Introduction: Sensorineural hearing loss is the most common sensory disorder in humans. Genetic analyses have greatly increased our understanding of the pathogenic mechanisms in play. Thus, characterization of audiologic phenotypes by the genetic etiology may aid elucidation of the etiologies of certain types of inherited hearing loss. Further, delineation of specific audiologic phenotypes based on the genetic etiology aids our understanding of some types of inherited hearing loss in terms of the prediction of clinical course, revelation of genotype-phenotype correlations, and application of appropriate audiologic rehabilitation. Here, we describe the interesting audiologic characteristics of LMX1A -associated deafness, which revealed significant asymmetry between two ears.

Methods: Among 728 probands of which genomic DNA went through exome sequencing regardless of any specific audiologic phenotypes, probands for which exome sequencing was performed and a causative LMX1A variant was found were all included. Five LMX1A -associated DFNA7 families (approximately 0.7%), the pedigrees of whom indicated autosomal-dominant hearing loss, were identified, and segregation was studied using Sanger sequencing. The affected individuals underwent comprehensive evaluations, including medical history reviews, physical examinations, imaging, and auditory phenotyping. We functionally characterized the novel LMX1A variants via computational structural modeling and luciferase reporter assays.

Results: Among 728 probands of which genomic DNA went through exome sequencing, we identified four novel LMX1A heterozygous variants related to DFNA7 (c.622C>T:p.Arg208*, c.719A>G:p.Gln240Arg, c.721G>A:p.Val241Met, and c.887dup:p.Gln297Thrfs*41) and one harboring a de novo heterozygous missense LMX1A variant (c.595A>G;p.Arg199Gly) previously reported. It is important to note that asymmetric hearing loss was identified in all probands and most affected individuals, although the extent of asymmetry varied. Structural modeling revealed that the two missense variants, p.Gln240Arg and p.Val241Met, affected conserved residues of the homeodomain, thus attenuating LMX1A-DNA interaction. In addition, Arg208*-induced premature termination of translation destroyed the structure of the LMX1A protein, including the DNA-binding homeodomain, and p.Gln297Thrfs*41 led to the loss of the C-terminal helix involved in LIM2 domain interaction. Compared with the wild-type protein, all mutant LMX1A proteins had significantly reduced transactivation efficiency, indicating that the ability to elicit transcription of the downstream target genes of LMX1A was severely compromised. Thus, in line with the American College of Medical Genetics and Genomics guideline specified to genetic hearing loss, the four novel LMX1A variants were identified as "pathogenic" (p.Arg208* and p.Gln297Thrfs*41), "likely pathogenic" (p.Val241Met), and as a "variant of uncertain significance'' (p.Gln240Arg).

Conclusion: For the first time, we suggest that LMX1A is one of the candidate genes which, if altered, could be associated with dominantly inherited asymmetric hearing loss. We also expand the genotypic spectrum of disease-causing variants of LMX1A causing DFNA7 by doubling the number of LMX1A variants reported thus far in the literature.