Autosomal Recessive Non-syndromic Hearing Impairment Research Articles

Identification of a novel compound heterozygous mutation in PTPRQ in a DFNB84 family with prelingual sensorineural hearing impairment.

With a prevalence of 0.1 %, hearing loss is among the most common sensory impairments and affects several million people around the world. Identification of deafness-related genes or loci may facilitate basic research and clinical translational research of the disorder. The PTPRQ gene encodes protein tyrosine phosphatase receptor Q, which is required for the formation of shaft connectors and the normal maturation and development of hair bundles in the mammalian cochlea. Here, we present the genetic and molecular characteristics of a Kazakh family with an autosomal recessive non-syndromic hearing impairment, DFNB84. Using whole-exome sequencing, we identified two mutations that together form a novel compound heterozygous mutation in PTPRQ. Sanger sequencing confirmed that the affected members inherited both the c.16_17insT (L8fsX18) and c.2714delA (E909fsX922) mutations. Both mutations lead to a frameshift and a truncated form of the protein. The novel compound heterozygous mutation co-segregated with hearing loss in this family, and neither of the two mutations was found in 200 healthy Kazakh controls or in any of the public databases. In the study, we identified novel mutations in PTPRQ responsible for DFNB84. This is the third report of PTPRQ mutations involved in deafness and the first report of familial deafness in China. The identification of novel mutations in PTPRQ presented here further confirms the essential role of PTPRQ in hearing development and auditory function. Our data provide additional molecular information for establishing a better genotype-phenotype understanding of DFNB84.

A canonical splice site mutation in GIPC3 causes sensorineural hearing loss in a large Pakistani family.

With homozygosity mapping we have identified two large homozygous regions on chromosome 3q13.11-q13.31 and chromosome 19p13.3-q31.32 in a large Pakistani family suffering from autosomal recessive nonsyndromic hearing impairment (arNSHI). The region on chromosome 19 overlaps with the previously described deafness loci DFNB15, DFNB72 and DFNB95. Mutations in GIPC3 have been shown to underlie the nonsyndromic hearing impairment linked to these loci. Sequence analysis of all exons and exon-intron boundaries of GIPC3 revealed a homozygous canonical splice site mutation, c.226-1G>T, in GIPC3. This is the first mutation described in GIPC3 that affects splicing. The c.226-1G>T mutation is located in the acceptor splice site of intron 1 and is predicted to affect the normal splicing of exon 2. With a minigene assay it was shown to result in the use of an alternative acceptor site in exon 2, resulting in a frameshift and a premature stop codon. This study expands the mutational spectrum of GIPC3 in arNSHI.

The promoter mutation c.-259C>T (-3438C>T) is not a common cause of non-syndromic hearing impairment in Austria.

The objective of this study was to investigate the relevance of routine assessment of c.-259C>T in the Austrian newborn screening program. Homozygous and compound heterozygous mutations in the coding region of the human gene encoding gap junction protein GJB2 (Connexin 26) cause up to 50 % of neonatal autosomal recessive non-syndromic hearing impairment identified in Caucasian newborn screening programs. More recently, a null mutation in the GC box of the GJB2 basal promoter c.-259C>T has been described which causes hearing impairment by completely suppressing GJB2 promoter activity. We determined the occurrence of c.-259C>T in cases of non-syndromic hearing impairment lacking known pathogenic alterations in GJB2 (n = 43), a non-syndromic hearing impaired patient group (n = 15) bearing the heterozygous GJB2 mutations c.35delG, c.[79G>A];[341A>G] (p. [V27I];[E114G]), c.109G>A (p.V37I), c.154G>C (p.V52L), c.262G>T (p.A88S), c.269T>C (p.L90P) and c.551G>C (p.R184P) and in a normal hearing group lacking alterations in GJB2 (n = 50). In the analyzed groups, no occurrence of c.-259C>T was found. The c.-259C>T mutation, previously described as -3438C>T, is not a common cause of non-syndromic hearing impairment alone or together with heterozygous pathogenic GJB2 mutations that are statistically overrepresented in non-syndromic hearing impaired patient groups. Screening of newborns for c.-259C>T is therefore unlikely to be commonly found in Austrian NSHI patients but could make a significant contribution to non-syndromic hearing impairment in other populations.

Progressive hearing loss and vestibular dysfunction caused by a homozygous nonsense mutation in CLIC5.

