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Tmc proteins are essential for zebrafish hearing where Tmc1 is not obligatory.

ABSTRACT: Perception of sound is initiated by mechanically gated ion channels at the tips of stereocilia. Mature mammalian auditory hair cells require transmembrane channel-like 1 (TMC1) for mechanotransduction, and mutations of the cognate genetic sequences result in dominant or recessive heritable deafness forms in humans and mice. In contrast, zebrafish lateral line hair cells, which detect water motion, require Tmc2a and Tmc2b. Here, we use standard and multiplex genome editing in conjunction with functional and behavioral assays to determine the reliance of zebrafish hearing and vestibular organs on Tmc proteins. Surprisingly, our approach using multiple mutant alleles demonstrates that hearing in zebrafish is not dependent on Tmc1, nor is it fully dependent on Tmc2a and Tmc2b. Hearing however is absent in triple-mutant zebrafish that lack Tmc1, Tmc2a and Tmc2b. These outcomes reveal a striking resemblance of Tmc protein reliance in the vestibular sensory epithelia of mammals to the maculae of zebrafish. Moreover, our findings disclose a logic of Tmc use where hearing depends on a complement of Tmc proteins beyond those employed to sense water motion.

SUBMITTER: Chen Z

PROVIDER: S-EPMC7390940 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Tmc proteins are essential for zebrafish hearing where Tmc1 is not obligatory.

Chen Zongwei Z Zhu Shaoyuan S Kindig Kayla K Wang Shengxuan S Chou Shih-Wei SW Davis Robin Woods RW Dercoli Michael R MR Weaver Hannah H Stepanyan Ruben R McDermott Brian M BM

Human molecular genetics 20200701 12

Perception of sound is initiated by mechanically gated ion channels at the tips of stereocilia. Mature mammalian auditory hair cells require transmembrane channel-like 1 (TMC1) for mechanotransduction, and mutations of the cognate genetic sequences result in dominant or recessive heritable deafness forms in humans and mice. In contrast, zebrafish lateral line hair cells, which detect water motion, require Tmc2a and Tmc2b. Here, we use standard and multiplex genome editing in conjunction with fun ...[more]

PMID: 32167554

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Project description:Variants in TMC1 (transmembrane channel-like 1) can cause both autosomal dominant and recessive hearing loss in human population. Mice with Tmc1 variants have been shown to be ideal animal models for gene therapy. In this article, we report four TMC1 variants in four different Chinese families and the follow-up auditory phenotype of a previously reported family. Four families with TMC1 variants, as well as a previously described family with TMC1 variant orthologous to the Beethoven mouse, were recruited in this study. A comprehensive auditory evaluation was performed on all ascertained family members. High-throughput sequencing was conducted using genomic DNA from the probands and other family members to identify probable deafness genes. We identified four TMC1 (NM_138691.2) variations, including two pathogenic variants, c.1714G>A, and c.1253T>A, one likely pathogenic variant, c.[797T>C];[797T>C], and one single nucleotide polymorphism (SNP), c.2276G>A. Among these variants, c.[797T>C];[797T>C] is a novel likely pathogenic variant, and c.1714G>A and c.1253T>A are known pathogenic variants at the DFNB7/11 (DFNA36) locus. Phenotype-genotype correlation analysis of TMC1 variants showed that the TMC1 dominant variation-related phenotype was late-onset, progressive, high frequency to all frequency sensorineural hearing loss, while the TMC1 recessive variant was related to congenital all frequency sensorineural hearing impairment. Two pathogenic, one likely pathogenic variants and one SNP of TMC1 were identified in four Chinese families with hereditary hearing loss, indicating that TMC1 may be a more frequent cause of hearing loss than expected. TMC1 variants related to hearing loss result in specific phenotypes. The TMC1 c.1253T>A (p.M418K) variation, homologous to the Tmc1 c. 1235 T> A (p.M412K) variant in Beethoven mice, was the second report of this variant in human patients with hearing loss, suggesting the possibility to translational gene therapy from Beethoven mice to human patients.

Project description: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.

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Tmc proteins are essential for zebrafish hearing where Tmc1 is not obligatory.

Publications

Tmc proteins are essential for zebrafish hearing where Tmc1 is not obligatory.

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