Enlargement of the vestibular aqueduct
(EVA; MIM 600709) is a common radiological inner ear malformation associated with sensorineural hearing loss (Valvassori and Clemis, 1978
). EVA is the radiologic finding of highest penetrance in the syndrome of recessive sensorineural hearing loss with goiter (Pendred syndrome [PDS]; MIM 274600) but can also be observed without goiter in nonsyndromic EVA (NSEVA) (DFNB4; MIM 600791) (Phelps et al.
). Mutations of SLC26A4
, which encodes pendrin, can cause either PDS or NSEVA (Everett et al.
; Li et al.
; Usami et al.
). Pendrin, a polytopic transmembrane protein, exchanges Cl−
across the apical membrane of thyroid follicular cells (Royaux et al.
; Gillam et al.
). It is also expressed in the inner ear, where it has been proposed to exchange Cl−
(Wangemann et al.
). Pendrin is also expressed in type B intercalated cells of the renal collecting duct (Royaux et al.
), and in respiratory and mammary epithelial cells (Rillema and Hill, 2003
; Pedemonte et al.
), although its absence in these tissues is not known to contribute to the phenotype of PDS.
The overlap of SLC26A4
genotypes and phenotypes in PDS and NSEVA has led to uncertainty in their nosology. This uncertainty is compounded by the detection of only one recessive mutant allele of SLC26A4
in many EVA patients and by the uncertain pathogenic potential of many SLC26A4
variants (Choi et al.
). Many of these variants are missense substitutions whose pathogenic potential may be clarified by evaluation of their genotypic and phenotypic context in the EVA patient as well as by functional characterization of their intracellular trafficking and anion exchange properties (Choi et al.
The expense of bidirectional DNA sequencing of all 21 exons of SLC26A4
and their adjacent splice sites is an obstacle to widespread molecular diagnostic testing of EVA patients. Alternative methods include denaturing high-performance liquid chromatography (DHPLC) (Prasad et al.
; Albert et al.
) and exon-specific strategies (Park et al.
). The latter can be incorporated into hierarchical screens that account for different identities and distributions of mutations of SLC26A4
among different ethnic populations (Tsukamoto et al.
; Park et al.
; Pryor et al.
; Wu et al.
; Albert et al.
; Wang et al.
). This type of strategy has been proposed for the comparatively restricted distribution of SLC26A4
mutations prevalent in East Asian EVA subjects (Park et al.
; Wu et al.
; Dai et al.
; Guo et al.
The purpose of this study was to refine the known SLC26A4 mutation spectrum in Korean EVA patients and to develop a DHPLC screen to detect these mutations. We show that a DHPLC screen can be integrated with direct nucleotide sequencing into a cost-effective hierarchical protocol for the detection of SLC26A4 mutations in Korean EVA patients. We validate the protocol by a simulated analysis of published EVA cohorts with known SLC26A4 genotypes from a variety of East Asian populations. We also demonstrate an evaluation of pathogenicity of two interesting novel missense substitutions, one of which is the second most frequent missense variant from our Korean cohort in this study.