In a consanguineous Turkish family diagnosed with autosomal recessive nonsyndromic hearing impairment (arNSHI), a homozygous region of 47.4 Mb was shared by the two affected siblings on chromosome 6p21.1-q15. This region contains 247 genes including the known deafness gene MYO6. No pathogenic variants were found in MYO6, neither with sequence analysis of the coding region and splice sites nor with mRNA analysis. Subsequent candidate gene evaluation revealed CLIC5 as an excellent candidate gene. The orthologous mouse gene is mutated in the jitterbug mutant that exhibits progressive hearing impairment and vestibular dysfunction. Mutation analysis of CLIC5 revealed a homozygous nonsense mutation c.96T>A (p.(Cys32Ter)) that segregated with the hearing loss. Further analysis of CLIC5 in 213 arNSHI patients from mostly Dutch and Spanish origin did not reveal any additional pathogenic variants. CLIC5 mutations are thus not a common cause of arNSHI in these populations. The hearing loss in the present family had an onset in early childhood and progressed from mild to severe or even profound before the second decade. Impaired hearing is accompanied by vestibular areflexia and in one of the patients with mild renal dysfunction. Although we demonstrate that CLIC5 is expressed in many other human tissues, no additional symptoms were observed in these patients. In conclusion, our results show that CLIC5 is a novel arNSHI gene involved in progressive hearing impairment, vestibular and possibly mild renal dysfunction in a family of Turkish origin.

Adenylate cyclase 1 (ADCY1) mutations cause recessive hearing impairment in humans and defects in hair cell function and hearing in zebrafish

Cyclic AMP (cAMP) production, which is important for mechanotransduction within the inner ear, is catalyzed by adenylate cyclases (AC). However, knowledge of the role of ACs in hearing is limited. Previously, a novel autosomal recessive non-syndromic hearing impairment locus DFNB44 was mapped to chromosome 7p14.1-q11.22 in a consanguineous family from Pakistan. Through whole-exome sequencing of DNA samples from hearing-impaired family members, a nonsense mutation c.3112C>T (p.Arg1038*) within adenylate cyclase 1 (ADCY1) was identified. This stop-gained mutation segregated with hearing impairment within the family and was not identified in ethnically matched controls or within variant databases. This mutation is predicted to cause the loss of 82 amino acids from the carboxyl tail, including highly conserved residues within the catalytic domain, plus a calmodulin-stimulation defect, both of which are expected to decrease enzymatic efficiency. Individuals who are homozygous for this mutation had symmetric, mild-to-moderate mixed hearing impairment. Zebrafish adcy1b morphants had no FM1-43 dye uptake and lacked startle response, indicating hair cell dysfunction and gross hearing impairment. In the mouse, Adcy1 expression was observed throughout inner ear development and maturation. ADCY1 was localized to the cytoplasm of supporting cells and hair cells of the cochlea and vestibule and also to cochlear hair cell nuclei and stereocilia. Ex vivo studies in COS-7 cells suggest that the carboxyl tail of ADCY1 is essential for localization to actin-based microvilli. These results demonstrate that ADCY1 has an evolutionarily conserved role in hearing and that cAMP signaling is important to hair cell function within the inner ear.

Homozygosity for the V37I GJB2 mutation in fifteen probands with mild to moderate sensorineural hearing impairment: Further confirmation of pathogenicity and haplotype analysis in Asian populations

Hearing impairment affects 1 in 650 newborns, making it the most common congenital sensory impairment. Autosomal recessive nonsyndromic sensorineural hearing impairment (ARNSHI) comprises 80% of familial hearing impairment cases. Mutations in GJB2 account for a significant number of ARNSHI (and up to 50% of documented recessive (e.g., more than 1 affected sibling) hearing impairment in some populations). Mutations in the GJB2 gene are amongst the most common causes of hearing impairment in populations of various ethnic backgrounds. Two mutations of this gene, 35delG and 167delT, account for the majority of reported mutations in Caucasian populations, especially those of Mediterranean and Ashkenazi Jewish background. The 235delC mutation is most prevalent in East Asian populations. Some mutations are of less well-characterized significance. The V37I missense mutation, common in Asian populations, was initially described as a polymorphism and later as a potentially pathogenic mutation. We report here on 15 unrelated individuals with ARNSHI and homozygosity for the V37I GJB2 missense mutation. Nine individuals are of Chinese ancestry, two are of unspecified Asian descent, one is of Japanese descent, one individual is of Vietnamese ancestry, one of Philippine background and one of Italian and Cuban/Caucasian background. Homozygosity for the V37I GJB2 mutation may be a more common pathogenic missense mutation in Asian populations, resulting in mild to moderate sensorineural hearing impairment. We report a presumed haplotype block specific to East Asian individuals with the V37I mutation encompassing the GJB2 gene that may account for the high prevalence in East Asian populations.

