Autosomal Recessive Non-syndromic Hearing Loss Research Articles

A Novel Mutation Located in the N-Terminal Domain of MYO15A Caused Sensorineural Hearing Loss.

MYO15A is one of the common genes of severe-to-profound sensorineural deafness. Mutations in this gene can cause both pre- and post-lingual hearing losses. In this study, a novel MYO15A variant (c.2482C>T) was identified to be associated with autosomal recessive non-syndromic hearing loss (ARNSHL) in a Chinese Uighur family. To examine the effects of the MYO15A mutation on the morphology and function of the derived hair cell-like cells, two iPSCs were generated separately from the proband and a mutation-negative family member and those were then induced to hair cell-like cells. Results showed that this homozygous MYO15A mutation (PVS1 + PM2 + PP1 + PP3), which is located in the N-terminal domain, displayed significant differences in the morphology and function of hair cell-like cells between the proband and the normal control, although it had no effect on the totipotency of iPSCs. Our study demonstrates that the novel variant c.2482C>T in the MYO15A gene may cause inner ear hair cell dysfunction and audiological disorders in this family.

A novel frameshift variant in the TMPRSS3 gene causes nonsyndromic hearing loss in a consanguineous family

BackgroundHearing Loss (HL) is the most common sensorineural condition in humans. Mutations in the TMPRSS3 gene (DNFB8/10 locus) have been linked to autosomal recessive non-syndromic hearing loss (ARNSHL).MethodsWhole-exome sequencing (WES) was utilized to identify disease-causing variants in a proband from Iran with ARNSHL who presented clinically with sensorineural, bilateral, and prelingual HL. The pathogenicity and novelty of the identified variant were assessed using various databases. A co-segregation study was also performed to confirm the presence of the variant in the proband’s parents. Additionally, the secondary and tertiary structures of the mutant TMPRSS3 protein were predicted using bioinformatics tools. Furthermore, a global mutational spectrum of TMPRSS3 was created and statistically analyzed. The Iranome database was also used to identify other putative mutations in the TMPRSS3 gene in the Iranian population.ResultsWe identified a novel homozygous single nucleotide deletion in TMPRSS3 (c.297delA, p.Asp100ThrfsTer52) in the proband. This is the first report of this mutation in a patient with ARNSHL. Sanger sequencing confirmed that this variant co-segregated from the proband’s parents. Bioinformatic tools classified this novel variant as likely pathogenic. Additionally, 49.55% of families with TMPRSS3-related HL patients were shown to have consanguinity, consistent with our study. The Iranome database also revealed the c.268G > A variant as a putative novel mutation in TMPRSS3.ConclusionThis research expanded the pool of evidence regarding the association between mutations in the TMPRSS3 gene and ARNSHL. The finding confirmed that a single nucleotide deletion caused HL in the proband, suggesting that genetic testing, such as WES, is a robust technique for diagnosing patients with this condition.

Segregation of Trans Mutations in the CDH23 Gene in an Emirati Family with Sensorineural Hearing Loss.

Hearing loss (HL) is a significant global health concern, affecting approximately 1 in every 1000 newborns, with over half of these cases attributed to genetic factors. This study focuses on identifying the genetic basis of autosomal recessive non-syndromic hearing loss (ARNSHL) in a consanguineous Emirati family. Clinical exome sequencing (CES) was performed on affected members of the family, followed by Sanger sequencing to validate the findings. Specific primers were used for PCR amplification of target CDH23 exons. Mutations were analyzed using various computational tools to assess their pathogenicity. We identified two heterozygous mutations in the CDH23 gene: a novel nonsense variant (c.264G>A, p.Trp88Ter) and a missense variant (c.5168G>A, p.Arg1723His). Both mutations were found in trans configuration, suggesting a compound heterozygous state contributing to the phenotype. In silico analysis predicted a significant impact on protein function, potentially leading to the observed ARNSHL. This study emphasizes the complexity of genetic factors in hearing loss, particularly in highly consanguineous populations. The identification of both nonsense and missense mutations in the CDH23 gene enhances understanding of its role in hearing loss and provides essential insights for genetic counseling and future therapeutic strategies.

