A novel nonsense mutation in MYO6 is associated with progressive nonsyndromic hearing loss in a Danish DFNA22 family

Autosomal dominant inheritance is described in about 20% of all nonsyndromic hearing loss with currently 54 distinct loci (DFNA1-54), and >20 different genes identified. Seven different unconventional myosin genes are involved in ten different types of syndromic and nonsyndromic hearing loss with different patterns of inheritance: MYO7A in DFNA11/DFNB2/USH1B, MYH9 in DFNA17, MYH14 in DFNA4, MYO6 in DFNA22/DFNB37, MYO3A in DFNB30, MYO1A in DFNA48, and MYO15A in DFNB3. Two missense mutations in MYO6 (p.C442Y and p.H246R) have been characterized in families of Italian and American Caucasian extraction with autosomal dominant hearing loss, respectively, and the latter was associated with cardiomyopathy in some patients. Three Pakistani families had homozygosity for three MYO6 mutations (c.36insT, p.R1166X, and p.E216V, respectively), and was in one instance associated with retinal degeneration. In the present study, we linked autosomal dominant hearing loss in a large Danish family to a 38.9 Mb interval overlapping with the DFNA22/DFNB37 locus on chromosome 6q13. A novel nonsense mutation in MYO6 exon 25 (c.2545C > T; p.R849X) was identified in the family. The mutation co-segregated with the disease and the mutant allele is predicted to encode a truncated protein lacking the coiled-coil and globular tail domains. These domains are hypothesized to be essential for targeting myosin VI to its cellular compartments. No other system was involved indicating nonsyndromic loss. In conclusion, a novel nonsense MYO6 mutation causes post-lingual, slowly progressive autosomal dominant nonsyndromic moderate to severe hearing loss in a Danish family.     

Variable hearing impairment in a DFNB2 family with a novel MYO7A missense mutation

Hildebrand MS, Thorne NP, Bromhead CJ, Kahrizi K, Webster JA, Fattahi Z, Bataejad M, Kimberling WJ, Stephan D, Najmabadi H, Bahlo M, Smith RJH. Variable hearing impairment in a DFNB2 family with a novel MYO7A missense mutation. Myosin VIIA mutations have been associated with non‐syndromic hearing loss (DFNB2; DFNA11) and Usher syndrome type 1B (USH1B). We report clinical and genetic analyses of a consanguineous Iranian family segregating autosomal recessive non‐syndromic hearing loss (ARNSHL). The hearing impairment was mapped to the DFNB2 locus using Affymetrix 50K GeneChips; direct sequencing of the MYO7A gene was completed. The Iranian family (L‐1419) was shown to segregate a novel homozygous missense mutation (c.1184G>A) that results in a p.R395H amino acid substitution in the motor domain of the myosin VIIA protein. As one affected family member had significantly less severe hearing loss, we used a candidate approach to search for a genetic modifier. This novel MYO7A mutation is the first reported to cause DFNB2 in the Iranian population and this DFNB2 family is the first to be associated with a potential modifier. The absence of vestibular and retinal defects, and less severe low frequency hearing loss, is consistent with the phenotype of a recently reported Pakistani DFNB2 family. Thus, we conclude this family has non‐syndromic hearing loss (DFNB2) rather than USH1B, providing further evidence that these two diseases represent discrete disorders.     

Whole-exome sequencing reveals diverse modes of inheritance in sporadic mild to moderate sensorineural hearing loss in a pediatric population

PurposeThis study was designed to delineate genetic contributions, if any, to sporadic forms of mild to moderate sensorineural hearing loss (SNHL) not related to GJB2 mutations (DFNB1) in a pediatric population.MethodsWe recruited 11 non-DFNB1 simplex cases of mild to moderate SNHL in children. We applied whole-exome sequencing to all 11 probands. We used a filtering strategy assuming that de novo variants of known autosomal dominant (AD) deafness genes, biallelic mutations in autosomal recessive (AR) genes, monoallelic mutations in X chromosome genes for males, and digenic inheritance could be associated. Candidate variants first were prioritized with allele frequency in public databases and confirmed by a phase or a segregation test in each family. Additional information from the literature or public databases was used to identify strong candidate variants.ResultsStrong candidate variants were detected in 5 of 11 probands (45.4%). A diverse mode of inheritance implicated the sporadic occurrence of the phenotype. AR mutations in OTOGL and SERPINB6 and digenic inheritance involving two deafness genes, GPR98 and PDZ7, were detected. A de novo AD mutation also was detected in TECTA and MYH14. No syndromic feature was detected in individuals with GPR98/PDZ7 or MYH14 variants in our cohort at this moment.ConclusionMild to moderate pediatric SNHL, even if sporadic, features a strong genetic etiology and can manifest via diverse modes of inheritance. In addition, a multidisciplinary approach should be used for a correct diagnosis.Genet Med17 11, 901–911.     