Mutations in KARS, Encoding Lysyl-tRNA Synthetase, Cause Autosomal-Recessive Nonsyndromic Hearing Impairment DFNB89

Previously, DFNB89, a locus associated with autosomal-recessive nonsyndromic hearing impairment (ARNSHI), was mapped to chromosomal region 16q21-q23.2 in three unrelated, consanguineous Pakistani families. Through whole-exome sequencing of a hearing-impaired individual from each family, missense mutations were identified at highly conserved residues of lysyl-tRNA synthetase (KARS): the c.1129G>A (p.Asp377Asn) variant was found in one family, and the c.517T>C (p.Tyr173His) variant was found in the other two families. Both variants were predicted to be damaging by multiple bioinformatics tools. The two variants both segregated with the nonsyndromic-hearing-impairment phenotype within the three families, and neither mutation was identified in ethnically matched controls or within variant databases. Individuals homozygous for KARS mutations had symmetric, severe hearing impairment across all frequencies but did not show evidence of auditory or limb neuropathy. It has been demonstrated that KARS is expressed in hair cells of zebrafish, chickens, and mice. Moreover, KARS has strong localization to the spiral ligament region of the cochlea, as well as to Deiters' cells, the sulcus epithelium, the basilar membrane, and the surface of the spiral limbus. It is hypothesized that KARS variants affect aminoacylation in inner-ear cells by interfering with binding activity to tRNA or p38 and with tetramer formation. The identification of rare KARS variants in ARNSHI-affected families defines a gene that is associated with ARNSHI.

Identification of a SNP in a Regulatory Region of GJB2 Associated With Idiopathic Nonsyndromic Autosomal Recessive Hearing Loss in a Multicenter Study

Additional genetic changes in the regulatory region of the human GJB2 gene encoding the gap junction protein (Connexin 26) may contribute to sensorineural hearing loss. Mutations in GJB2 cause up to 50% of autosomal recessive nonsyndromic hearing impairment (NSHI). In the present study, we screened the putative 5' GJB2 regulatory region for novel alterations. In idiopathic familial cases of NSHI lacking known pathogenic alterations in GJB2, we identified a T→C transition (refSNP: rs117685390) in a putative transcription factor binding sequence 228 bp proximal to the transcriptional start site at a homozygous frequency of 0.125 (n = 40), significantly overrepresented in comparison to the homozygous allele frequencies of 0.043 in the normal-hearing Caucasian population (n = 211; p < 0.001). In a NSHI family, inheritance of the rs117685390 C allele segregated on independent chromosomes with NSHI in conjunction with heterozygous inheritance of c.35delG, the most common Caucasian mutation in the GJB2 coding region. In a patient group (n = 32) bearing heterozygous pathogenic c.35delG mutations, - rs117685390 C allele homozygosity was also highly overrepresented (0.25; p < 0.001) and not exclusively linked to the c.35delG mutation in cis in patients homozygous for c.35delG. However, in the majority of NSHI homozygous c.35delG chromosomes examined (91/94), c.35delG homozygosity was linked to the rs117685390 C allele in cis. These results suggest that the rs117685390 C allele could represent a biomarker for the development of NSHI in Caucasian populations and may be included in risk assessment for the development of NSHI.

Genome-wide analysis reveals a novel autosomal-recessive hearing loss locus DFNB80 on chromosome 2p16.1-p21

Hearing impairment (HI) is the decreased ability to hear and discriminate among sounds. It is one of the most common birth defects. Epidemiological data show that more than one child in 1000 is born with HI, whereas more than 50% of prelingual HI cases are found to be hereditary. So far, 95 published autosomal-recessive nonsyndromic HI (ARNSHI) loci have been mapped, and 41 ARNSHI genes have been identified. In this study, we performed a genome-wide linkage study in a consanguineous Tunisian family, and report the mapping of a novel ARNSHI locus DFNB80 to chromosome 2p16.1-p21 between the two single-nucleotide polymorphisms rs10191091 and rs2193485 with a maximum multipoint logarithm of odds score of 4.1. The screening of seven candidate genes, failed to reveal any disease-causing mutations.

