatic and presented with skewed X-chromosome inactivation, indicating silencing of the pathogenic allele. Affected individuals presented minor phenotypic overlap with Shashi syndrome, indicating a different disease-causing mechanism. Investigation of the variant effect in a neuronal cell line (SH-SY5Y) revealed differentially expressed genes enriched for transcription factors involved in RNA polymerase II transcription. Prediction tools and a fluorescence polarization assay imply a novel SH3-binding motif of hnRNP G, and potentially a reduced affinity to SH3 domains caused by the deletion. In conclusion, we present a novel in-frame deletion in RBMX segregating with Gustavson syndrome, leading to disturbed RNA polymerase II transcription, and potentially reduced SH3 binding. The results indicate that disruption of different protein domains affects the severity of RBMX-associated intellectual disabilities.

We present the case of a male patient who was ultimately diagnosed with Becker muscular dystrophy (BMD; MIM# 300376) after the onset of muscle weakness in his teens progressively led to significant walking difficulties in his twenties. A genetic diagnosis was pursued but initial investigation revealed no aberrations in the dystrophin gene (DMD), although immunohistochemistry and Western blot analysis suggested the diagnosis of dystrophinopathy. Eventually, after more than 10 years, an RNA analysis captured abnormal splicing where 154 nucleotides from intron 43 were inserted between exon 43 and 44 resulting in a frameshift and a premature stop codon. Normal splicing of the DMD gene was also observed. Additionally, a novel variant c.6291–13537A>G in DMD was confirmed in the genomic DNA of the patient. The predicted function of the variant aligns with the mRNA results. To conclude, we here demonstrate that mRNA analysis can guide the diagnosis of non-coding genetic variants in DMD.

The Nexilin F-Actin Binding Protein (Nexilin) encoded by NEXN is a cardiac Z-disc protein important for cardiac function and development in humans, zebrafish, and mice. Heterozygote variants in the human NEXN gene have been reported to cause dilated and hypertrophic cardiomyopathy. Homozygous variants in NEXN cause a lethal form of human fetal cardiomyopathy, only described in two patients before. In a Swedish, four-generation, non-consanguineous family comprising 42 individuals, one female had three consecutive pregnancies with intrauterine fetal deaths caused by a lethal form of dilated cardiomyopathy. Whole-exome sequencing and variant analysis revealed that the affected fetuses were homozygous for a NEXN variant (NM_144573:c.1302del;p.(Ile435Serfs*3)). Moreover, autopsy and histology staining declared that they presented with cardiomegaly and endocardial fibroelastosis. Immunohistochemistry staining for Nexilin in the affected fetuses revealed reduced antibody staining and loss of striation in the heart, supporting loss of Nexilin function. Clinical examination of seven heterozygote carriers confirmed dilated cardiomyopathy (two individuals), other cardiac findings (three individuals), or no cardiac deviations (two individuals), indicating incomplete penetrance or age-dependent expression of dilated cardiomyopathy. RNA sequencing spanning the variant in cDNA blood of heterozygote individuals revealed nonsense-mediated mRNA decay of the mutated transcripts. In the current study, we present the first natural course of the recessively inherited lethal form of human fetal cardiomyopathy caused by loss of Nexilin function. The affected family had uneventful pregnancies until week 23-24, followed by fetal death at week 24-30, characterized by cardiomegaly and endocardial fibroelastosis.

ALG2 mutations are extremely rare causes of congenital myasthenic syndromes (CMS). The clinical phenotype and treatment response is therefore not well described. We present the case of a baby who immediately after birth presented with pronounced truncal hypotonia, proximal muscle weakness and feeding difficulties. Single fibre electromyography showed neuromuscular transmission failure and salbutamol and ephedrine treatment improved both muscle weakness and neuromuscular transmission. Genetic analysis revealed a likely pathogenic variant c.1040del, p.(Gly347Valfs*27) in exon 2 and a variant of uncertain significance, c.239G>A, p.(Gly80Asp) in exon 1 of the ALG2 gene. Western blot in whole cell lysates of HEK293 cells transfected with p.Gly80Asp, or p.Gly347Valfs*27 expression constructs indicated that p.Gly347Valfs*27 is likely a null allele and p.Gly80Asp is pathogenic through marked reduction of ALG2 expression. This case highlights the utility of functional studies in clarifying variants of unknown significance, in suspected cases of CMS. (C) 2021 The Author(s). Published by Elsevier B.V.

