Ultrarare heterozygous pathogenic variants of genes causing dominant forms of early-onset deafness underlie severe presbycusis

Abstract

Presbycusis, or age-related hearing loss (ARHL), is a major public health issue. About half the phenotypic variance has been attributed to genetic factors. Here, we assessed the contribution to presbycusis of ultrarare pathogenic variants, considered indicative of Mendelian forms. We focused on severe presbycusis without environmental or comorbidity risk factors and studied multiplex family age-related hearing loss (mARHL) and simplex/sporadic age-related hearing loss (sARHL) cases and controls with normal hearing by whole-exome sequencing. Ultrarare variants (allele frequency [AF] < 0.0001) of 35 genes responsible for autosomal dominant early-onset forms of deafness, predicted to be pathogenic, were detected in 25.7% of mARHL and 22.7% of sARHL cases vs. 7.5% of controls (P = 0.001); half were previously unknown (AF < 0.000002). MYO6, MYO7A, PTPRQ, and TECTA variants were present in 8.9% of ARHL cases but less than 1% of controls. Evidence for a causal role of variants in presbycusis was provided by pathogenicity prediction programs, documented haploinsufficiency, three-dimensional structure/function analyses, cell biology experiments, and reported early effects. We also established Tmc1^N321I/+ mice, carrying the TMC1:p.(Asn327Ile) variant detected in an mARHL case, as a mouse model for a monogenic form of presbycusis. Deafness gene variants can thus result in a continuum of auditory phenotypes. Our findings demonstrate that the genetics of presbycusis is shaped by not only well-studied polygenic risk factors of small effect size revealed by common variants but also, ultrarare variants likely resulting in monogenic forms, thereby paving the way for treatment with emerging inner ear gene therapy.

Keywords: Tmc1; age-related hearing loss; monogenic disorder; presbycusis; ultrarare variants.

Fig. 1.

Schematic workflow for the identification of genetic variants involved in presbycusis. WES data from mARHL and sARHL cases and from controls were filtered on the basis of AF in the European population: below 5, 0.1, and 0.01%. Only the variants predicted to be pathogenic by CADD, MutationTaster, PolyPhen-2, and SIFT, with a maximal score, were selected for further analysis. Indel, insertion/deletion.

Fig. 2.

Distribution of ultrarare variants of DFNA and ADSD genes in mARHL and sARHL cases. Venn diagram of the variants (number in parentheses) present in DFNA and ADSD genes in sARHL (green) and mARHL (blue) cases. The variants in the intersection correspond to genes affected in both mARHL and sARHL cases. Variants of the genes in bold accounted for one-third of mARHL cases. *An ultrarare variant of the corresponding gene was also detected in the control group (SI Appendix, Table S5).

Fig. 3.

Functional validation of the Tmc1N321I mutation as responsible for ARHL. (A) Pedigree of the PAR093 family (dots indicate individuals included in the genetic analysis) and DNA sequencing chromatograms of unaffected (Lower Left) and affected (Lower Right) individuals showing the variant in the heterozygous state in the ARHL patient (arrow). Nucleotide positions are based on the NM_138691.2 transcript used as a reference. (B) Audiograms of PAR093 family members: individuals with ARHL (III.6 and III.7: 63 and 59 y old, respectively) and an individual with normal hearing (IV.1: 51 y old). Dashed lines show the hearing threshold defining severe ARHL in age- and sex-matched members of the general population (in black for 51-y-old women, in dark blue for 63-y-old women, and in light blue for 59-y-old men) (SI Appendix, Patients and Methods). (C) Interspecies conservation of the asparagine residue in position 327 (boxed in red) through evolution. (D) Schematic diagram of the predicted human TMC1 protein with 10 TM domains (1 to 10), according to Pan et al. (59). (E–H) Results from Tmc1 mutant mice (in blue, Tmc1^N321I/+; in purple, Tmc1^N321I/N321I; in black, Tmc1^+/+). (E) MET current amplitude of OHCs from Tmc1^+/+ (n = 17), Tmc1^N321I/+ (n = 25), and Tmc1^N321I/N321I (n = 21) mice recorded on P8. The amplitude (mean ± SD) of the MET current in response to stimulation is significantly lower in cells from Tmc1^N321I/N321I mice than in cells from Tmc1^+/+mice. (F) MET current as a function of hair bundle displacement for Tmc1^+/+, Tmc1^N321I/+, and Tmc1^N321I/N321I mice. For sensitivity measurements, the mean value (± SD) of the three-state derivative of the Boltzmann equation was calculated for displacements corresponding to P_o values between 0.2 and 0.8. The P_o(X) curves were superimposed, indicating that MET sensitivity to hair bundle displacement is unaffected in Tmc1^N321I/+ and Tmc1^N321I/N321I mutants. (G) Pure tone-evoked ABR thresholds (mean ± SD) in Tmc1^+/+, Tmc1^N321I/+, and Tmc1^N321I/N321I mice at 3 (Left) and 7 mo (Right). (H) ABR thresholds (mean ± SD) at 20 kHz (Left) and 40 kHz (Right) in mice aged P21 (Tmc1^+/+, n = 5; Tmc1^N321I/+, n = 10; Tmc1^N321I/N321I, n = 7), 3 mo (Tmc1^+/+, n = 12; Tmc1^N321I/+, n = 7; Tmc1^N321I/N321I, n = 8), and 7 mo (Tmc1^+/+, n = 7; Tmc1^N321I/+, n = 7; Tmc1^N321I/N321I, n = 6). n.s., nonsignificant; dBSPL, décibel sound pressure level. *P < 0.05; **P < 0.005; ***P < 0.0005.