**2.3 Genes involved in Progressive Hearing Loss**

There are a few genes mutations in which have been described to cause progressive loss in hearing (*MYO3A*, *LOXHD1*, *PTPRQ* and *SERPINB6*). Additionally, some mutations in genes which cause stable and profound deafness, for example *TMC1* and *TMPRSS3* also cause progressive hearing loss. Many different mutations have been described in *TMC1*, *TMPRSS3*, *GRXRC1*, *PJVK*, *GIPC3*, *TPRN* and *PTPRQ* in affected individuals from different countries. However, it is noteworthy that apart from *DFNB8* (*TMPRSS3*), *DFNB59* (*PJVK*)) and *DFNB84* (*PTPRQ*) only one family is reported to have progression of deafness for each locus. All other mutations in these genes have been described to cause stable, moderate to profound deafness. This could be due to either of two reasons; the hearing loss associated with other mutations in these genes may also be progressive (slow or rapid progression) and has not been documented since many patients undergo audiometry at the time of enrollment in a genetic study. Alternatively, some mutations affecting a gene may cause a progressive hearing loss due to the type of mutation as for example those in *TMPRSS3*. Identical mutations may also cause stable or progressive hearing loss depending on the genetic background of the individuals.

#### *TMC1 (DFNB7)*

Homozygous inactivating mutations in *TMC1* cause severe to profound prelingual hearing impairment at the *DFNB7* locus. However, a Dutch family with autosomal recessive hearing loss was reported in which affected individuals had a postlingual onset, progressive hearing loss due to a mutation near the donor splice site of intron 19 (c.1763+3A→G) in *TMC1* (de Heer et al., 2011). The hearing loss initially affected the high frequencies and by second decade of life the hearing loss progressed to profound degree. This was documented as a "corner audiogram". Both normally spliced and aberrantly spliced *TMC1* transcripts were detected in blood of the patients. The presence of some wild type protein may account for the late onset of deafness and residual hearing in the patients, or alternately, the truncated mutant protein may have residual function (de Heer et al., 2011).

TMC1 is a transmembrane protein present in hair cells and may be involved in intracellular protein trafficking. Additionally it is also proposed to play a role in differentiation of hair cells into functional auditory receptors. The *deafness* mice mutants carry a homozygous genomic deletion in *Tmc1* and are profoundly deaf (Kurima et al., 2002). Currently there is no mouse model which mimics the recessively inherited progressive hearing loss due to mutations in *Tmc1* though a model, *Beethoven* exists for dominant deafness *DFNA36* and the mice have a missense mutation in *Tmc1* (Vreugde et al., 2002).

#### *TMPRSS3 (DFNB8)*

254 Hearing Loss

also exist which may be involved in lipoprotein transport (Borck et al., 2011b). Hearing loss due to mutations of this gene varies from moderate to profound in different individuals while it is severe in degree for one family with a mutation affecting the start codon of *ILDR1* (Borck et al., 2011b)*. Ildr1* is expressed in the developing mouse cochlea but the expression is low at birth. It increases gradually by P4 and P10. The pillar and Hensen cells have the highest expression of *Ildr1* while it can also be detected in other cells in organ of Corti

*TRIC* or *MARVELD2* encodes a tight junction protein with a ubiquitous expression in the epithelial junctions throughout the body tissues. In the inner ear, TRIC is specifically expressed in the tricellular junctions of sensory epithelia as well as those between supporting cells and the hair cells (Riazuddin et al., 2006). Affected individuals with identical mutations in *TRIC* show a wide range of variability in severity of deafness ranging from moderate to severe hearing loss to profound deafness (Chishti et al., 2008; Riazuddin et al., 2006). All mutations described in *TRIC* are predicted to produce truncated proteins and

