Identifying new genes responsible for non-syndromic hearing loss remains a critical goal, as many individuals with hereditary deafness still lack a molecular diagnosis despite comprehensive genetic testing. The tectorial membrane (TM) is a specialized, collagen-rich, acellular matrix of the inner ear, essential for stimulating mechanosensitive hair cell bundles during sound transduction, and its structural integrity is critical for frequency tuning and auditory sensitivity. Although mutations in genes encoding a number of non-collagenous proteins found in the TM (TECTA, CEACAM16, OTOG, OTOGL) have been identified as deafness genes, definitive evidence implicating β-tectorin (TECTB) in human hearing loss has been lacking.

Here, we present multiple lines of genetic and experimental evidence linking a missense variant in TECTB (c.674G>A, p.Cys225Tyr) to autosomal dominant, non-syndromic hearing loss in a multigenerational family. The variant alters one of eight highly conserved cysteines present within the zona pellucida (ZP) domain of TECTB and is predicted to disrupt protein folding and matrix assembly. Using a Tectb-C225Y knock-in mouse model, we show that homozygous animals exhibit severe hearing loss and profound disruption of TM morphology, while heterozygote animals display decreased matrix content within the TM and increased susceptibility to noise-induced hearing loss—despite normal auditory thresholds.

These findings identify TECTB as a novel human deafness gene, further elucidate its structural role in maintaining TM integrity, and highlight its contribution to resilience against environmental and age-related auditory decline.

The authors have declared no competing interest.

This work was supported by NIH NIDCD R01DC020190, R01DC017166 and R01DC021795 to A.A.I., NIH NIDCD F31DC021855 and the Speech and Speech and Hearing Bioscience and Technology Program Training grant T32 DC000038 to E.H.; B.V. is supported by the German Research Foundation (DFG) VO 2138/7-1 grant 469177153, via the DFG Heisenberg program VO 2138/8-1 grant 543719215, and the DFG Collaborative Research Center 1690: Disease Mechanisms and Functional Restoration of Sensory and Motor Systems (Project A03). B.V. is a member of the European Reference Network on Rare Congenital Malformations and Rare Intellectual Disability (ERN-ITHACA) [EU Framework Partnership Agreement ID: 3HP-HP-FPA ERN-01-2016/739516]. RG and GR were supported by grants from the Biotechnology and Biological Sciences Research Council (BBSRC) (BB/T016337/1) and a RNID Translational Research in Hearing grant (T6) during the course of this work. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Human studies were approved by the institutional review board of Dartmouth-Hitchcock Medical Center (No. 22289) and the Nicaraguan Ministry of Health.

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The source data used to generate the graphs in this manuscript are available in the supplementary data file. Further imaging data that support the findings of this study are available from the corresponding authors upon request.