1 INTRODUCTION

Hearing loss (HL) is a prevalent sensory disorder that affects over 5% of the global population, according to the World Health Organization. In 75% of cases, HL is the only clinical finding, which is referred to as nonsyndromic hearing loss (NSHL). NSHL is typically congenital or prelingual in onset when inherited with autosomal recessive inheritance. In contrast, autosomal dominant nonsyndromic hearing loss (ADNSHL) typically has post-lingual onset and is often progressive. While ADNSHL accounts for a smaller proportion of NSHL cases, it can still have a significant impact on affected individuals and their families (Alde et al., 2023; Smith et al., 2005).

MYH14 (MIM 608568) is among the 50 genes that have been implicated in causing ADNSHL (Donaudy et al., 2004). This gene is highly expressed in the Organ of Corti, where it plays a critical role in protecting against overstimulation by acoustic signals. Loss of this protective function can result in progressive HL (Fu et al., 2016). Therefore, MYH14 is included in gene panels for clinical testing (Abou Tayoun et al., 2016; Peart et al., 2023).

To date, 86 variants reported in MYH14, at Human Gene Mutation Database (HGMD) (https://www.hgmd.cf.ac.uk/ac/index.php; last accessed:1/3/2024). Out of 86 variants, 59 of them associated with HL. Among these variants, only 13 pathogenic or likely pathogenic missense and nonsense variants have been reported in families from various regions of the world (Choi et al., 2011; Donaudy et al., 2004; Hiramatsu et al., 2021; Kim et al., 2017; Qing et al., 2014; Shearer et al., 2010; Smith et al., 2005; Yang et al., 2005). The remaining variants have been classified as conflicting or benign due to incomplete penetrance and lack of functional or segregation data. There are 1036 entries for MYH14 in ClinVar (https://www.ncbi.nlm.nih.gov/clinvar/; last accessed: 1/3/2024) and 13 of them are interpreted as pathogenic or likely pathogenic without conflict.

In this study, we identified five MYH14 variants in six families. We carefully evaluated the variants and classified them according to American College of Medical Genetics (ACMG) and Hearing Loss Expert panel (HLEP) guidelines (Oza et al., 2018; Richards et al., 2015). Our findings support the role of MYH14 in ADNSHL and broaden the spectrum of this gene as a contributor to post-lingual severe to profound deafness.

2 METHODS

3 RESULTS

We found a total of five variants in MYH14 in probands of six families after ES (Table 1) out of which three were novel. The genetic findings in all affected individuals are shown in Table S1. Pedigree analysis of families with novel MYH14 variants suggested ADNSHL (Figure 1 and Figure S1). No other clinical findings were detected in affected individuals. Audiometric re-evaluation showed stable HL in all affected individuals from family 1. All the affected individuals from this family had post-lingual, severe, nonprogressive HL (Figure S2). Family 2 had eight affected individuals, we only had mother's (II:3) and proband's (III:2) DNA. Mother had severe while the proband presented with moderate HL. We were not able to access the audiograms for the rest of the families. Family 1 and 2 have different substitutions at the same nucleotide position (c.571G > C and c.571G > A, respectively), which co-segregated with the phenotype in family 1 and 2. These two variants were classified as likely pathogenic. Moreover, the protein residue at position 191 is highly conserved among different vertebrates except two species, Scarlet macaw (Ara macao) and Spiny softshell turtle (Apalone spinifera) (Figure 1b). A novel missense variant c.5380G > A, lying in the tail domain of MYH14 was identified in a simplex family 3 and classified as VUS. We were not able to confirm this variant as a de novo since parents were not available (Figure S1). For family 4 the variant c.526G > A was confirmed by Sanger sequencing in the proband. She has moderate HL. However, there were no additional DNA samples from the family available to test segregation. In ClinVar database (http://www.ncbi.nlm.nih.gov/clinvar/) this variant is classified as variant of uncertain significance (VUS) and together with the in-silico predictions and our analyses, it still remains as VUS (Chen et al., 2016). In family 5 there were two affected individuals, and the variant c.547C > T in MYH14 co-segregated with HL (Figure S3). The similar variant was identified in the proband of family 6 along with a common mitochondrial pathogenic variant m.1555A > G.