MYO15A (DFNB3) mutations in Turkish hearing loss families and functional modeling of a novel motor domain mutation

Myosin XVA is an unconventional myosin which has been implicated in autosomal recessive nonsyndromic hearing impairment (ARNSHI) in humans. In Myo15A mouse models, vestibular dysfunction accompanies the autosomal recessive hearing loss. Genomewide homozygosity mapping and subsequent fine mapping in two Turkish families with ARNSHI revealed significant linkage to a critical interval harboring a known deafness gene MYO15A on chromosome 17p13.1-17q11.2. Subsequent sequencing of the MYO15A gene led to the identification of a novel missense mutation, c.5492G-->T (p.Gly1831Val) and a novel splice site mutation, c.8968-1G-->C. These mutations were not detected in additional 64 unrelated ARNSHI index patients and in 230 Turkish control chromosomes. Gly1831 is a conserved residue located in the motor domains of the different classes of myosins of different species. Molecular modeling of the motor head domain of the human myosin XVa protein suggests that the Gly1831Val mutation inhibits the powerstroke by reducing backbone flexibility and weakening the hydrophobic interactions necessary for signal transmission to the converter domain.     

Genetic causes of moderate to severe hearing loss point to modifiers

The genetic underpinnings of recessively inherited moderate to severe sensorineural hearing loss are not well understood, despite its higher prevalence in comparison to profound deafness. We recruited 92 consanguineous families segregating stable or progressive, recessively inherited moderate or severe hearing loss. We utilized homozygosity mapping, Sanger sequencing, targeted capture of known deafness genes with massively parallel sequencing and whole exome sequencing to identify the molecular basis of hearing loss in these families. Variants of the known deafness genes were found in 69% of the participating families with the SLC26A4, GJB2, MYO15A, TMC1, TMPRSS3, OTOF, MYO7A and CLDN14 genes together accounting for hearing loss in 54% of the families. We identified 20 reported and 21 novel variants in 21 known deafness genes; 16 of the 20 reported variants, previously associated with stable, profound deafness were associated with moderate to severe or progressive hearing loss in our families. These data point to a prominent role for genetic background, environmental factors or both as modifiers of human hearing loss severity.     

A splice-site mutation and overexpression of MYO6 cause a similar phenotype in two families with autosomal dominant hearing loss

Hearing loss is the most common sensory disorder, affecting 1 in 650 newborns. Linkage analysis revealed linkage to locus DFNA22 in two Belgian families 1 and 2 with autosomal dominant sensorineural hearing loss. As MYO6 has previously been reported as responsible for the hearing loss at loci DFNA22 and DFNB37, respectively, DNA sequencing of the coding region and the promoter of MYO6 was performed but this analysis did not reveal any mutations. However, only in patients of family 2, an insertion of 108 bp was identified in the mRNA of the gene. The inserted fragment was part of intron 23 and sequencing of this intron revealed a new splice-site mutation c.IVS23+2321T>G, segregating with the hearing loss in the family. The mutation causes a frameshift and a premature termination codon, but real-time PCR revealed that only 15-20% of the mRNA is degraded by nonsense-mediated decay, while the other part may give rise to an aberrant protein. In family 1, a quantitative real-time PCR experiment revealed a 1.5-1.8-fold overexpression of MYO6 in patients compared to controls. The possible presence of a gene duplication could be excluded by real-time PCR on genomic level. Most likely, the overexpression is caused by a mutation in an unidentified regulatory region of the gene. This study indicates that the inner ear hair cells are sensitive to changes in expression levels of MYO6.     

SNaPshot reveals high mutation and carrier frequencies of 15 common hearing loss mutants in a Chinese newborn cohort

Genetic causes account for more than half of congenital hearing loss cases. The most frequent mutations found in non‐syndromic hearing loss patients occur in GJB2 and SLC26A4. Mitochondrial genome mutations are also prevalent. However, the frequency of common hearing loss mutations in the Chinese population has not yet been well estimated. Here, we implemented the SNaPshot genotyping method to investigate the carrier frequency of 15 commonly reported hearing loss mutations in GJB2, SLC26A4 and the mitochondrial genome based on a cohort of 5800 neonates in China. Up to 15.9% (923/5800) of the newborns carry at least one mutant allele. The top three were GJB2‐c.109G>A, GJB2‐c.235delC, and SLC26A4‐c.919A>G, with notably high carrier frequencies of 1/10, 1/53 and 1/62 respectively, and mt‐7444G>A with 1/141 was the most frequent allele in the mitochondrial genome. In this cohort, 0.48% (28/5800) of neonates were genetically diagnosed with hearing loss, from which seven cases failed an OAE test. This is the first epidemiological study of non‐syndromic hearing loss in Chinese newborns indicating a notably high carrier frequency (1 per 6.3 newborns) among these 15 mutant alleles. Our carrier frequency data also aid in effective risk assessment and genetic counseling for hearing loss patients in the Chinese population.     