Molecular and Clinical Characterization of a Cohort of Autosomal Recessive Sensorineural Hearing Loss in Egyptian Patients

Hearing loss (HL) is one of the most common health problems worldwide. Autosomal recessive non-syndromic sensorineural hearing loss (ARNSHL) represents a large portion of congenital hereditary HL. Our study was conducted on 13 patients from 13 unrelated families. The majority of patients presented with congenital severe to profound bilateral sensorineural HL. All patients were subjected to detailed family history and three-generation pedigree analysis to exclude any environmental cause and to ensure an autosomal recessive mode of inheritance. Molecular analysis was performed using the whole exome sequencing (WES) technique for the recruited patients. Three variants in the MYO7A and OTOF genes were reported for the first time in patients with ARNSHL (one nonsense, one frameshift, and one splice variant). Ten previously reported variants were detected in seven genes (GJB2, MYO15A, BSND, OTOF, CDH23, SLC26A4, and TMIE). They varied between missense, nonsense, frameshift, and splice variants. This study expands the molecular spectrum of two types of autosomal recessive deafness (types 2 and 9).

A Village in the Southeastern Region of Iran Harboring the c.716T>A (p.Val239Asp) Mutation in SLC26A4.

After GJB2, SLC26A4 is the second most common contributor to autosomal recessive nonsyndromic hearing loss (ARNSHL) worldwide. In this study, we used Exome Sequencing (ES) to present a village with 31 individuals affected by hereditary hearing loss (HHL) in southeastern Iran near the border of Pakistan. The village harbored the known pathogenic missense SLC26A4 (NM_000441.2):c.716T>A (p.Val239Asp) mutation, which has a founder effect attributed to Pakistan, Iran's southeastern neighbor. Our findings, in addition to unraveling the molecular cause of non-syndromic hearing loss in these patients and further confirming the common ancestry and migration story between the people of this region and Pakistan, provide further insight into the genetic background of this region and highlight the importance of understanding the mutation spectrum of GJB2 and SLC26A4 in different regions to choose cost-effective strategies for molecular genetic testing.

Genetic analysis of TRIOBP and MYO15A variants in Iranian families with autosomal recessive non-syndromic hearing loss

BackgroundDeafness is a prevalent sensory and neurological disorder that impacts over 466 million individuals globally. Congenital deafness is the most prevalent birth defect, occurring in approximately 2–3 out of every 1000 births. It is recognized as a highly diverse condition. >50% of congenital deafness has genetic causes and the rest is due to environmental causes or both. With the development of next-generation sequencing and bioinformatics tools, whole-exome sequencing has been proposed as one of the effective methods for diagnosing genetics of hearing loss. MethodFive Iranian Autosomal recessive non-syndromic hearing loss (ARNSHL) families negative for GJB2 (NM_004004.6) gene mutations from Sistan and Baluchestan province were selected for further study by whole-exome sequencing analysis. The analysis procedure was performed using multiple bioinformatics tools and websites after filling the consent form and extracting DNA from whole blood using the salting out method. After detecting the variants in priority and confirming them in the probands by Sanger sequencing, other family members were studied to confirm the variant within the family. ResultsAfter analyzing the families recruited for this study, four known genes along with known and novel variants were discovered. Mutations found in the MYO15A, SLC26A4, TRIOBP and TECTA genes among which, the variants found in TRIOBP (NM_001039141.3, p.R283X) and MYO15A (NM_016239.4, p.P2880Rfs*19) were novel. Other known variants were TECTA (NM_005422.4, p.W1534X) and SLC26A4 (NM_000441.2, p.V239D). No genes or variants that might contribute to hearing loss have been identified within one of the families. ConclusionOur study's findings support previous research that has identified SLC26A4, MYO15A, and TECTA genes as common genetic factors following GJB2 in our population.

Heterozygous variants in transmembrane channel-like 1 gene cause autosomal recessive nonsyndromic hearing loss.

