Ann Lab Med.  2020 May;40(3):224-231. 10.3343/alm.2020.40.3.224.

A Novel Heterozygous Missense Variant (c.667G>T;p.Gly223Cys) in USH1C That Interferes With Cadherin-Related 23 and Harmonin Interaction Causes Autosomal Dominant Nonsyndromic Hearing Loss

  • 1GC Genome, Yongin, Korea.
  • 2Unité de génétique et physiologie de l'audition, Institut Pasteur, Paris, France.
  • 3UMRS 1120, Inserm, Paris, France.
  • 4Sorbonne Universités, Paris, France.
  • 5Department of Otolaryngology - Head and Neck Surgery, Stanford University, Stanford, California, USA.
  • 6College de France and Institut Pasteur, Paris, France.
  • 7Department of Ophthalmology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
  • 8Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
  • 9Korean Bioinformation Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.
  • 10Department of Bioinformatics, University of Sciences and Technology, Daejeon, Korea.


Pathogenic variants of USH1C, encoding a PDZ-domain-containing protein called harmonin, have been known to cause autosomal recessive syndromic or nonsyndromic hearing loss (NSHL). We identified a causative gene in a large Korean family with NSHL showing a typical pattern of autosomal dominant (AD) inheritance.
Exome sequencing was performed for five affected and three unaffected individuals in this family. Following identification of a candidate gene variant, segregation analysis and functional studies, including circular dichroism and biolayer interferometry experiments, were performed.
A novel USH1C heterozygous missense variant (c.667G>T;p.Gly223Cys) was shown to segregate with the NSHL phenotype in this family. This variant affects an amino acid residue located in the highly conserved carboxylate-binding loop of the harmonin PDZ2 domain and is predicted to disturb the interaction with cadherin-related 23 (cdh23). The affinity of the variant PDZ2 domain for a biotinylated synthetic peptide containing the PDZ-binding motif of cdh23 was approximately 16-fold lower than that of the wild-type PDZ2 domain and that this inaccessibility of the binding site was caused by a conformational change in the variant PDZ2 domain.
A heterozygous variant of USH1C that interferes with the interaction between cdh23 and harmonin causes novel AD-NSHL.


Harmonin; Nonsyndromic hearing loss; USH1C; Heterozygous variant

MeSH Terms

Binding Sites
Circular Dichroism
Hearing Loss*


  • Fig. 1 Clinical and genetic findings in a Korean family with NSHL. (A) Pedigree of a large Korean family with HL; the proband is indicated with an arrow. Individuals who participated in the molecular genetic testing are marked with an asterisk on the right. Whole exome sequencing was performed for eight individuals (shaded in yellow). The presence (+) or absence (−) of the USH1C c.667G>T variant is indicated underneath each individual. (B) Fundus autofluorescence photographs and (C) full-field electroretinogram showing normal findings. (D) Pure tone audiometry result showing moderate to moderately severe HL, with an average of 53 and 56 dB for the right and left ears, respectively. (E) Caloric test result showing normal vestibular function bilaterally. (F) Sequencing chromatograms of the c.667G>T variant of USH1C in individuals who participated in the molecular genetic testing. Nomenclature is based on RefSeq NM_005709.3, with nucleotide number +1 designating the A of the start codon ATG.Abbreviations: HL, Hearing loss; NSHL, nonsyndromic hearing loss; Lt, left; Rt, right.

  • Fig. 2 Functional analysis and protein modeling of harmonin. (A) The affinity of the WT PDZ2 (Kd=37.6±4.1 µM) was approximately 16-fold higher than that of PDZ2 carrying the p.Gly223Cys variant (Kd=621±88 µM). (B) Circular dichroism spectra for PDZ2 WT and the PDZ2 p.Gly223Cys variant demonstrated the global secondary structure of the WT and variant PDZ2 domain and showed that the p.Gly223Cys variant causes a conformational change. (C) The normalized elution profile of the full-length p.Gly223Cys variant protein showed a left-shifted peak compared with WT full-length harmonin, and a mixture of the full-length p.Gly223Cys variant and WT harmonin revealed a further left-shifted peak. (D) The structure of the PDZ2 domain of harmonin in a complex with the PBM of cdh23 and conformational analysis of the p.Gly223Cys variant demonstrated the interaction between the PDZ2 domain of harmonin and the PDZ-binding motif of cdh23 through the GLGF motif, a highly conserved carboxylate-binding loop in a groove between the αB and βB structural elements (left). The third glycine residue in the loop region is fully conserved and adopts a left-handed α-helical conformation. Replacement of the glycine residue with cysteine leads to a conformational change with a structural modification (right).Abbreviations: WT, wild-type; SEC, size-exclusion chromatography; PBM, PDZ-binding motif.


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