Unveiling a novel GJB2 dominant K22T mutation in a Chinese family with hearing loss

Abstract

Hearing loss constitutes one of the most prevalent conditions within the field of otolaryngology. Recent investigations have revealed that mutations in deafness-associated genes, including point mutations and variations in DNA sequences, can cause hearing impairments. With the ethology of deafness remaining unclear for a substantial portion of the affected population, further screenings for pathogenic mutations are imperative to unveil the underlying mechanisms. On this study, by using next-generation sequencing, we examine 129 commonly implicated deafness-related genes in a Chinese family with hearing loss, revealing a novel heterozygous dominant mutation in the GJB2 gene (GJB2: c.65T>G: p. Lys22Thr). This mutation consistently occurs in affected family members but is not detected in unaffected individuals, strongly suggesting its causative role in hearing loss. Structural analysis indicates potential disruption to the Cx26 gap junction channel's hydrogen bond and electrostatic interactions, aligning with predictions from the PolyPhen and SIFT algorithms. In conclusion, our study provides conclusive evidence that the identified heterozygous GJB2 mutation (GJB2: c.65T>G: p. Lys22Thr), specifically the K22T alteration, is the primary determinant of the family's deafness. This contribution enhances our understanding of the interplay between common deafness-associated genes and hearing loss, offering valuable insights for diagnostic guidance and the formulation of therapeutic strategies for this condition.

Keywords: deafness; next-generation sequencing; point mutations.

Figure 1

Pedigree and PTA of the Chinese family The circle symbols represent women and squares indicate men. Open symbols mean unaffected patients and close symbols mean affected patients. The proband is indicated by an arrow and deceased persons are indicated by a diagonal line through the symbol. (A–F) The PTA was detected in affected members in the Chinese family. Frequency in hertz (Hz) is plotted on the x-axis and the auditory threshold in decibels (dB) on the y-axis. Round, right ear; square, left ear.

Figure 2

Mutation validation, conservation analysis and protomer structure of GJB2 (A) The identified mutation was confirmed by Sanger sequencing. The boxes indicate the location of the nucleotide changes. (B) Sequence alignment showing the Lys22 is a conserved residue among different species. NTH, N-terminal helix; TM1~TM4, transmembrane helices1~4; E1~E2, extracellular loops 1~2. (C) Topological diagram of the Cx26. The mutated site is indicated by arrow.

Figure 3

Structure views of Cx26, Cx30.3 and their mutations (A) Sequence alignment of Cx26 and Cx30. The sequence similarity is 71.9% while the sequence identity is 51.1%. (B) Structure views of Cx26 (left) and mutant Cx26-K22T (right). The mutant structure was constructed using Discover Studio. Main residues surrounding Lys22 within 10 Å, in grey; hydrogen bonds, green dotted lines. The H-bond between Tyr136 and Lys22 disappears when Lys22 is mutated into Thr22. The substitution of lysine with threonine also disrupts the possible electrostatic interaction between Lys22 and the neighbor residue Glu209. (C) Structure views of Cx30.3 (left) and Cx30.3-E204A (right). The homologous structure of Cx30.3 was constructed using SWISS-MODEL server. Further mutation was performed in Discover Studio. Main residues surrounding Glu204 within 10 Å. The H-bonds between Glu204 and the neighbor residues Ser21, Arg22 and Leu 200 are absent in E204A. The substitution of glutamate with alanine also disrupts the electrostatic interaction between Glu204 and the neighbor residue Arg22.