There are a few genes mutations in which have been described to cause progressive loss in hearing (*MYO3A*, *LOXHD1*, *PTPRQ* and *SERPINB6*). Additionally, some mutations in genes which cause stable and profound deafness, for example *TMC1* and *TMPRSS3* also cause progressive hearing loss. Many different mutations have been described in *TMC1*, *TMPRSS3*, *GRXRC1*, *PJVK*, *GIPC3*, *TPRN* and *PTPRQ* in affected individuals from different countries. However, it is noteworthy that apart from *DFNB8* (*TMPRSS3*), *DFNB59* (*PJVK*)) and *DFNB84* (*PTPRQ*) only one family is reported to have progression of deafness for each locus. All other mutations in these genes have been described to cause stable, moderate to profound deafness. This could be due to either of two reasons; the hearing loss associated with other mutations in these genes may also be progressive (slow or rapid progression) and has not been documented since many patients undergo audiometry at the time of enrollment in a genetic study. Alternatively, some mutations affecting a gene may cause a progressive hearing loss due to the type of mutation as for example those in *TMPRSS3*. Identical mutations may also cause stable or progressive hearing loss depending on the

Homozygous inactivating mutations in *TMC1* cause severe to profound prelingual hearing impairment at the *DFNB7* locus. However, a Dutch family with autosomal recessive hearing loss was reported in which affected individuals had a postlingual onset, progressive hearing loss due to a mutation near the donor splice site of intron 19 (c.1763+3A→G) in *TMC1* (de Heer et al., 2011). The hearing loss initially affected the high frequencies and by second decade of life the hearing loss progressed to profound degree. This was documented as a "corner audiogram". Both normally spliced and aberrantly spliced *TMC1* transcripts were detected in blood of the patients. The presence of some wild type protein may account for the late onset of deafness and residual hearing in the patients, or alternately, the truncated

mutant protein may have residual function (de Heer et al., 2011).

including the hair cells (Borck et al., 2011b).

consequent inability to bind several scaffolding proteins.

**2.3 Genes involved in Progressive Hearing Loss** 

genetic background of the individuals.

*TMC1 (DFNB7)* 

*TRIC (DFNB49)* 

The gene encoding serine protease, TMRSS3 is expressed in supporting cells, stria vasularis as well as in the spiral ganglion in the inner ear. Mutations in *TMPRSS3* are responsible for deafness at the *DFNB8* locus. Most mutations in *TMPRSS3* result in severe to profound deafness in many world populations. However there are many mutations in *TMPRSS3*  causing less severe hearing loss which is progressive in nature. The first family with progression in hearing loss due to a *TMPRSS3* mutation was reported from Pakistan. Affected members in this family suffered from progressive deafness with onset of hearing loss in childhood and had a mutation at a splice acceptor site in intron 4 (Scott et al., 2001; Veske et al., 1996). This is predicted to create an alternative splice acceptor site which on use introduces a frameshift in the open reading frame of *TMPRSS3*. It is hypothesized that this mutation may allow limited normal splicing since the actual splice site remains unchanged. Thus some normal TMPRSS3 could be produced accounting for progressive hearing loss observed in the affected individuals.

There are other reports of mutations in *TMPRSS3* causing post-lingual progressive deafness in British, Turkish, German and Dutch families (Elbracht et al., 2007; Hutchin et al., 2005; Wattenhofer et al., 2005; Weegerink et al., 2011). One of the first studies reported a British family with two affected individuals who were homozygous for a missense mutation in *TMPRSS3* and suffered from moderate to severe hearing loss (Hutchin et al., 2005). A recent study involved 8 small nuclear families from Holland and affected individuals were compound heterozygous for different mutations of *TMPRSS3* including missense and frameshift mutations (Weegerink et al., 2011). The higher frequencies were affected first resulting in a distinctive "ski-slope" audiometric configuration followed by low frequency hearing loss resulting in a flat audiogram (Weegerink et al., 2011). It was hypothesized that some mutations in *TMPRSS3* result in creation of hypomorphic alleles accounting for the less severe loss in hearing and progression of deafness observed in affected individuals (Weegerink et al., 2011).

*Tmprss3* was shown to be important for hearing in mice as well since mutants homozygous for a nonsense mutation in the gene suffer from deafness. The hair cells start to degenerate at P12 from basal to apical turn of the cochlea and the degeneration is complete by P14 (Fasquelle et al., 2011). However, no mouse model has been reported which mimics the progressive hearing loss as observed in humans.