For three novel p.Asp191His (Family 1), p.Asp191Asn (Family 2) and p.Gly1794Arg (Family 3) variants we performed the structural modeling (Figure 2). In silico analysis revealed that these changes are unfavorable for protein stability. The substitution of Histidine at position 191 produces a long protruding chain affecting the proper folding of the protein. Similarly, Glycine is a small neutral amino acid substituted for an acidic residue at position 191. It disrupts hydrogen bonding and ionic interactions with the neighboring molecules. Glycine at position 1794 is changed to arginine which is a basic amino acid thus altering the interaction with the adjacent molecules in the chain.

4 DISCUSSION AND CONCLUSIONS

In this study, we identified five MYH14 variants two of which were novel and interpreted as likely pathogenic in patients with ADNSHL. Interpretation of the autosomal dominant variants is difficult due to incomplete penetrance, lack of functional studies, and large families. To date only 13 variants have been reported in HGMD in families with more than two affected individuals. Our findings expand the knowledge of MYH14 variants associated with HL.

The likely pathogenic p.Asp191His (c.571G > C) variant in Family 1 and p.Asp191Asn (c.571G > A) variant in Family 2, affects the aspartic acid 191 residue in the motor domain of MYH14. This residue is highly conserved between species indicating its importance for the protein's function. Only two species, Scarlet macaw (Ara macao) and, Spiny softshell turtle (Apalone spinifera) has Asn instead Asp in the same residue. The in-silico structural analysis of the p.Asp191His and p.Asp191Asn variants revealed that both variants cause the disruption of the hydrogen and ionic bonding between the neighboring amino acids in the chain. As aspartic acid is polar and acidic, and histidine is polar and basic, while asparagine is polar and neutral, it is possible that the phenotype of the affected individuals with these variants may differ due to the varying physicochemical properties of the amino acids. A p. Asp191Gly variant affecting same codon has been reported in a proband and affected father as a cause of prelingual severe and progressive autosomal dominant NSHL (Kim et al., 2017).

We have identified a novel p.Gly1794Arg variant in proband of the Family 3. The variant is rare and amino acid residue is not conserved in many vertebrates (Figure S1). In the Family 4 we identified a p.Ala176Thr variant in proband (Figure S3) which was previously reported in a case with hereditary HL (Chen et al., 2016). In ClinVar database this variant reported as VUS and our interpretation remained same.

We have identified the p.Arg183Trp variant in Family 5 and 6, which was previously reported by Retterer et al (Retterer et al., 2016). In family 5, it was segregated in affected father and son with HL. In family 6 proband and unaffected father has the variant. Proband also carries m.1555A > G variant which is inherited from unaffected mother. The proband does not have aminoglycoside usage history. We have interpreted the variant p.Arg183Trp as VUS.

Some MYH14 variants may be mediated by environmental influences, while others may be tolerated due to compensatory mechanisms. In a knockout mice study by Fu et al., 2016, Myh14 knockout mice did not exhibit significant HL until 5 months of age, but outer hair cell loss was observed after acoustic trauma. The authors suggested that MYH10 can compensate for most of the functions of MYH14 in the cochlea and could explain the milder phenotypes in mice. Additionally, Myh14 was shown to play a protective role in noise-induced damage of outer hair cells, suggesting that some MYH14 variants may be more susceptible to environmental factors (Fu et al., 2016).

In conclusion, this study broadens the spectrum of MYH14 gene variants associated with ADNSHL and supports its role as a contributor to post-lingual severe to profound deafness. The identified variants can aid in genetic counseling and facilitate early diagnosis and intervention for affected individuals and their families. Further functional studies are warranted to elucidate the pathogenic mechanisms underlying MYH14-related HL.

AUTHOR CONTRIBUTIONS

D.D., M.R., G.B., and MT designed the study. A.S, A.M., L.P., S.S., K.I., B.B., M.T.K. and M.T. evaluated cases, collected the clinical data, and assisted with specimen collection. D.D., M.R., G.B., S.G. and MT analyzed the data. D.D. wrote the first draft of the manuscript. M.R., G.B., and M.T. revised the manuscript. All authors read and approved the final version of the manuscript.

FUNDING INFORMATION

This work was supported by the National Institutes of Health grants R01DC009645 and R01DC012836 to M.T and by TUBITAK (1059B191801475) to D.D. Authors declare that there are no conflicts of interest to report.

CONFLICT OF INTEREST STATEMENT

The authors declare that there are no conflicts of interest.