PKHD1L1, A Gene Involved in the Stereocilia Coat, Causes Autosomal Recessive Nonsyndromic Hearing Loss

Abstract

AbstractIdentification of genes associated with nonsyndromic hearing loss is a crucial endeavor given the substantial number of individuals who remain without a diagnosis after even the most advanced genetic testing.PKHD1L1was established as necessary for the formation of the cochlear hair-cell stereociliary coat and causes hearing loss in mice and zebrafish when mutated. We sought to determine if biallelic variants inPKHD1L1also cause hearing loss in humans.Exome sequencing was performed on DNA of four families segregating autosomal recessive nonsyndromic sensorineural hearing loss. Compound heterozygous p.[(Gly129Ser)];p.[(Gly1314Val)] and p.[(Gly605Arg)];p[(Leu2818TyrfsTer5)], homozygous missense p.(His2479Gln) and nonsense p.(Arg3381Ter) variants were identified inPKHD1L1that were predicted to be damaging usingin silicopathogenicity prediction methods.In vitrofunctional analysis of two missense variants was performed using purified recombinant PKHD1L1 protein fragments. We then evaluated protein thermodynamic stability with and without the missense variants found in one of the families and performed a minigene splicing assay for another variant.In silicomolecular modelling using AlphaFold2 and protein sequence alignment analysis were carried out to further explore potential variant effects on structure.In vitrofunctional assessment indicated that both engineered PKHD1L1 p.(Gly129Ser) and p.(Gly1314Val) mutant constructs significantly reduced the folding and structural stabilities of the expressed protein fragments, providing further evidence to support pathogenicity of these variants. Minigene assay of the c.1813G>A p.(Gly605Arg) variant, located at the boundary of exon 17, revealed exon skipping leading to an in-frame deletion of 48 amino acids.In silicomolecular modelling exposed key structural features that might suggest PKHD1L1 protein destabilization.Multiple lines of evidence collectively associatePKHD1L1with nonsyndromic mild-moderate to severe sensorineural hearing loss.PKHD1L1testing in individuals with mild-moderate hearing loss may identify further affected families.