#### *CDH23 (DFNB12)*

Most mutations of *CDH23* cause severe phenotypes of deafness or *USH1D*. However, a few missense mutations in *CDH23* when present together in compound heterozygosity are reported to cause moderate to severe hearing loss or severe to profound deafness which is

Genetics of Nonsyndromic Recessively Inherited

nonsense and a splice site mutation.

function and ultimate degeneration of hair cells.

hearing loss but no audiograms were provided (Borck et al., 2011a).

*PJVK (DFNB59)* 

function only in the neurons.

Moderate to Severe and Progressive Deafness in Humans 257

frequencies. By the age of 50 there was moderate hearing loss at low frequencies while the high and middle frequencies were severely affected. All individuals were equally affected by the sixth decade of life. The three mutations identified in *MYO3A* included a nonsense and two splice site mutations. Hearing loss was significantly worse in affected individuals homozygous for the nonsense mutation than those who were compound heterozygous for a

MYO3A is present in the hair cells stereocilia at the tips in a characteristic pattern described as "thimble-like" (Schneider et al., 2006) with presence at the stereocilia tip and also extending further down into the shaft of the stereocilia. Mice created as models for *DFNB30* mimic the hearing loss phenotype observed in humans. *Myo3aKI/KI* mice have an engineered nonsense mutation in the gene and they exhibit a hearing loss which progresses from mild to moderate, and to moderate to severe between ages of 2.5 months to 13 months. More severe loss in hearing is observed for sounds of high frequencies. In *Myo3aKI/KI* mice, development of hair cells stereocilia is normal which is followed by a gradual loss of hair cells from basal to apical turn of the cochlea (Walsh et al., 2011). It is hypothesized that loss of MYO3A may result in failure of transport of essential components within stereocilia (Walsh et al., 2011). This affects signal transduction in the inner ear, leading to loss of

Mutations in *PJVK* cause hearing loss with or without auditory neuropathy. The hearing loss is severe to profound and stable in all individuals reported so far except in members of one Moroccan, one Iranian and three Arab families (Borck et al., 2011a; Ebermann et al., 2007b; Schwander et al., 2007). Three affected individuals in the Moroccan family had deafness with onset by age of 4 in one individual and congenital hearing loss was observed in the other two individuals. The loss in hearing is severe in degree in the former and profound in the latter two individuals. Audiometry performed over a period of three years revealed a progressive nature of hearing loss for all three affected individuals. Unlike other families with *PJVK* mutations which have missense mutations in the gene, this family had a frameshift mutation in exon 2 of the 7 exon gene and the mutation is predicted to severely truncate the protein. Hearing loss is also reported to be progressive in nature and moderate to profound in degree in a family from Iran with a frameshift mutation of *PJVK* but audiometric profiles were not provided (Schwander et al., 2007). Additionally, deafness due to a nonsense mutation in three Arab families is reported to cause stable moderate to severe

PJVK exhibits 32% identity and 54% similarity over a stretch of 250 amino acids to DFNA5 protein. It is expressed in the hair cells and spiral ganglion. Mice with a targeted knock-in missense mutation in *Pjvk*, *Dfnb59tm1Ugds/tm1Ugd* have a moderate to severe non-progressive hearing loss which is elevated at high frequencies (Delmaghani et al., 2006) unlike *sirtaki* mice which have a nonsense mutation in *Pjvk* (Schwander et al., 2007). The *sirtaki* mice have outer hair cell functional defects as apparent by absent DPOAE and also suffer from progressive deafness. No morphological defects are apparent in the ears of the *sirtaki* or knock-in mutant mice. It is hypothesized that that functional null allele of *Pjvk* inactivates protein function in both hair cells and neurons, while the missense mutation affect its

progressive in nature (Astuto et al., 2002). In two German siblings with deafness, the age of onset was also different, with onset of hearing loss at 4 and 6 years respectively. The degree of hearing loss was variable and there was asymmetric hearing loss in the older sibling.

In the inner ear, *CDH23* is expressed in the hair cells and Reissner's membrane. Mice lacking CDH23 are profoundly deaf and suffer from developmental defects of the stereocilia (Di Palma et al., 2001). In contrast, ENU induced *salsa* mutants have a recessively inherited missense mutation in *Cdh23* which affects the tip links of the stereocilia (Schwander et al., 2009). These mice have a progressive hearing loss which increases from severe in degree to profound deafness by three months of age as a consequence of gradual loss of tip links and eventual hair cell death (Schwander et al., 2009).