Mutations of the Gene Encoding Otogelin Are a Cause of Autosomal-Recessive Nonsyndromic Moderate Hearing Impairment

Already 40 genes have been identified for autosomal-recessive nonsyndromic hearing impairment (arNSHI); however, many more genes are still to be identified. In a Dutch family segregating arNSHI, homozygosity mapping revealed a 2.4 Mb homozygous region on chromosome 11 in p15.1-15.2, which partially overlapped with the previously described DFNB18 locus. However, no putative pathogenic variants were found in USH1C, the gene mutated in DFNB18 hearing impairment. The homozygous region contained 12 additional annotated genes including OTOG, the gene encoding otogelin, a component of the tectorial membrane. It is thought that otogelin contributes to the stability and strength of this membrane through interaction or stabilization of its constituent fibers. The murine orthologous gene was already known to cause hearing loss when defective. Analysis of OTOG in the Dutch family revealed a homozygous 1bp deletion, c.5508delC, which leads to a shift in the reading frame and a premature stop codon, p.Ala1838ProfsX31. Further screeningof 60 unrelated probands from Spanish arNSHI families detected compound heterozygous OTOG mutations in one family, c.6347C>T (p.Pro2116Leu) and c. 6559C>T (p.Arg2187X). The missense mutation p.Pro2116Leu affects a highly conserved residue in the fourth von Willebrand factor type D domain of otogelin. The subjects with OTOG mutations have a moderate hearing impairment, which can be associated with vestibular dysfunction. The flat to shallow "U" or slightly downsloping shaped audiograms closely resembled audiograms of individuals with recessive mutations in the gene encoding α-tectorin, another component of the tectorial membrane. This distinctive phenotype may represent a clue to orientate the molecular diagnosis.

A Mutation in PNPT1, Encoding Mitochondrial-RNA-Import Protein PNPase, Causes Hereditary Hearing Loss

A subset of nuclear-encoded RNAs has to be imported into mitochondria for the proper replication and transcription of the mitochondrial genome and, hence, for proper mitochondrial function. Polynucleotide phosphorylase (PNPase or PNPT1) is one of the very few components known to be involved in this poorly characterized process in mammals. At the organismal level, however, the effect of PNPase dysfunction and impaired mitochondrial RNA import are unknown. By positional cloning, we identified a homozygous PNPT1 missense mutation (c.1424A>G predicting the protein substitution p.Glu475Gly) of a highly conserved PNPase residue within the second RNase-PH domain in a family affected by autosomal-recessive nonsyndromic hearing impairment. Invitro analyses in bacteria, yeast, and mammalian cells showed that the identified mutation results in a hypofunctional protein leading to disturbed PNPase trimerization and impaired mitochondrial RNA import. Immunohistochemistry revealed strong PNPase staining in the murine cochlea, including the sensory hair cells and the auditory ganglion neurons. In summary, we show that a component of the mitochondrial RNA-import machinery is specifically required for auditory function.

The c.242G&gt;A mutation in LRTOMT gene is responsible for a high prevalence of deafness in the Moroccan population

Congenital hearing impairment (HI) affects one in 1,000 newborns and has a genetic cause in 50 % of the cases. Autosomal recessive non-syndromic hearing impairment is responsible for 70-80 % of all hereditary cases of HI. Recently, it has been demonstrated that, mutations of LRTOMT are associated with profound nonsyndromic hearing impairment at the DFNB63 locus. The objective of this study is to evaluate the carrier frequency of c.242G>A mutation in LRTOMT gene and define the contribution of this gene in the etiology of deafness in Moroccan population. We screened 105 unrelated Moroccan families with non-syndromic HI and 120 control individuals for mutation in the exon 8 of the LRTOMT gene, by sequencing and PCR-RFLP. The Homozygous c.242G>A mutation was found in 8.75 % of the families tested and in 4.16 % of control in the heterozygous state. Our results show that after the GJB2 gene mutation in LRTOMT gene is the second cause of congenital hearing impairment in Moroccan patients. This finding should facilitate diagnosis of congenital deafness of the affected subjects in Morocco.

