A Novel Splicing Variant in the TMC1 Gene Causes Non-Syndromic Hearing Loss in a Chinese Family
Hearing loss is the most prevalent sensory disorder globally, affecting approximately 1 in 1,000 newborns, with half of these cases attributed to genetic causes. The condition exhibits significant genetic heterogeneity, with over 110 genes implicated in non-syndromic hearing loss (NSHL). Among these, the TMC1 gene (transmembrane channel-like 1; OMIM: 606706) is notable for its role in both autosomal recessive (ARNSHL) and autosomal dominant (ADNSHL) forms of hearing loss. TMC1, located on chromosome 9q21.13, encodes a protein critical for cochlear hair cell function. Mutations in this gene disrupt auditory signal transduction, leading to sensorineural hearing loss. This study identifies a novel splicing variant in TMC1 associated with severe congenital hearing loss in a Chinese family, expanding the mutational spectrum of TMC1-related disorders.
Clinical Presentation and Diagnostic Evaluation
The proband, a 2-year-old boy, presented with congenital bilateral hearing loss. His parents, non-consanguineous and phenotypically normal, reported no family history of hearing impairment or exposure to ototoxic agents. Audiological assessments revealed profound hearing loss: auditory brainstem response (ABR) thresholds measured 85 dBnHL (left ear) and 90 dBnHL (right ear). Auditory steady-state response (ASSR) tests showed binaural thresholds ranging from 75–90 dBnHL. Distortion product otoacoustic emissions (DPOAEs) were absent bilaterally, confirming outer hair cell dysfunction. Radiological evaluations, including temporal bone computed tomography (CT) and brain magnetic resonance imaging (MRI), ruled out structural abnormalities. These findings aligned with the characteristic DFNB7/11 phenotype associated with TMC1-related ARNSHL, typically involving congenital, prelingual, severe-to-profound hearing loss without vestibular dysfunction.
Genetic Analysis and Variant Identification
Targeted high-throughput sequencing of 129 known deafness genes was performed on the proband and parents. A homozygous variant, c.2002A>G (NM_138691.3), was identified in TMC1 exon 20. Sanger sequencing confirmed the variant’s segregation: the proband was homozygous, while both parents were heterozygous carriers. The variant, absent from population databases (ExAC and gnomAD; ACMG PM2 criterion), substitutes adenine for guanine at position 2002, potentially disrupting the canonical splice site.
In Silico and Functional Validation of the Variant
Bioinformatic tools predicted significant splicing disruption. MaxEntScan analysis indicated altered splice site efficiency, while dbscSNV’s ADA (adaptive boosting) and RF (random forest) scores (>0.97) strongly supported pathogenicity. To validate these predictions, a minigene assay was designed to assess exon 20 splicing. The construct included intron 19 (197 bp), exon 20 (240 bp), intron 20 (129 bp), and universal flanking sequences (exon A–intron A; intron B–exon B). Wild-type and mutant plasmids were transfected into HEK293T cells, and RNA splicing patterns were analyzed via RT-PCR.
In wild-type constructs, two transcripts were observed: a predominant 629-bp product retaining exon 20 and a minor 389-bp product skipping exon 20. In contrast, mutant constructs produced only the 389-bp transcript, indicating complete exon 20 skipping (Figure 1E–F). This finding confirmed that c.2002A>G abolishes exon 20 inclusion, leading to a frameshift and premature termination codon. The resulting truncated protein lacks critical transmembrane domains required for mechanoelectrical transduction in cochlear hair cells.
Pathogenicity Classification and Genotype-Phenotype Correlation
The c.2002A>G variant was classified as “likely pathogenic” per ACMG/AMP guidelines. Key criteria included:
- PM2: Absence in population databases.
- PM3: Homozygosity in the proband and carrier status in parents.
- PS3: Functional evidence from the minigene assay.
- PM5: Co-location with a known pathogenic variant (c.2004T>G, p.Ser668Arg).
This variant represents the 19th TMC1 mutation reported in Chinese NSHL populations, contributing to a growing database of 120 global TMC1 variants. Most TMC1-related ARNSHL cases in China present with congenital, severe-to-profound hearing loss, consistent with the proband’s phenotype. In contrast, ADNSHL-linked TMC1 mutations (e.g., DFNA36) typically manifest as postlingual, progressive high-frequency loss, highlighting the gene’s dual role in recessive and dominant inheritance.
Broader Implications and Mechanistic Insights
TMC1 encodes a six-transmembrane domain protein essential for hair cell mechanotransduction. Exon 20 skipping due to c.2002A>G disrupts the protein’s C-terminal region, abolishing ion channel function. This mechanism aligns with prior studies showing that TMC1 truncations impair auditory neurotransmission. The minigene assay’s utility in splicing validation underscores its importance in variant classification, particularly for non-canonical splice site mutations.
Notably, phenotypic variability exists among TMC1 mutations. For example, a Dutch family with a homozygous splicing variant (c.1763+3A>G) exhibited progressive postlingual hearing loss, contrasting with the congenital presentation in this study. Such differences may arise from residual protein function in hypomorphic alleles or modifier gene effects, warranting further investigation.
Conclusion
This study identifies c.2002A>G in TMC1 as a novel pathogenic splicing variant causing ARNSHL in a Chinese family. Comprehensive clinical, genetic, and functional analyses validated its role in exon skipping and protein truncation. The findings reinforce TMC1’s prominence in hereditary hearing loss and highlight the necessity of integrating splicing assays into diagnostic pipelines. Continued discovery of TMC1 variants will enhance genetic counseling, prognostic accuracy, and therapeutic development for affected families.
DOI: https://doi.org/10.1097/CM9.0000000000001966
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