Учебники / Genetic Hearing Loss Willems 2004

the GATA3 gene. Subsequent mutation analysis in HDR patients confirmed that haploinsu ciency for GATA3 is the underlying mechanism of the human HDR syndrome (5). Thus far, di erent mutations have been found including missense and nonsense mutations, intragenic deletions, insertions, and whole-gene deletions (5,15) (Table 1). These mutations result in a truncated protein that fails to bind the DNA, suggesting a loss of function.

II.CLINICAL CHARACTERISTICS

Clinically, the hypoparathyroidism is characterized by low serum calcium as a result of a deficient parathyroid hormone (PTH) secretion. PTH levels measured in serum of HDR patients range from low normal to undetectable. This leads to symptomatic or subclinical manifestations such as hypocalcemic seizures, calcium deposits, and bone demineralization. The renal anomalies observed in HDR patients belong to a spectrum of malformations including renal hypoand dysplasia, vesicoureteral reflux, and agenesis of the kidney (16). The deafness found in HDR patients is of the sensorineural type, bilateral, and present at birth. The auditory loss is more pronounced at the higher frequencies and the severity ranges from moderate to severe, necessitating the use of hearing aids. Further deterioration of the hearing loss with advancing age is rarely the case. Phenotypic variability has been observed for the renal anomalies and the deafness. This might be due to variable penetrance of these anomalies; however, lack of appropriate investigations and possible phenotypic changes with age may account for the observed variability. Thus far, no mutations have been found in four families presenting with HDR syndrome, of which two also had additional features (5,15) (Table 1). It remains to be clarified whether a mutation in promotor or intron sequences remains undetected or whether the HDR syndrome is a heterogeneous disorder.

III.GATA3 EXPRESSION

The HDR phenotype is consistent with the observed expression of GATA3 in the developing kidney, inner ear, and parathyroids in human as well as in mouse embryos (17,18). GATA3 expression can already be observed in the otic placode in the mouse at E8. From then on, GATA3 is expressed in the main components of the developing auditory system, including the sensory epithelium, the a erent and e erent nerves, and the mesenchymal and ectodermal cells of the developing middle and outer ear (17,19). Based on

its expression pattern, it has been suggested that GATA3 is involved in the controlled suppression of hair cell di erentiation in the mouse, to form the adult sensory cell pattern in the cochlea (20). Insu cient levels of GATA3 and its regulated proteins could interfere with these strictly controlled processes and lead to a decreased auditory function. Besides expression in the cochlea, studies have shown that GATA3 expression is downstream of Hoxb1 expression during mouse hindbrain development (21). Lack of GATA3 expression in rhombomere 4 inhibits the projection of contralateral vestibuloacoustic e erent neurons. This indicates that GATA3 may play a role in the di erentiation of the inner ear e erent neurons. However, clarification of these roles requires further studies, including identification of downstream target genes in these organs.

A homozygous Gata3 knockout mouse has been reported by Pandolfi et al. (8) and displays multiple organ abnormalities, especially of the central nervous system together with a total block of T-cell di erentiation and massive internal bleeding, lethal at midgestation. In contrast to human GATA3 haploinsu ciency, heterozygous Gata3 knockout mice were reported to be normal. However, careful reexamination of these animals showed the presence of sensorineural deafness in the heterozygous mutants (F. Grosveld, personal communication). This finding was already suspected from detailed GATA3 expression studies during ear morphogenesis in normal and mutant mice (22).

REFERENCES

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formations, and cervical sinuses/cysts/fistula occur together, reflecting the embryological origin of these structures from the first and second branchial arches.

The prevalence of BOR syndrome (MIM#113650; URL http://www. ncbi.nlm.nih.gov/Omim) is 1:40,000 and has been reported to be as high as 2% of the deaf population (27). However, the prevalence of BOR syndrome may be much higher than that reported earlier. In the general population, ear pits and ear tags are the most common ear malformations, which occur with a frequency of 5–6/1000 live births (28,29). It is not clear what proportion of those cases (ear anomalies, tags/pits) are later diagnosed as BOR. In the past, the families with single or only two major phenotypes were considered of di erent genetic origin than that of BOR (presence of three or more major clinical features). However, EYA1 mutations (see below for details) have been detected in many familial and isolated cases with symptoms of preauricular sinuses and/or hearing as well as families with typical BOR loss (30–37). In the light of new findings reported in the last few years, such as identification of the EYA1 gene and localization of the second BOR gene, more detailed studies are needed to determine the frequency of BOR syndrome.

II.CLINICAL ASPECTS OF BOR

Several studies have been conducted in the past to determine the frequency of di erent clinical phenotypes associated with BOR. Among all the anomalies reported, the hearing impairment was observed to be the most common feature (90%). Other symptoms associated with BOR were preauricular pits (77%), branchial cysts or fistulas usually found on the external lower third of the neck (63%), and renal anomalies (13%). In another study, renal anomalies were identified in 67% of individuals examined by ultrasound or excretory uregraphy (26). More recently, hypoplasia and dysplasia of the cochlea were described in several BOR patients by radiological techniques using temporal bones. An enlarged vestibular aqueduct, cochlear hypoplasia, and a widened vestibular sac were also reported (38–40). Other malformations included external (atretic, stenotic, or tortuous external canal) and middle ear anomalies (malformed enlarged or small) (26,38). Many other features, although not very common, such as abnormalities of the face, palate, ureters, and bladder, dysfunction of the lacrimal system, otitis media, palatoschisis, meatal atresia, malformation of facial nerve or paralysis, intracranial aneurysms, congenital cholesteatoma, and shoulder abnormalities, have also been associated with BOR (11,12,18,26,41–48). The

overlapping clinical features associated with branchial anomalies are presented in Table 1.