A synonymous variant in MYO15A enriched in the Ashkenazi Jewish population causes autosomal recessive hearing loss due to abnormal splicing

Nonsyndromic hearing loss is genetically heterogeneous. Despite comprehensive genetic testing, many cases remain unsolved because the clinical significance of identified variants is uncertain or because biallelic pathogenic variants are not identified for presumed autosomal recessive cases. Common synonymous variants are often disregarded. Determining the pathogenicity of synonymous variants may improve genetic diagnosis. We report a synonymous variant c.9861 C > T/p.(Gly3287=) in MYO15A in homozygosity or compound heterozygosity with another pathogenic or likely pathogenic MYO15A variant in 10 unrelated families with nonsyndromic sensorineural hearing loss. Biallelic variants in MYO15A were identified in 21 affected and were absent in 22 unaffected siblings. A mini-gene assay confirms that the synonymous variant leads to abnormal splicing. The variant is enriched in the Ashkenazi Jewish population. Individuals carrying biallelic variants involving c.9861 C > T often exhibit progressive post-lingual hearing loss distinct from the congenital profound deafness typically associated with biallelic loss-of-function MYO15A variants. This study establishes the pathogenicity of the c.9861 C > T variant in MYO15A and expands the phenotypic spectrum of MYO15A-related hearing loss. Our work also highlights the importance of multicenter collaboration and data sharing to establish the pathogenicity of a relatively common synonymous variant for improved diagnosis and management of hearing loss.Subject terms: Genetic testing, Genetic testing     

A novel missense mutation in the ESRRB gene causes DFNB35 hearing loss in a Tunisian family

Autosomal recessive non-syndromic hearing loss (ARNSHL) is a genetically heterogenous disorder with 41 genes so far identified. Among these genes, ESRRB whose mutations are responsible for DFNB35 hearing loss in Pakistani and Turkish families. This gene encodes the estrogen-related receptor beta. In this study, we report a novel mutation (p.Y305H) in the ESRRB gene in a Tunisian family with ARNSHL. This mutation was not detected in 100 healthy individuals. Molecular modeling showed that the p.Y305H mutation is likely to alter the conformation of the ligand binding-site by destabilizing the coactivator binding pocket. Interestingly, this ligand-binding domain of the ESRRB protein has been affected in 5 out of 6 mutations causing DFNB35 hearing loss. Using linkage and DHPLC analysis, no more mutations were detected in the ESRRB gene in other 127 Tunisian families with ARNSHL indicating that DFNB35 is most likely to be a rare type of ARNSHL in the Tunisian population.     

Novel heterozygous nonsense mutation of the OPTN gene segregating in a Danish family with ALS

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder. About 10% of ALS cases are familial (FALS) and the genetic defect is known only in approximately 20%-30% of these cases. The most common genetic cause of ALS is SOD1 (superoxide dismutase 1) mutation. Very recently, mutations of the optineurin gene (OPTN), which is involved in open-angle glaucoma, were identified in 3 Japanese patients/families with ALS, and subsequently in a few FALS patients of European descent. We found a heterozygous nonsense mutation (c.493C>T, p.Gln165X, exon 6) in the OPTN gene in a Danish patient with ALS, and the mutation segregated from his affected father. The p.Gln165X mutation could not be detected in 1070 healthy Danish controls, in 1000 Danish individuals with metabolic phenotypes or in 64 sporadic ALS (SALS) cases. The p.Gln165X mutation described in this study is the first mutation reported in a Danish family and is likely involved in disease pathogenesis. Until now, only few OPTN mutations have been associated with ALS. As the underlying genetic defect is known only in approximately 20%-30% of FALS families, further screening of these cases is necessary for establishing the contribution of OPTN mutations in disease pathogenesis.     

A novel DFNA5 mutation does not cause hearing loss in an Iranian family

Mutations in DFNA5 lead to autosomal dominant non-syndromic sensorineural hearing loss that starts at the high frequencies. To date, only three DFNA5 mutations have been described, and although different at the genomic DNA level, all lead to exon 8 skipping at the mRNA level. This remarkable fact has led towards the hypothesis that DFNA5-associated hearing loss is caused by a gain-of-function mutation and not by haplo-insufficiency as previously thought. Here, we describe a fourth DFNA5 mutation: the insertion of a cytosine at nucleotide position 640 (AF073308.1:_c.640insC, AAC69324.1:_p. Thr215HisfsX8). Unlike the previously described mutations, this frameshift mutation truncates the protein in exon 5 of the gene. Although the mutation was found in an extended Iranian family with hereditary hearing loss, it does not segregate with the hearing loss phenotype and is even present in persons with normal hearing. This fact provides further support for the hypothesis that DFNA5-associated hearing loss is caused by a gain-of-function mutation.     