### *GRXCR1 (DFNB25)*

Splice site mutations were identified in *GRXCR1* in two Dutch families while three missense and a nonsense mutation were identified in *GRXRC1* in families from Pakistan and Iran which segregated with hearing loss (Odeh et al., 2010; Schraders et al., 2010a). Two mutations create alternative splice sites within *GRXCR1* which on usage are predicted to create frameshifts in the open reading frame of the gene. The presence of alternatively spliced transcripts was demonstrated for one of the splice site mutations by an *in vivo* assay on lymphoblastoid cell lines derived from patients' blood.

The single affected individual in one of the Dutch families with a splice site mutation had a significantly milder phenotype of moderate to severe hearing loss which was not progressive in nature. Additionally, the hearing loss in affected members of another Dutch family and Pakistani families varied from moderate to profound in degree. The loss in hearing was progressive from moderate to profound in the Dutch family, while it was severe in the two families from Pakistan (Schraders et al., 2010a). The audiometric profile varied from flat to a slight U-shape and was down-sloping (Schraders et al., 2010a). Data was not provided about the audiometric profile of affected individuals in families from Iran, although hearing loss was reported to be severe to profound in degree (Odeh et al., 2010).

*GRXCR1* is predicted to contain a GRX-like domain*.* These domains take part in reversible Sglutathionylation of proteins by which they are predicted to control activity or localization of the proteins*.* In the inner ear, *Grxcr1* is expressed in the hair cells and is localized along the lengths of stereocilia exhibiting a differential pattern in levels of expression in young and adult mice (Odeh et al., 2010). There are five mutant alleles of *Grxcr1* which cause profound deafness in the *pirouette* mice (Odeh et al., 2010). The absence of GRXCR1 results in formation of relatively short and thin stereocilia and cytocauds indicating actin abnormalities. This suggests that GRXCR1 plays an active role in development of actin architecture in the stereocilia (Beyer et al., 2000; Odeh et al., 2010).

#### *MYO3A (DFNB30)*

A class III myosin, *MYO3A* is required for normal hearing as shown by a single family originating from Iraq in which affected individuals had a progressive hearing loss with onset in second decade of life (Walsh et al., 2002). Interestingly, three different mutations were identified in this family which were either present in homozygosity or individuals were compound heterozygous for any of the mutations. The hearing loss first affected high frequencies. By the age of 50 there was moderate hearing loss at low frequencies while the high and middle frequencies were severely affected. All individuals were equally affected by the sixth decade of life. The three mutations identified in *MYO3A* included a nonsense and two splice site mutations. Hearing loss was significantly worse in affected individuals homozygous for the nonsense mutation than those who were compound heterozygous for a nonsense and a splice site mutation.

MYO3A is present in the hair cells stereocilia at the tips in a characteristic pattern described as "thimble-like" (Schneider et al., 2006) with presence at the stereocilia tip and also extending further down into the shaft of the stereocilia. Mice created as models for *DFNB30* mimic the hearing loss phenotype observed in humans. *Myo3aKI/KI* mice have an engineered nonsense mutation in the gene and they exhibit a hearing loss which progresses from mild to moderate, and to moderate to severe between ages of 2.5 months to 13 months. More severe loss in hearing is observed for sounds of high frequencies. In *Myo3aKI/KI* mice, development of hair cells stereocilia is normal which is followed by a gradual loss of hair cells from basal to apical turn of the cochlea (Walsh et al., 2011). It is hypothesized that loss of MYO3A may result in failure of transport of essential components within stereocilia (Walsh et al., 2011). This affects signal transduction in the inner ear, leading to loss of function and ultimate degeneration of hair cells.

#### *PJVK (DFNB59)*

256 Hearing Loss

progressive in nature (Astuto et al., 2002). In two German siblings with deafness, the age of onset was also different, with onset of hearing loss at 4 and 6 years respectively. The degree of hearing loss was variable and there was asymmetric hearing loss in the older sibling.

