Identification of a Novel de novo Splicing Mutation in Duchenne Muscular Dystrophy Gene in an Iranian Family.

Introduction: Duchenne muscular dystrophy (DMD) (NM_004006.3) is one of the most notable neuromuscular disorders of early years. The majority of DMD cases are caused by deletions or duplications in dystrophin, while point mutations are less prevalent in dystrophin abnormalities. It is a common knowledge that the severity of the disease depends on the effect of the mutation on the translational reading frame of the dystrophin mRNA. Case Report: We studied an 8-year-old boy with relevant clinical presentations for DMD. Deletion/duplication screening was performed by using multiplex ligation-dependent probe amplification, and whole-exome sequencing was conducted in order to identify potential variants. A novel de novo splice site variant was identified in the DMD gene (DMD: c.8548-2A>G). To explore the effect of a novel variant in DMD, various in silico analyses were carried out to investigate the pathogenicity of the causative variant. To study the structure of a DMD protein and information on how the genetic variant impacts splicing site in models of wild-type and mutated DMD, we carried out different computational studies. Sanger sequencing was performed for the purpose of variant confirmation and familial segregation analysis. Discussion: This novel de novo variant was predicted to have an effect on splicing, which leads to DMD due to its significant impacts on dystrophin functionality. The novel mutation would be expected to disrupt the protein structure.

Next-generation sequencing reveals a novel pathogenic variant in the ATM gene.

INTRODUCTION: Ataxia telangiectasia (A-T) is a rare autosomal recessive, multisystemic disease. Patients with the A-T syndrome present a broad spectrum of disease phenotypes. The ATM (ataxia telangiectasia mutated) gene, the only causative gene for A-T. METHOD: A patient of Persian origin presenting with typical A-T was referred to our genetics centre for specialized genetic counselling and testing. Targeted next-generation sequencing (NGS) was applied. Sanger sequencing was used to confirm the candidate variant. Modelling was performed using the SWISS-MODEL server. RESULTS: A homozygous stop-gain variant c.829G > T (p.E277*) was found in the ATM gene. This variant was confirmed by Sanger sequencing and modelling of native structure, and truncated structure was performed. CONCLUSION: To date, very few pathogenic variants of the ATM gene have been reported from the Iranian population. The finding has implications in molecular diagnostic for A-T in Iran.

A novel pathogenic variant of BRAT1 gene causes rigidity and multifocal seizure syndrome, lethal neonatal.

INTRODUCTION: Lethal neonatal rigidity and multifocal seizure syndrome (RMFSL) is a severe autosomal recessive epileptic encephalopathy characterized by microcephaly, rigidity, intractable focal seizures, apnea, and bradycardia at or soon after birth. RMFSL is related to BRCA1-associated ATM activator 1 (BRAT1) gene mutations. METHODS: An Iranian couple with history of infant death due to RMFSL was referred to our genetics lab for specialized genetic counseling and testing. Whole Exome Sequencing (WES) was applied. Following WES, Sanger sequencing was performed to confirm the candidate variant. RESULT: A novel nonsense variant (c.2041G > T, p. E681X) was identified in exon 14 of the BRAT1 gene. Based on the American College of Medical Genetics and Genomics guideline this variant was classified as a pathogenic variant. CONCLUSION: This research expands the spectrum of BRAT1 pathogenic variants in RMFSL syndrome and demonstrates the utility of WES in genetic diagnostic.

A pathogenic variant in SLC26A4 is associated with Pendred syndrome in a consanguineous Iranian family.

Objective: Hearing loss (HL) is a common sensory deficit with high phenotypic and genotypic heterogeneity. A large Iranian family with HL was genetically assessed in this study. Design: A proband from a consanguineous multiplex HL family from Iran was examined via Targeted Next-Generation Sequencing (TNGS). Sanger sequencing allowed the segregation analysis of the variant of interest and the investigation of its presence in a cohort of 50 ethnicity-matched healthy control individuals. The gene was previously associated with HL. Therefore, to determine whether the variant was specifically associated with Pendred Syndrome (PDS) or DFNB4, biochemical analyses, PTA, thyroid scans by Tc99m, perchlorate discharge test and high-resolution CT scan of the temporal bone were carried out on the affected family members. Study sample: Ten members of a large multiplex Iranian family with HL were recruited in this study. In addition, 50 unrelated healthy controls of the same ethnic group were randomly selected to genotype the variant. Results: A homozygous missense variant (NM_000441.1: c.1211C > T/p.Thr404Ile) in exon 10 was found segregating in the family. Based on the ACMG's guidelines, the variant was classified as pathogenic. Conclusion: This study expands the spectrum of SLC26A4 pathogenic variants in hearing loss.

A novel pathogenic variant in an Iranian Ataxia telangiectasia family revealed by next-generation sequencing followed by in silico analysis.

Ataxia telangiectasia (A-T) is a neurodegenerative autosomal recessive disorder with the main characteristics of progressive cerebellar degeneration, sensitivity to ionizing radiation, immunodeficiency, telangiectasia, premature aging, recurrent sinopulmonary infections, and increased risk of malignancy, especially of lymphoid origin. Ataxia Telangiectasia Mutated gene, ATM, as a causative gene for the A-T disorder, encodes the ATM protein, which plays an important role in the activation of cell-cycle checkpoints and initiation of DNA repair in response to DNA damage. Targeted next-generation sequencing (NGS) was performed on an Iranian 5-year-old boy presented with truncal and limb ataxia, telangiectasia of the eye, Hodgkin lymphoma, hyper pigmentation, total alopecia, hepatomegaly, and dysarthria. Sanger sequencing was used to confirm the candidate pathogenic variants. Computational docking was done using the HEX software to examine how this change affects the interactions of ATM with the upstream and downstream proteins. Three different variants were identified comprising two homozygous SNPs and one novel homozygous frameshift variant (c.80468047delTA, p.Thr2682ThrfsX5), which creates a stop codon in exon 57 leaving the protein truncated at its C-terminal portion. Therefore, the activation and phosphorylation of target proteins are lost. Moreover, the HEX software confirmed that the mutated protein lost its interaction with upstream and downstream proteins. The variant was classified as pathogenic based on the American College of Medical Genetics and Genomics guideline. This study expands the spectrum of ATM pathogenic variants in Iran and demonstrates the utility of targeted NGS in genetic diagnostics.