Autosomal recessive non-syndromic hearing loss (ARNSHL) can cause severe or very severe pre-speech hearing loss. Transmembrane channel-like 1 (TMC1) gene is the sixth deafness gene discovered, but the precise extent of its protein structure and function is unknown. First, history collection, audiology examination and imaging examination were performed on the proband and his family members. Peripheral blood of proband and family members was collected, genomic DNA was extracted, exon high-throughput sequencing technology was used to detect the deafness gene mutation of the proband, and Sanger sequencing was performed to verify the TMC1 gene of the proband's parents. The proband was born with hearing impairment, normal tympanic function, inability to induce acoustic reflex in both ears (acoustic reflex threshold is 100 dBHL), and severe sensorineural deafness. One of his sisters has severe sensorineural hearing loss, and neither his parents nor his other sister is hearing impaired. High-throughput sequencing of the proband identified mutations at c.741+3_741+6delAAGT (splicing) and c.884C>T (p.A295V) of the TMC1 gene, two of which were heterozygous mutations. Sanger sequencing confirmed that the c.884C > T mutation was inherited from the mother, while the c.741+3_741+6delAAGT mutation was derived from the father. Prediction of amino acid function suggested that both mutations were pathogenic mutations. In conclusion, we found a new pathogenic complex heterozygous mutation of the TMC1 gene, which enriched the mutation spectrum of the TMC1 gene and provided a basis for genetic counseling and prenatal diagnosis of ARNSHL.

Unusual phenotype in 35delG mutation: a case report

BackgroundMutations in the GJB2 gene, which encodes the protein connexin 26 and is involved in inner ear homeostasis, are identified in approximately 50% of patients with autosomal recessive nonsyndromic hearing loss, making it one of the primary causes of prelingual nonsyndromic hearing loss in various populations. The 35delG mutation, one of the most common mutations of the GJB2 gene, usually causes prelingual, bilateral mild to profound, nonprogressive sensorineural hearing loss.Case presentationWe present an unusual case of an 18-year-old Turkish female with heterozygous 35delG mutation and postlingual, profound-sloping, progressive and fluctuating unilateral sensorineural hearing loss. The phenotype is different from the usual findings.ConclusionsThe 35delG mutation causing hearing loss may not always be reflected in the phenotype as expected and therefore may have different audiologic manifestations.

Targeted Linked-Read Sequencing for Direct Haplotype Phasing of Parental GJB2/SLC26A4 Alleles: A Universal and Dependable Noninvasive Prenatal Diagnosis Method Applied to Autosomal Recessive Nonsyndromic Hearing Loss in At-Risk Families

Noninvasive prenatal diagnosis (NIPD) for autosomal recessive nonsyndromic hearing loss (ARNSHL) has been rarely reported until recent years. Additionally, the existing method can not be used for challenging genome loci (eg, copy number variations, deletions, inversions, or gene recombinants) or on families without proband genotype. This study assessed the performance of relative haplotype dosage analysis (RHDO)-based NIPD for identifying fetal genotyping in pregnancies at risk of ARNSHL. Fifty couples carrying pathogenic variants associated with ARNSHL in either GJB2 or SLC26A4 were recruited. The RHDO-based targeted linked-read sequencing combined with whole gene coverage probes was used to genotype the fetal cell-free DNA of 49 families who met the quality control standard. Fetal amniocyte samples were genotyped using invasive prenatal diagnosis (IPD) to assess the performance of NIPD. The NIPD results showed 100% (49/49) concordance with those obtained through IPD. Two families with copy number variation and recombination were also successfully identified. Sufficient specific informative single-nucleotide polymorphisms for haplotyping, as well as the fetal cell-free DNA concentration and sequencing depth, are prerequisites for RHDO-based NIPD. This method has the merits of covering the entire genes of GJB2 and SLC26A4, qualifying for copy number variation and recombination analysis with remarkable sensitivity and specificity. Therefore, it has clinical potential as an alternative to traditional IPD for ARNSHL.

Autosomal recessive non-syndromic hearing loss genes in Pakistan during the previous three decades.

Hearing loss is a clinically and genetically heterogeneous disorder, with over 148 genes and 170 loci associated with its pathogenesis. The spectrum and frequency of causal variants vary across different genetic ancestries and are more prevalent in populations that practice consanguineous marriages. Pakistan has a rich history of autosomal recessive gene discovery related to non-syndromic hearing loss. Since the first linkage analysis with a Pakistani family that led to the mapping of the DFNB1 locus on chromosome 13, 51 genes associated with this disorder have been identified in this population. Among these, 13 of the most prevalent genes, namely CDH23, CIB2, CLDN14, GJB2, HGF, MARVELD2, MYO7A, MYO15A, MSRB3, OTOF, SLC26A4, TMC1 and TMPRSS3, account for more than half of all cases of profound hearing loss, while the prevalence of other genes is less than 2% individually. In this review, we discuss the most common autosomal recessive non-syndromic hearing loss genes in Pakistani individuals as well as the genetic mapping and sequencing approaches used to discover them. Furthermore, we identified enriched gene ontology terms and common pathways involved in these 51 autosomal recessive non-syndromic hearing loss genes to gain a better understanding of the underlying mechanisms. Establishing a molecular understanding of the disorder may aid in reducing its future prevalence by enabling timely diagnostics and genetic counselling, leading to more effective clinical management and treatments of hearing loss.