In the inner ear, *CDH23* is expressed in the hair cells and Reissner's membrane. Mice lacking CDH23 are profoundly deaf and suffer from developmental defects of the stereocilia (Di Palma et al., 2001). In contrast, ENU induced *salsa* mutants have a recessively inherited missense mutation in *Cdh23* which affects the tip links of the stereocilia (Schwander et al., 2009). These mice have a progressive hearing loss which increases from severe in degree to profound deafness by three months of age as a consequence of gradual loss of tip links and

Splice site mutations were identified in *GRXCR1* in two Dutch families while three missense and a nonsense mutation were identified in *GRXRC1* in families from Pakistan and Iran which segregated with hearing loss (Odeh et al., 2010; Schraders et al., 2010a). Two mutations create alternative splice sites within *GRXCR1* which on usage are predicted to create frameshifts in the open reading frame of the gene. The presence of alternatively spliced transcripts was demonstrated for one of the splice site mutations by an *in vivo* assay

The single affected individual in one of the Dutch families with a splice site mutation had a significantly milder phenotype of moderate to severe hearing loss which was not progressive in nature. Additionally, the hearing loss in affected members of another Dutch family and Pakistani families varied from moderate to profound in degree. The loss in hearing was progressive from moderate to profound in the Dutch family, while it was severe in the two families from Pakistan (Schraders et al., 2010a). The audiometric profile varied from flat to a slight U-shape and was down-sloping (Schraders et al., 2010a). Data was not provided about the audiometric profile of affected individuals in families from Iran, although hearing loss was reported to be severe to profound in degree (Odeh et al., 2010). *GRXCR1* is predicted to contain a GRX-like domain*.* These domains take part in reversible Sglutathionylation of proteins by which they are predicted to control activity or localization of the proteins*.* In the inner ear, *Grxcr1* is expressed in the hair cells and is localized along the lengths of stereocilia exhibiting a differential pattern in levels of expression in young and adult mice (Odeh et al., 2010). There are five mutant alleles of *Grxcr1* which cause profound deafness in the *pirouette* mice (Odeh et al., 2010). The absence of GRXCR1 results in formation of relatively short and thin stereocilia and cytocauds indicating actin abnormalities. This suggests that GRXCR1 plays an active role in development of actin

A class III myosin, *MYO3A* is required for normal hearing as shown by a single family originating from Iraq in which affected individuals had a progressive hearing loss with onset in second decade of life (Walsh et al., 2002). Interestingly, three different mutations were identified in this family which were either present in homozygosity or individuals were compound heterozygous for any of the mutations. The hearing loss first affected high

eventual hair cell death (Schwander et al., 2009).

on lymphoblastoid cell lines derived from patients' blood.

architecture in the stereocilia (Beyer et al., 2000; Odeh et al., 2010).

*GRXCR1 (DFNB25)* 

*MYO3A (DFNB30)* 

Mutations in *PJVK* cause hearing loss with or without auditory neuropathy. The hearing loss is severe to profound and stable in all individuals reported so far except in members of one Moroccan, one Iranian and three Arab families (Borck et al., 2011a; Ebermann et al., 2007b; Schwander et al., 2007). Three affected individuals in the Moroccan family had deafness with onset by age of 4 in one individual and congenital hearing loss was observed in the other two individuals. The loss in hearing is severe in degree in the former and profound in the latter two individuals. Audiometry performed over a period of three years revealed a progressive nature of hearing loss for all three affected individuals. Unlike other families with *PJVK* mutations which have missense mutations in the gene, this family had a frameshift mutation in exon 2 of the 7 exon gene and the mutation is predicted to severely truncate the protein. Hearing loss is also reported to be progressive in nature and moderate to profound in degree in a family from Iran with a frameshift mutation of *PJVK* but audiometric profiles were not provided (Schwander et al., 2007). Additionally, deafness due to a nonsense mutation in three Arab families is reported to cause stable moderate to severe hearing loss but no audiograms were provided (Borck et al., 2011a).