III.MOLECULAR GENETICS

BOR is inherited as an autosomal dominant disorder. Similar to other dominant developmental disorders, the phenotype is quite variable and not all the features of the syndrome are expressed in all carriers of BOR mutations. In the late seventies, many studies suggested that BO (branchio-oto), BOR, and BR (branchio-renal) are the result of mutations in a single gene. In 1992, the gene for BOR syndrome was localized to chromosome 8q (57,58). The EYA1 gene was identified as the cause of BOR syndrome (59) and subsequently more than 30 mutations have been reported (30–37). Among many EYA1 mutations reported in BOR cases, there are some mutations also identified in patients who had only eye defects (60), suggesting that the human Eya1 gene is also involved in eye morphogenesis. EYA1 is the human homolog of the Drosophila eye-absent (Eya) gene (61). More recent results indicate that families with branchial and hearing anomalies with no renal manifestation are not allelic to the BOR gene (62–64), suggesting the involvement of other gene(s). Recently, the second gene associated with branchial anomalies was localized to chromosome 1q (65).

The human Eya1 cDNA is approximately 3.6 kb in length. The predicted protein of 539 amino acids is encoded by a 16-exon gene that spans approximately 156 kb of the genome and generates a 5.5-kb transcript (30). The human introns ranged in size from 0.1 to 27.5 kb in length. The transcript for EYA1 and the encoded gene product have wide distribution in human tissues, including kidney, brain, lung, heart, and skeletal muscle (30).

Murine homologs of the Drosophila eya gene have also been identified (66). The new family of vertebrate genes designated EYA1, EYA2, EYA3, and EYA4 share a high degree of sequence similarity with the Drosophila Eya gene. In mouse, Eya protein sequences reveal two distinct domains, a highly conserved 271-amino-acid carboxyl (C-) terminal region and a nonconserved amino (N-) terminal region. Eya1 is widely expressed in cranial placodes, in the branchial arches and the CNS, and in complementary or overlapping patterns during organogenesis (67). The EYA1 gene is a transcriptional coactivator and has been shown to interact with Six-family proteins (68) and possibly SIX, DACH, and/or G proteins are involved in the pathogenesis processes (69). The murine EYA1 is coexpressed with Six1 in early branchial and otic development and with Six2 during early kidney development (70). These results suggest that despite enormous morphological

di erences, the conserved Eya-Six regulatory pathway is used during mammalian organogenesis.

A recent study has been conducted to inactivate the EYA1 gene in mice to understand the developmental pathogenesis of organs a ected in BOR syndromes (67,70,71). The knockout mice as well as a mouse strain with spontaneous mutation in the EYA1 gene have cochlear and kidney anomalies (67,70,72). The phenotypic defects in mouse are strikingly similar to those of BOR syndrome. Using genetically well-defined mice with varying genetic background, linkage backcross mice bearing EYA1 mutations will permit identification of genes that modify the phenotypic manifestation of the EYA1 mutation. Also, they may provide insights into causes of the variable expressivity of human BOR syndrome. The mouse mutations also provide the means to study mutant EYA1 gene expression during embryonic development and to examine the role of EYA1 in molecular pathways leading to these common morphogenetic events.

Until recently, it was di cult to interpret whether syndromes with overlapping clinical phenotypes, such as BO, BOR, BOF (branchio-oculo-facial), BO, BOU (branchio-oto-uretral), or OFC (otofaciocervial), associated with branchial anomalies were caused by a single gene or more than one gene. After the localization of two loci associated with BOR syndrome and the identification of the EYA1 gene, several reports were published on BOF (50,51) and BO indicating genetic heterogeneity (62,64). However, these reports should be interpreted with caution where the undetected EYA1 mutation is not supported by genetic linkage studies. Recent reports indicate that BOF syndrome is not allelic to BO or BOR (50,51) and the OFC syndrome is a contiguous gene deletion syndrome involving the EYA1 gene (56).

Many of our BOR families screened for EYA1 do not show (approximately 60%) mutations in the EYA1 gene (33). It is possible that most of the mutations lie in the noncoding regions of the gene or involve deletions and/or complex rearrangements. More than 30 mutations have now been reported in the EYA1 gene (30–37). These mutations include nonsense, missense, frame shift, aberrant splicing, exon skipping, and large or small deletions. Most of the mutations are found in the highly conserved carboxyterminal region of the gene, indicating that this region may play a predominant role in the structure and/or function of the EYA1 gene.

IV. MUTATION IDENTIFICATION

Mutations in EYA1 have been reported in approximately one-third of persons with phenotypic features of BOR syndromes. Only a few mutations,