A capture-based method of prenatal cell-free DNA screening for autosomal recessive non-syndromic hearing loss.

This study aimed to develop and validate a prenatal cell-free DNA (cfDNA) screening method that uses capture-based enrichment to genotype fetal autosomal recessive disorders. This method was applied in pregnancies at high risk of autosomal recessive non-syndromic hearing loss (ARNSHL) to assess its accuracy and effectiveness. This assay measured the allele counts in both white blood cell DNA and cfDNA from the blood samples of pregnant women using a capture-based next-generation sequencing method. It then applied a binomial model to infer the fetal genotypes with the maximum likelihood. Ninety-four pregnant couples that were carriers of variants of ARNSHL in GJB2 or SLC26A4 were enrolled. The fetal genotypes deduced using this screening method were compared with the results of genetic diagnosis using amniocentesis. Of the 94 couples, 65 carried more than one variant, resulting in 170 single-nucleotide polymorphism (SNP) loci to be inferred in the fetuses. Of the 170 fetal SNP genotypes, 150 (88.2%) had high confidence calls and 139 (92.7%) of these matched the genotypes obtained by amniocentesis result. Out of the remaining 20 (11.8%) cases with low-confidence calls, only 14 (70.0%) were concordant with genetic diagnosis using amniocentesis. The concordance rate was 100% for sites where the maternal genotype was wild-type homozygous. The discordance was site-biased, with each locus showing a consistent direction of discordance. Genetic diagnosis identified a total of 19 wild-type homozygotes, 46 heterozygotes, 19 compound heterozygotes, and 10 pathogenic homozygotes. This screening method correctly genotyped 81.9% (77/94) of fetuses and demonstrated a sensitivity of 89.7% and a specificity of 89.2% for correctly identifying ARNSHL. This capture-based method of prenatal screening by cfDNA demonstrated strong potential for fetal genotyping of autosomal recessive disorders.

PKHD1L1, a gene involved in the stereocilia coat, causes autosomal recessive nonsyndromic hearing loss

Identification of genes associated with nonsyndromic hearing loss is a crucial endeavor given the substantial number of individuals who remain without a diagnosis after even the most advanced genetic testing. PKHD1L1 was established as necessary for the formation of the cochlear hair-cell stereociliary coat and causes hearing loss in mice and zebrafish when mutated. We sought to determine if biallelic variants in PKHD1L1 also cause hearing loss in humans. Exome sequencing was performed on DNA of four families segregating autosomal recessive nonsyndromic sensorineural hearing loss. Compound heterozygous p.[(Gly129Ser)];p.[(Gly1314Val)] and p.[(Gly605Arg)];p[(Leu2818TyrfsTer5)], homozygous missense p.(His2479Gln) and nonsense p.(Arg3381Ter) variants were identified in PKHD1L1 that were predicted to be damaging using in silico pathogenicity prediction methods. In vitro functional analysis of two missense variants was performed using purified recombinant PKHD1L1 protein fragments. We then evaluated protein thermodynamic stability with and without the missense variants found in one of the families and performed a minigene splicing assay for another variant. In silico molecular modeling using AlphaFold2 and protein sequence alignment analysis were carried out to further explore potential variant effects on structure. In vitro functional assessment indicated that both engineered PKHD1L1 p.(Gly129Ser) and p.(Gly1314Val) mutant constructs significantly reduced the folding and structural stabilities of the expressed protein fragments, providing further evidence to support pathogenicity of these variants. Minigene assay of the c.1813G>A p.(Gly605Arg) variant, located at the boundary of exon 17, revealed exon skipping leading to an in-frame deletion of 48 amino acids. In silico molecular modeling exposed key structural features that might suggest PKHD1L1 protein destabilization. Multiple lines of evidence collectively associate PKHD1L1 with nonsyndromic mild–moderate to severe sensorineural hearing loss. PKHD1L1 testing in individuals with mild–moderate hearing loss may identify further affected families.