PJVK exhibits 32% identity and 54% similarity over a stretch of 250 amino acids to DFNA5 protein. It is expressed in the hair cells and spiral ganglion. Mice with a targeted knock-in missense mutation in *Pjvk*, *Dfnb59tm1Ugds/tm1Ugd* have a moderate to severe non-progressive hearing loss which is elevated at high frequencies (Delmaghani et al., 2006) unlike *sirtaki* mice which have a nonsense mutation in *Pjvk* (Schwander et al., 2007). The *sirtaki* mice have outer hair cell functional defects as apparent by absent DPOAE and also suffer from progressive deafness. No morphological defects are apparent in the ears of the *sirtaki* or knock-in mutant mice. It is hypothesized that that functional null allele of *Pjvk* inactivates protein function in both hair cells and neurons, while the missense mutation affect its function only in the neurons.

Genetics of Nonsyndromic Recessively Inherited

*TPRN (DFNB79)* 

et al., 2010).

*PTPRQ (DFNB84)* 

post natal day 90 which include hair cell loss (Grillet et al., 2009).

degree of deafness (Khan et al., 2009; Rehman et al., 2010).

available about progression of deafness in the Palestinian family*.* 

Moderate to Severe and Progressive Deafness in Humans 259

domains or nonsense mediated decay of the transcript could lead to absence of LOXHD1. In contrast, the only other mutation reported in *LOXHD1* is a founder mutation, p.R1572X, in the Ashkenazi Jews and causes prelingual profound degree of deafness (Edvardson et al., 2011). Normally LOXHD1 is present along the lengths of hair cell stereocilia plasma membrane while it cannot be detected at the tips of the stereocilia (Grillet et al., 2009). Interestingly, the hair cells bodies have no expression of *LOXHD1*. ENU induced *samba* mutants homozygous for a missense mutation in *Loxhd1* acquire hearing loss by 3 weeks of age and are completely deaf by eight weeks. The stereocilia develop normally. However, the hair cells have functional defects and DPOAE cannot be elicited from ears of *samba* mutants. Morphological defects are also observed in the inner ears of *samba* mutants with fused stereocilia and ruffled membranes at the apical cell surfaces. Additional degenerative changes are visible by

Mutations of *TPRN* cause different degrees of hearing loss in humans. In a Dutch family, hearing loss is documented as moderate to severe in degree till 11 years of age and by 15 years it progresses to profound deafness (Li et al., 2010). In contrast, hearing loss in a Moroccan family is severe in degree even in the third decade of life (Li et al., 2010). Affected members of the Dutch family had a nonsense mutation in exon 1 of *TPRN* while the affected individuals in the Moroccan family were homozygous for a frameshift mutation in exon 1. It is interesting to note that the same frameshift mutation as observed in the Moroccan family was also identified in a Pakistani family. However, individuals in this family had profound

TPRN is present in the supporting cells as well as at the base of hair cells' stereocilia in the organ of Corti at the taper region of each stereocilium just above the cuticular plate (Rehman

Congenital hearing loss which has been shown to be progressive in nature is associated with mutations in *PTPRQ*, a gene which encodes a phosphatase specific for phoshatidyl inositol, PI(4,5)P2 (Schraders et al., 2010b; Seifert et al., 2003). PI(4,5)P2 plays an important part in actin remodeling. Hearing loss due to mutations *PTPRQ* is severest in a Dutch family with a p.Y497X mutation and hearing loss was self reported to have progressed to profound by the third to fourth decade of life. However in affected individuals of a Moroccan family, a p.R457G mutation in PTPRQ caused a less severe loss in hearing. The loss in hearing was moderate which had deteriorated with age (Schraders et al., 2010b). Otoacoustic emissions were normal at the age of 13 months. The 1 to 2 KHz frequencies seemed to be more affected comparable to that observed in members of a Palestinian family with moderate to severe hearing loss also with a mutation of *PTPRQ* (Shahin et al., 2010a). In the Palestinian family, there were four affected individuals with considerable variation of hearing loss. A nonsense mutation, p.Q429X, was observed in *PTPRQ* for this family (Shahin et al., 2010a). Data is not

*Ptprq* is transiently expressed over a period of first three weeks in mouse hair cells at the basal turn of the cochlea (Goodyear et al., 2003). MYO6 plays a role in localization of PTPRQ