Homozygous M34T mutation of the GJB2 gene associates with an autosomal recessive nonsyndromic sensorineural hearing impairment in Finnish families

Conclusion: The genetic and audiological data support the hypothesis that the p.M34T is a pathogenic mutation in the Finnish population. The p.M34T mutation displays an autosomal recessive pattern of inheritance and is associated with mild to moderate nonsyndromic sensorineural hearing impairment (SNHI) in the homozygous state. The audiograms often display a hearing impairment notch at 2–4 kHz in young patients, which may aid in the early diagnosis. Objectives: The aim of the study was to assess whether the p.M34T mutation in the GJB2 gene may associate with nonsyndromic SNHI. Methods: We systematically reviewed the families with children diagnosed with nonsyndromic SNHI caused by a homozygous p.M34T mutation at the Kuopio and Oulu University Hospital Clinics. The children were re-examined and audiological and genetic data were obtained from their parents and healthy siblings to study genotype–phenotype correlation. Results: We describe 11 patients from 6 families including 5 sibling pairs from 6 to 23 years of age with homozygous p.M34T genotype all having mild nonsyndromic SNHI. In addition, we found three patients with compound p.M34T mutation also exhibiting mild to moderate SNHI.

A Review on Autosomal Recessive Non-syndromic Hearing Impairment

Hearing impairment (HI) or deafness is the most common sensory disorder in humans. The prevalence of bi- lateral hearing loss is 1.6 in 1,000 in Pakistan. HHI may be syndromic or non-syndromic, prelingual or postlingual, and conductive or sensorineural. Non-syndromic HHI results in sensorineural hearing loss. About 40 genes are identified to cause non-syndromic autosomal recessive deafness of 120 cloned genes of deafness. HHI is 75-80% transmitted through an autosomal recessive trait. Total 95 autosomal recessive non-syndromic deafness loci (DFNB) have been identified. Up till now, 28 loci and 15 genes involved in deafness have been mapped in Pakistan. Different mutations in the same gene may cause different phenotypes. Most cases of recessive HHI involve mutation in a single locus. But cases are found in which mutation in two different genes (GJB2/GJB6) or two different mutations in a single allele (GJB2) are involved. The GJB2 is the most important gene responsible for 50% of the cases of prelingual, non-syndromic autosomal recessive hearing loss. Others include SLC26A4, MYO15A, OTOF, CDH23 and TMC1. These genes are the most frequent cause of autosomal re- cessive non-syndromic hearing impairment. In the present report, we have reviewed these genes, their functions, mutations, and their contribution to genetic deafness. Cochlear implant is one of the solutions for cure of congenital sensorineural deafness. Gene therapy is under clinical trials to treat the defective gene involved in hearing impairment.

Molecular analysis of the TMPRSS3 gene in Moroccan families with non-syndromic hearing loss

Autosomal recessive non-syndromic hearing impairment (ARNSHI) is the most common type of inherited hearing impairment, accounting for approximately 80% of inherited prelingual hearing impairment. Hearing loss is noted to be both phenotypically and genetically heterogeneous. Mutations in the TMPRSS3 gene, which encodes a transmembrane serine protease, are known to cause autosomal recessive non-syndromic hearing impairment DFNB8/10.In order to elucidate if the TMPRSS3 gene is responsible for ARNSHI in 80 Moroccan families with non-syndromic hearing impairment, the gene was sequenced using DNA samples from these families. Nineteen TMPRSS3 variants were found, nine are located in the exons among which six are missense and three are synonymous. The 10 remaining variations are located in non-coding regions. Missense variants analysis show that they do not have a significant pathogenic effect on protein while pathogenicity of some variant remains under discussion. Thus we show that the TMPRSS3 gene is not a major contributor to non-syndromic deafness in the Moroccan population.

Autosomal Recessive Nonsyndromic Hearing Impairment due to a Novel Deletion in the RDX Gene

The RDX gene anchors cytoskeletal actin of stereocilia to hair cell transmembrane and is responsible for autosomal recessive nonsyndromic hearing impairment (ARNSHI) due to DFNB24. A genome scan was performed using DNA samples from a consanguineous Pakistani family with ARNSHI. A significant maximum two-point LOD score of 4.5 (θ = 0) and multipoint LOD score of 5.8 were achieved at marker D11S1998 (chr11 : 117.20 Mb). The region of homozygosity is bounded by markers D11S2000 (105.06 Mb) and D11S4464 (123.13 Mb) and contains the NSHI genes TECTA and RDX. Although no potentially causal variants were identified in the TECTA gene, within the RDX gene a novel deletion c.1076_1079delTTAA (p.Ile359Lysfs∗6) was identified. The RDX deletion segregates with ARNSHI within the family and was not observed in 500 control chromosomes. It is predicted to cause premature truncation of radixin at the α-helical domain and to result in nonfunctional transcripts within the cochlea. RDX isoforms which encode the coiled-coil region of the α-helical domain are deemed necessary for proper function of hair cell stereocilia.