Mutation Detection in MYO15A Gene in an Iranian Family with Non-Syndromic Hearing Loss

Hearing loss, recognized as one of the most prevalent sensory disorders, encompasses both syndromic and non-syndromic manifestations, with the identification of 87 genes and over 100 genetic loci in autosomal recessive non-syndromic hearing loss marking significant progress in understanding its genetic basis. In this case report, we showcase a non-syndromic hearing loss scenario involving a 21-year-old man experiencing progressive hearing loss. Through whole-exome sequencing, we unveiled a previously unreported homozygous mutation, c.1178_1179delAC; p.Tyr393Serfs*38, located in exon 2 (NM_016239.4) of the MYO15A gene in the proband. The newly identified mutation, causing a new reading frame (p.Tyr393Serfs*38), results in an early encounter with a stop codon, leading to the formation of a shortened protein. These findings advance our understanding of the molecular mechanisms involved in autosomal recessive non-syndromic hearing loss, contributing to broader scientific knowledge and potential breakthroughs in hearing loss research.

Genetic heterogeneity in hereditary hearing loss: Potential role of kinociliary protein TOGARAM2.

Hearing loss (HL) is a heterogenous trait with pathogenic variants in more than 200 genes that have been discovered in studies involving small and large HL families. Over one-third of families with hereditary HL remain etiologically undiagnosed after screening for mutations in the recognized genes. Genetic heterogeneity complicates the analysis in multiplex families where variants in more than one gene can be causal in different individuals even in the same sibship. We employed exome or genome sequencing in at least two affected individuals with congenital or prelingual-onset, severe to profound, non-syndromic, bilateral sensorineural HL from four multiplex families. Bioinformatic analysis was performed to identify variants in known and candidate deafness genes. Our results show that in these four families, variants in a single HL gene do not explain HL in all affected family members, and variants in another known or candidate HL gene were detected to clarify HL in the entire family. We also present a variant in TOGARAM2 as a potential cause underlying autosomal recessive non-syndromic HL by showing its presence in a family with HL, its expression in the cochlea and the localization of the protein to cochlear hair cells. Conclusively, analyzing all affected family members separately can serve as a good source for the identification of variants in known and novel candidate genes for HL.

Whole Exome Sequencing of Non-Syndromic Hearing Loss Patients.

Hearing loss is the second most common disease after mental retardation in Iran. Autosomal recessive non-syndromic hearing loss (ARNSHL) is an extreme and highly heterogeneous disease, for which more than 70 genes have been identified. Considering the frequency of family marriage as well as the importance of ARNSHL in Iran, we evaluated the genetic factors involved in this type of deafness. We performed the whole exome sequencing (WES) of eight Iranian subjects with severe nonsyndromic hearing loss selected from 110 well-characterized subjects with non-syndromic hearing loss from 2017-2019. The patients with mutated GJB2 and GJB6 genes were excluded from the study. The use of the whole exome sequencing method revealed 10 different mutations in 7 genes, including SLC26A4 (c.1234G>T), FGF3 (c.45DelC, c.466T>C), ADGRV1 (c.12528-2A>C, c.16226-16227insAGTC), OTOG (c.7454delG), OTOF (c.3570+2T>C), ESPN (c.992G>A), OTOA (c.2359G>T, c.2353A>C). Seven new variants were observed in seven families including SLC26A4 (c.1234G>T), FGF3 (c.45DelC), ADGRV1 (c.12528-2A>C), OTOG (c.7454delG), ADGRV1 (c.16226-16227insAGTC), OTOF (c.3570+2T>C). The causal mutation of ARNSHL was found in all patients using the WES. Meta-analysis studies can help to identify common mutations causing deafness in any population to facilitate identification of carriers and subjects with deafness.