#### *GIPC3 (DFNB72/15/95)*

*GIPC3* encodes a PDZ domain containing protein which is important in peripheral auditory signal transmission. Mutations in this gene cause both profound deafness and progressive hearing loss. In a small Dutch hearing loss followed a different course in the two affected siblings. In the oldest affected individual, the loss in hearing was 70 dB HL at 11 months which progressed to 110 dB HL by 12 years of age. The hearing loss in second individual was 80 dB HL at 3 months which seemed to be stable as it only progressed to 90 dB HL by age of 14 years. (Charizopoulou et al., 2011). The mutation in *GIPC3* identified in the Dutch family introduces a stop codon in exon 6 (p.W301X) truncating the C-terminal. It is hypothesized that since this mutation is in the last exon it will not cause nonsense mediated decay and will allow production of a mutant protein retaining some function.

*GIPC3* mutations have also been reported in seven other large families from Pakistan which include one framsehift and and six missense mutations (Rehman et al., 2011). The younger affected individuals in these families (7 and 9 years old) had better hearing as compared to the older affected individuals (20 and 25 years old). However, information is not available as yet whether hearing loss in any of the affected individuals is stable or progressive.

A mutation affecting PDZ domain of GIPC3 has also been reported in mice which causes progressive hearing loss, *ahl5* (Charizopoulou et al., 2011). The mice have a moderate hearing loss at one month of age which progresses to profound in degree by 1 year of age. The higher frequencies are affected first with eventual involvement of all frequencies. DPOAE and the endocochlear potential are gradually affected as well. The stereocilia are defective and degenerate together with the spiral ganglion cells.

#### *BSND (DFNB73)*

*BSND* encodes barttin which is an essential subunit for two chloride channels. Heteromeric channels formed by barttin and the chloride channels play an important role in potassium recycling in the inner ear (Estevez et al., 2001)*.* A missense mutation, p.I12Tof BSND segregates with hearing loss in three families from Pakistan while in a fourth family it is present with a nonsense mutation in compound heterozygosity (Riazuddin et al., 2009). The younger individuals have a less severe hearing loss as compared to the older affected individuals suggesting that *BSND* may be involved in aetiology of progressive deafness. Functional analysis of the p.I12T mutation of *BSND* in HEK293T cells have demonstrated that function of the channels is unaffected. However, the number of channels on the surface membrane is reduced which results in a decreased current amplitude (Riazuddin et al., 2009).

#### *LOXHD1 (DFNB77)*

The 15 PLAT (polycystin/lipoxygenase/a-toxin) domains encoding gene *LOXHD1* was found to be mutated in an Iranian family with postlingual onset of hearing loss at ages ranging from 7 to 8 years. Affected individuals in the family had preserved hearing at low frequencies in the beginning. The loss was mild to moderate at frequencies of 0.5 to 2 KHz. Hearing loss worsened during childhood and adolescence to moderate to severe at mid and high frequencies. All frequencies were affected eventually (Grillet et al., 2009). A nonsense mutation, p.R670X was identified which introduces a premature stop codon at the Cterminal end of f the fifth PLAT domain. This could either lead to a protein lacking 10 PLAT domains or nonsense mediated decay of the transcript could lead to absence of LOXHD1. In contrast, the only other mutation reported in *LOXHD1* is a founder mutation, p.R1572X, in the Ashkenazi Jews and causes prelingual profound degree of deafness (Edvardson et al., 2011).

Normally LOXHD1 is present along the lengths of hair cell stereocilia plasma membrane while it cannot be detected at the tips of the stereocilia (Grillet et al., 2009). Interestingly, the hair cells bodies have no expression of *LOXHD1*. ENU induced *samba* mutants homozygous for a missense mutation in *Loxhd1* acquire hearing loss by 3 weeks of age and are completely deaf by eight weeks. The stereocilia develop normally. However, the hair cells have functional defects and DPOAE cannot be elicited from ears of *samba* mutants. Morphological defects are also observed in the inner ears of *samba* mutants with fused stereocilia and ruffled membranes at the apical cell surfaces. Additional degenerative changes are visible by post natal day 90 which include hair cell loss (Grillet et al., 2009).
