Evaluation of the Contribution of the <i>EYA4</i> and <i>GRHL2</i> Genes in Korean Patients with Autosomal Dominant Non-Syndromic Hearing Loss

<div><p><i>EYA4</i> and <i>GRHL2</i> encode transcription factors that play an important role in regulating many developmental stages. Since <i>EYA4</i> and <i>GRHL2</i> were identified as the transcription factors for the DFNA10 and DFNA28, 8 <i>EYA4</i> mutations and 2 <i>GRHL2</i> mutations have been reported worldwide. However, these genes have been reported in few studies of the Korean population. In this study, we performed a genetic analysis of <i>EYA4</i> and <i>GRHL</i>2 in 87 unrelated Korean patients with autosomal dominant non-syndromic hearing loss (NSHL). A total of 4 genetic variants in the <i>EYA4</i> gene were identified, including the 2 nonsense mutations p.S288X and p.Q393X. The novel mutation p.Q393X (c.1177C>T) resulted in a change in the codon at amino acid position 393 from a glutamine to a stop codon. The p.Q393X allele was predicted to encode a truncated protein lacking the entire C-terminal Eya homolog region (Eya HR), which is essential for the interaction with the transcription factor SIX3. The p.S288X (c.863C>A) mutation was found in a Korean family from a previous study. We analyzed p.S288X-linked microsatellite markers and determined that p.S288X might be a founder mutation and a hotspot mutation in Koreans. In <i>GRHL2</i>, a total of 4 genetic variants were identified, but none were associated with hearing loss in Korean patients. This suggests that <i>GRHL2</i> may not be a main causal gene for autosomal dominant NSHL in Korean patients. In conclusion, our data provide fundamental information to predict the genotypes of Korean patients diagnosed with autosomal dominant NSHL.</p></div

Mutation analysis of the <i>EYA4</i> gene in the YS-151 family.

<p>(A) Pedigree of a Korean family with autosomal dominant inheritance (upper panel). A three-generation pedigree that includes 8 members is presented. The filled symbols and open symbols indicate affected and unaffected individuals, respectively. The arrow designates the proband. Pure tone audiogram for the left and right ears of the YS-151 patient (III-2) (lower panel). The circles and crosses indicate unmasked air conduction thresholds for the right and left ears, respectively. (B) DNA sequencing analysis of <i>EYA4</i> exon 13 shows the c.1177C>T change in an affected family member (III-2) and a normal control. The arrow indicates the changed base. (C) Multiple alignments of the amino acid sequence encoded by the <i>EYA4</i> gene including the HR domain in vertebrate species. The arrow marks the position of the p.Q393X mutation.</p

Location of the p.S288X mutation in <i>EYA4</i> and linked STR markers on chromosome 6q.

<p>The physical map marks the location of 5 microsatellite markers and positions.</p

SNPs of the <i>GRHL2</i> gene identified in this study.

<p>SNPs of the <i>GRHL2</i> gene identified in this study.</p

Summary of <i>EYA4</i> gene mutations identified in previous and current studies.

<p>* N/A, Not available.</p><p>Summary of <i>EYA4</i> gene mutations identified in previous and current studies.</p

Mutation analysis of the <i>EYA4</i> gene in the HL-01 family.

<p>(A) Pedigree of the Korean family with autosomal dominant inheritance (upper panel). A three-generation pedigree including 10 members is presented. The filled symbols and open symbols indicate affected and unaffected individuals, respectively. The arrow designates the proband. Pure tone audiogram for the left and right ears of the HL-01 patient (III-2) (lower panel). The circles and crosses indicate unmasked air conduction thresholds for the right and left ears, respectively. (B) DNA sequencing analysis of <i>EYA4</i> exon 11 shows the c.863C>A change in an affected member (III-2) of the HL-01 family and a normal control. The arrow indicates the changed base. (C) Multiple alignments of the amino acid sequence encoded by the <i>EYA4</i> gene including the VR domain in vertebrate species. The arrow marks the position of the p.S288X mutation.</p

SNPs of the <i>CLDN14</i> gene identified in this study.

<p>*Novel variation of <i>CLDN14</i> gene identified in this study.</p

Clinical and genetic information relevant to the KNUF25 (a) and SY-149 (b) families with the p.T1188A mutation of the <i>TJP2</i> gene.

<p>(A) The top panel shows the pedigree and genotype of each family. The arrows indicate the proband (KNUF25 II-1, YS-149 II-6), and filled symbols represent affected individuals from these families. The asterisks denote individuals screened in this study. The bottom panel shows pure tone audiograms of affected individuals (KNUF25 II-1, KNUF25 I-2, YS-149 II-6). The red (circles) and black (crosses) lines represent unmasked air conduction thresholds for the right and left ears, respectively. (B) The top panel shows partial DNA sequences of the <i>TJP2</i> gene with the genotypes of affected and unaffected individuals within the family. The position of a changed nucleotide is indicated by a square. The bottom panel indicates the evolutionary conservation of amino acids, including threonine 1188, in vertebrate species. The arrow and box designate the location of the mutation.</p

Protein structures of the PDZ 1 domain, including protein position 112 in the <i>TJP2</i> protein.

<p>(A, B) Overview of the wild type (p.A112). (C, D) Differences between the alanine (C) and threonine (D) at amino acid position 112.</p

SNPs of the <i>TJP2</i> gene identified in this study.

<p>*Novel variation of <i>TJP2</i> gene identified in this study.</p