Unraveling the Diversity of GJB2 Mutations in Nonsyndromic Hearing Loss: A Comprehensive Study in the Moroccan Population

Introduction: Despite the high genetic heterogeneity of hearing loss, mutations in the GJB2 gene are a major cause of autosomal recessive nonsyndromic hearing loss (NSHL) worldwide. However, the mutation profile of GJB2 in NSHL is under-investigated in Morocco, especially among simplex cases. This study aimed to identify the spectrum and frequency of GJB2 mutations in the Moroccan population among simplex and multiplex families with NSHL. Methods: Moroccan families with NSHL were selected according to well-defined criteria. Selected families were screened for GJB2 gene variants using direct sequencing of the entire coding region of GJB2. Results: A total of 145 affected individuals from 115 families with NSHL were included in this study (49 simplex, 66 multiplex). Mutations in the GJB2 gene were noted in 28.69% of the families (33/115), of which 75.75% were multiplex families and 24.24% were simplex. In total, seven different mutations were detected: c.35delG(p.G12fs), c.551G>A(p.R184Q), c.139G>T(p.E47X), c.109G>A(p.V37I), c.167delT(p.L56fs), c.617A>G(p.N206S), c.94C>T(p.R32C). The last three mutations have not previously been reported in Morocco. The most common GJB2 mutation was c.35delG (21.73%), followed by p.V37I (2.60%) and p.E47X (1.73%). Conclusions: Our study confirms a high prevalence of GJB2 variants in the Moroccan population, particularly the c.35delG mutation. Additionally, we have identified previously unreported or rarely reported mutations, revealing a greater diversity of GJB2 mutations. These findings emphasize the importance of comprehensive screening beyond the 35delG mutation for patients with NSHL, regardless of their family history. Integrating this approach into clinical care will enhance diagnosis and management of hearing loss in the Moroccan population.

Novel cis compound heterozygous variants in MYO6 causes early onset of non-syndromic hearing loss in a Chinese family.

Background: Mutations in the MYO6 gene have been associated with both autosomal dominant non-syndromic hearing loss (ADNSHL) and autosomal recessive non-syndromic hearing loss (ARNSHL), with a cumulative identification of 125 pathogenic variants. To investigate the underlying genetic factor within a Chinese family affected with heriditary hearing loss, prompted the utilization of high-throughput sequencing. Method: A detailed clinical investigation was performed. Genetic testing was performed by using target panel sequencing, and Sanger sequencing. Targeted sequencing identified the variants and Sanger sequencing was employed to validate segregation of the identified variants within family. Additionally, bioinformatics analysis was performed to strengthen our findings. Results: Clinical investigation revealed the family members were affected by progressive and sensorineural hearing loss with an onset around 8-10 years old. Furthermore, genetic testing identified novel MYO6 variants, c.[2377T>G; 2382G>T] p.[Trp793Gly; Lys794Asn], positioned in a cis pattern, as plausible pathogenic contributors to early-onset hearing loss characterized by a severe and progressive course. Moreover, bioinformatics analysis showd disruptin in hydrogen bonding of mutant amino acids with interactive amino acids. Conclusion: Our research uncovered a relationship between mutations in the MYO6 gene and non-syndromic hearing loss. We identified two variants, c.[2377T>G; 2382G>T] p.[Trp793Gly; Lys794Asn] in MYO6 as strong candidates responsible for the observed progressive hereditary hearing loss. This study not only adds to our knowledge about hearing problems related to MYO6 but also reveals the presence of monogenic compound heterozygosity. Our study will provide a new sight for genetic diagnosis in such patients and their management for future use.

Novel compound heterozygous MYO15A splicing variants in autosomal recessive non-syndromic hearing loss.

Hereditary hearing loss is a highly heterogeneous disorder. This study aimed to identify the genetic cause of a Chinese family with autosomal recessive non-syndromic sensorineural hearing loss (ARNSHL). Clinical information and peripheral blood samples were collected from the proband and its parents. Two-step high-throughput next-generation sequencing on the Ion Torrent platform was applied to detect variants as follows. First, long-range PCR was performed to amplify all the regions of the GJB2, GJB3, SLC26A4, and MT-RNR1 genes, followed by next-generation sequencing. If no candidate pathogenetic variants were found, the targeted exon sequencing with AmpliSeq technology was employed to examine another 64 deafness-associated genes. Sanger sequencing was used to identify variants and the lineage co-segregation. The splicing of the MYO15A gene was assessed by in silico bioinformatics prediction and minigene assays. Two candidate MYO15A gene (OMIM, #602,666) heterozygous splicing variants, NG_011634.2 (NM_016239.3): c.6177 + 1G > T and c.9690 + 1G > A, were identified in the proband, and these two variants were both annotated as pathogenic according to the American College of Medical Genetics and Genomics (ACMG) guidelines. Further bioinformatic analysis predicted that the c.6177 + 1G > T variant might cause exon skipping and that the c.9690 + 1G > A variant might activate a cryptic splicing donor site in the downstream intronic region. An in vitro minigene assay confirmed the above predictions. We identified a compound heterozygous splicing variant in the MYO15A gene in a Han Chinese family with ARNSHL. Our results broaden the spectrum of MYO15A variants, potentially benefiting the early diagnosis, prevention, and treatment of the disease.