Pathogenic variants in the LRP2 gene, encoding the multiligand receptor megalin, cause a rare autosomal recessive syndrome: Donnai-Barrow/Facio-Oculo-Acoustico-Renal (DB/FOAR) syndrome. Because of the rarity of the syndrome, the long-term consequences of the tubulopathy on human renal health have been difficult to ascertain, and the human clinical condition has hitherto been characterized as a benign tubular condition with asymptomatic low-molecularweight proteinuria. We investigated renal function and morphology in a murine model of DB/FOAR syndrome and in patients with DB/FOAR. We analyzed glomerular filtration rate in mice by FETC-inulin clearance and clinically characterized six families, including nine patients with DB/FOAR and nine family members. Urine samples from patients were analyzed by Western blot analysis and biopsy materials were analyzed by histology. In the mouse model, we used histological methods to assess nephrogenesis and postnatal renal structure and contrast-enhanced magnetic resonance imaging to assess glomerular number. In megalin-deficient mice, we found a lower glomerular filtration rate and an increase in the abundance of injury markers, such as kidney injury molecule-1 and N-acetyl-11-n-glucosaminidase. Renal injury was validated in patients, who presented with increased urinary kidney injury molecule-1, classical markers of chronic kidney disease, and glomerular proteinuria early in life. Megalin-deficient mice had normal nephrogenesis, but they had 19% fewer nephrons in early adulthood and an increased fraction of nephrons with disconnected glomerulotubular junction. In conclusion, megalin dysfunction, as present in DB/FOAR syndrome, confers an increased risk of progression into chronic kidney disease.

Intellectual Disability (ID) is a clinically heterogeneous condition that affects 2-3% of population worldwide. In recent years, exome sequencing has been a successful strategy for studies of genetic causes of ID, providing a growing list of both candidate and validated ID genes. In this study, exome sequencing was performed on 28 ID patients in 27 patient-parent trios with the aim to identify de novo variants (DNVs) in known and novel ID associated genes. We report the identification of 25 DNVs out of which five were classified as pathogenic or likely pathogenic. Among these, a two base pair deletion was identified in the PUF60 gene, which is one of three genes in the critical region of the 8q24.3 microdeletion syndrome (Verheij syndrome). Our result adds to the growing evidence that PUF60 is responsible for the majority of the symptoms reported for carriers of a microdeletion across this region. We also report variants in several genes previously not associated with ID, including a de novo missense variant in NAA15. We highlight NAA15 as a novel candidate ID gene based on the vital role of NAA15 in the generation and differentiation of neurons in neonatal brain, the fact that the gene is highly intolerant to loss of function and coding variation, and previously reported DNVs in neurodevelopmental disorders.

Vestibular disorders comprise a heterogeneous group of diseases with transient or permanent loss of vestibular function. Vestibulopathy is in most cases associated with migraine, Meniere disease, hereditary ataxias, or sensorineural hearing loss. We identified two brothers and their first cousin affected by hearing loss and episodic vertigo. The brothers were homozygous STRC nonsense variant [c.4027 C> T, p.(Q1343*)], whereas their first cousin was compound heterozygous for the STRC nonsense variant and a 97 kb deletion spanning the entire STRC gene. Clinical investigations confirmed pathological vestibular responses in addition to a characteristic DFNB16 hearing loss. The STRC gene encodes Stereocilin in the cochlea and in the vestibular organ where it ensheathes the kinocilium of the otolithic membranes. Stereocilin is associated with the gel overlaying the vestibular kinocilia, suggesting a role for the protein in sensing balance and spatial orientation. Our findings support such a function for Stereocilin in the vestibular organ and expand the phenotype associated with DFNB16.

Mutations in genes involved in the cilium-centrosome complex are called ciliopathies. Meckel-Gruber syndrome (MKS) is a ciliopathic lethal autosomal recessive syndrome characterized by genetically and clinically heterogeneous manifestations, including renal cystic dysplasia, occipital encephalocele and polydactyly. Several genes have previously been associated with MKS and MKS-like phenotypes, but there are still genes remaining to be discovered. We have used whole exome sequencing (WES) to uncover the genetics of a suspected autosomal recessive Meckel syndrome phenotype in a family with two affected fetuses. RNA studies and histopathological analysis was performed for further delineation. WES lead to identification of a homozygous nonsense mutation c.256C>T (p.Arg86*) in CEP55 (centrosomal protein of 55 kDa) in the affected fetus. The variant has previously been identified in carriers in low frequencies, and segregated in the family. CEP55 is an important centrosomal protein required for the mid-body formation at cytokinesis. Our results expand the list of centrosomal proteins implicated in human ciliopathies and provide evidence for an essential role of CEP55 during embryogenesis and development of disease.