Septo-optic dysplasia and WS1 in the proband of a WS1 family segregating for a novel mutation in PAX3 exon 7.

A four generation family (UoM1) was ascertained with Waardenburg syndrome type 1 (WS1). The proband exhibited both WS1 and septo-optic dysplasia. A G to C transversion was identified in PAX3 exon 7 in four subjects affected with WS1 in this family including the proband. This glutamine to histidine missense mutation at position 391 may also affect splicing. There are over 50 mutations characterised in PAX3 in WS1 patients; however, this is the first example of a WS1 mutation in exon 7 of PAX3.     

A novel heterozygous nonsense mutation of the OPTN gene segregating in a Danish family with ALS

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder. About 10% of ALS cases are familial (FALS) and the genetic defect is known only in approximately 20%-30% of these cases. The most common genetic cause of ALS is SOD1 (superoxide dismutase 1) mutation. Very recently, mutations of the optineurin gene (OPTN), which is involved in open-angle glaucoma, were identified in 3 Japanese patients/families with ALS, and subsequently in a few FALS patients of European descent. We found a heterozygous nonsense mutation (c.493C>T, p.Gln165X, exon 6) in the OPTN gene in a Danish patient with ALS, and the mutation segregated from his affected father. The p.Gln165X mutation could not be detected in 1070 healthy Danish controls, in 1000 Danish individuals with metabolic phenotypes or in 64 sporadic ALS (SALS) cases. The p.Gln165X mutation described in this study is the first mutation reported in a Danish family and is likely involved in disease pathogenesis. Until now, only few OPTN mutations have been associated with ALS. As the underlying genetic defect is known only in approximately 20%-30% of FALS families, further screening of these cases is necessary for establishing the contribution of OPTN mutations in disease pathogenesis.     

A novel deletion mutation in CENPJ gene in a Pakistani family with autosomal recessive primary microcephaly

Autosomal recessive primary microcephaly (MCPH) is a rare human genetic disorder in which the head circumference is reduced because of abnormality in fetal brain growth. To date, six loci and four genes have been identified for this condition. Our study of primary MCPH led to the identification of 33 Pakistani families with different ethnic backgrounds. Most of these families showed linkage to MCPH5 locus on chromosome 1q31. Only one family with Pashtoon origin from a remote region in Pakistan linked to MCPH6 locus on chromosome 13q12.12–q12.13. Sequence analysis of exon 11 of CENPJ gene, located at MCPH6 locus, revealed a novel four base pair deletion mutation, which is predicted to be protein truncating.     

A novel mutation in the HPGD gene causing primary hypertrophic osteoarthropathy with digital clubbing in a Pakistani family

Primary hypertrophic osteoarthropathy (PHO) is a congenital multisystemic entity characterized by three major clinical symptoms: pachydermia, periostosis, and digital clubbing. Recently it has been reported that pathogenic mutations in two genes are known to be associated with PHO: HPGD and SLCO2A1. In the present study, a five‐generation consanguineous Pakistani family harboring primary hypertrophic osteoarthropathy in autosomal‐recessive pattern was ascertained. Whole genome single nucleotide polymorphisms (SNPs) genotyping and sequence analysis revealed a novel homozygous missense mutation (c.577T?C) of the human HPGD gene in all affected members of the family. The study presented here demonstrate the first case of primary hypertrophic osteoarthropathy reported in Pashtun population.     

A deafness mutation isolates a second role for the tectorial membrane in hearing

Alpha-tectorin (encoded by Tecta) is a component of the tectorial membrane, an extracellular matrix of the cochlea. In humans, the Y1870C missense mutation in TECTA causes a 50- to 80-dB hearing loss. In transgenic mice with the Y1870C mutation in Tecta, the tectorial membrane's matrix structure is disrupted, and its adhesion zone is reduced in thickness. These abnormalities do not seriously influence the tectorial membrane's known role in ensuring that cochlear feedback is optimal, because the sensitivity and frequency tuning of the mechanical responses of the cochlea are little changed. However, neural thresholds are elevated, neural tuning is broadened, and a sharp decrease in sensitivity is seen at the tip of the neural tuning curve. Thus, using Tecta(Y1870C/+) mice, we have genetically isolated a second major role for the tectorial membrane in hearing: it enables the motion of the basilar membrane to optimally drive the inner hair cells at their best frequency.