Novel autosomal dominant TMC1 variants linked to hearing loss: insight into protein-lipid interactions

BackgroundTMC1, which encodes transmembrane channel-like protein 1, forms the mechanoelectrical transduction (MET) channel in auditory hair cells, necessary for auditory function. TMC1 variants are known to cause autosomal dominant (DFNA36) and autosomal recessive (DFNB7/11) non-syndromic hearing loss, but only a handful of TMC1 variants underlying DFNA36 have been reported, hampering analysis of genotype-phenotype correlations.MethodsIn this study, we retrospectively reviewed 338 probands in an in-house database of genetic hearing loss, evaluating the clinical phenotypes and genotypes of novel TMC1 variants associated with DFNA36. To analyze the structural impact of these variants, we generated two structural models of human TMC1, utilizing the Cryo-EM structure of C. elegans TMC1 as a template and AlphaFold protein structure database. Specifically, the lipid bilayer-embedded protein database was used to construct membrane-embedded models of TMC1. We then examined the effect of TMC1 variants on intramolecular interactions and predicted their potential pathogenicity.ResultsWe identified two novel TMC1 variants related to DFNA36 (c.1256T > C:p.Phe419Ser and c.1444T > C:p.Trp482Arg). The affected subjects had bilateral, moderate, late-onset, progressive sensorineural hearing loss with a down-sloping configuration. The Phe419 residue located in the transmembrane domain 4 of TMC1 faces outward towards the channel pore and is in close proximity to the hydrophobic tail of the lipid bilayer. The non-polar-to-polar variant (p.Phe419Ser) alters the hydrophobicity in the membrane, compromising protein-lipid interactions. On the other hand, the Trp482 residue located in the extracellular linker region between transmembrane domains 5 and 6 is anchored to the membrane interfaces via its aromatic rings, mediating several molecular interactions that stabilize the structure of TMC1. This type of aromatic ring-based anchoring is also observed in homologous transmembrane proteins such as OSCA1.2. Conversely, the substitution of Trp with Arg (Trp482Arg) disrupts the cation-π interaction with phospholipids located in the outer leaflet of the phospholipid bilayer, destabilizing protein-lipid interactions. Additionally, Trp482Arg collapses the CH-π interaction between Trp482 and Pro511, possibly reducing the overall stability of the protein. In parallel with the molecular modeling, the two mutants degraded significantly faster compared to the wild-type protein, compromising protein stability.ConclusionsThis results expand the genetic spectrum of disease-causing TMC1 variants related to DFNA36 and provide insight into TMC1 transmembrane protein-lipid interactions.

Homozygous Missense Variants in FOXI1 and TMPRSS3 Genes Associated with Non-syndromic Deafness in Moroccan Families.

One of the most prevalent sensorineural disorders, autosomal recessive non-syndromic hearing loss (ARNSHL) which can affect all age groups, from the newborn (congenital) to the elderly (presbycusis). Important etiologic, phenotypic, and genotypic factors can cause deafness. So far, the high genetic variability that explains deafness makes molecular diagnosis challenging. In Morocco, the GJB2 gene is the primary cause of non-syndromic hereditary deafness, while the existence of a variant in the LRTOMT gene is the second cause of this condition. After excluding these two frequently occurring GJB2 and LRTOMT variants, whole-exome sequencing was carried out in two Moroccan consanguineous families with hearing loss. As a result, two novel variants in the TMPRSS3 (c.1078G>A, p. Ala 360Thr) and FOXI1 (c.6C>G, p. Ser 2Arg) genes have been discovered in deaf patients and the pathogenic effect has been anticipated by several bioinformatics and molecular modeling systems. For the first time, these variants are identified in the Moroccan population, showing the population heterogeneity and demonstrating the value of the WES in hearing loss diagnosis.