A Novel ESRRB Deletion Is a Rare Cause of Autosomal Recessive Nonsyndromic Hearing Impairment among Pakistani Families

Mutations in the estrogen-related receptor beta (ESRRB) gene is the underlying cause of autosomal recessive nonsyndromic hearing impairment (ARNSHI) due to the DFNB35 locus which maps to 14q24.3. A genome scan of a large consanguineous Pakistani pedigree with ARNSHI established linkage with a maximum multipoint LOD score of 4.2 to the 14q24 region and the region of homozygosity contained the ESRRB gene. Sequencing of the ESRRB gene using DNA samples from hearing-impaired family members uncovered a novel three-nucleotide deletion c.1018_1020delGAG (p.Glu340del). The deletion segregates with hearing impairment in the pedigree and was not observed in 500 control chromosomes. The deletion of glutamic acid residue occurs in the ligand-binding domain of ESRRB protein. It is expected that the deletion affects the ligand-binding activity of the domain in ESRRB, which leads to the ARNSHI.

A new autosomal recessive nonsyndromic hearing impairment locus DFNB96 on chromosome 1p36.31–p36.13

A novel locus for autosomal recessive nonsyndromic hearing impairment (ARNSHI), DFNB96, was mapped to 1p36.31-p36.13. A whole genome linkage scan was performed using DNA samples from a consanguineous family from Pakistan with ARNSHI. A maximum two-point LOD score of 3.2 was obtained at marker rs8627 (chr1:8.34Mb) at θ=0 and a significant maximum multipoint LOD score of 3.8 was achieved at 15 contiguous markers from rs630075 (9.3 Mb) through rs10927583 (15.13 Mb). The 3-unit support interval and the region of homozygosity were both delimited by markers rs3817914 (6.42 Mb) and rs477558 (18.09 Mb) and contain 11.67 Mb. Of the 125 genes within the DFNB96 interval, the previously identified ARNSHI gene for DFNB36, ESPN and two genes that cause Bartter syndrome, CLCNKA and CLCNKB, were sequenced, but no potentially causal variants were identified.

High frequency of autosomal‐recessive DFNB59 hearing loss in an isolated Arab population in Israel

Autosomal-recessive non-syndromic hearing impairment (DFNB) is usually of prelingual onset with a moderate to profound degree of hearing loss. More than 70 DFNB loci have been mapped and ~40 causative genes have been identified. Non-syndromic hearing impairment caused by mutations of DFNB59 (encoding pejvakin) has been described in a couple of families in which affected individuals presented with either auditory neuropathy or hearing loss of cochlear origin. We have identified and clinically evaluated three consanguineous families of Israeli Arab origin with prelingual non-syndromic hearing impairment and absent otoacoustic emissions in a total of eight affected individuals. All the families originate from the same village and bear the same family name. We have identified a c.406C>T (p.R136X) nonsense mutation in the DFNB59 gene in affected individuals from these families. Among the inhabitants of the village, we found an exceptionally high carrier frequency of ~1 in 12 individuals (7/85; 8.2%). The high prevalence of hearing impairment can be explained by a founder effect and the high consanguinity rate among the inhabitants of this village.

Novel TMPRSS3 variants in Pakistani families with autosomal recessive non‐syndromic hearing impairment

Mutations in the TMPRSS3 gene are known to cause autosomal recessive non-syndromic hearing impairment (ARNSHI). After undergoing a genome scan, 10 consanguineous Pakistani families with ARNSHI were found to have significant or suggestive evidence of linkage to the TMPRSS3 region. In order to elucidate if the TMPRSS3 gene is responsible for ARNSHI in these families, the gene was sequenced using DNA samples from these families. Six TMPRSS3 variants were found to cosegregate in 10 families. None of these variants were detected in 500 control chromosomes. Four novel variants, three of which are missense [c.310G>A (p.Glu104Lys), c.767C>T (p.Ala256Val) and c.1273T>C (p.Cys425Arg)] and one nonsense [c.310G>T (p.Glu104Stop)], were identified. The pathogenicity of novel missense variants was investigated through bioinformatics analyses. Additionally, the previously reported deletion c.208delC (p.His70ThrfsX19) was identified in one family and the known mutation c.1219T>C (p.Cys407Arg) was found in five families, which makes c.1219T>C (p.Cys407Arg) as the most common TMPRSS3 mutation within the Pakistani population. Identification of these novel variants lends support to the importance of elements within the low-density lipoprotein receptor A (LDLRA) and serine protease domains in structural stability, ligand binding and proteolytic activity for proper TMPRSS3 function within the inner ear.