Resolving the Molecular Mechanisms of Inherited Deafness caused by Missense Mutations in Cadherin-23

Abstract

Cadherin-23 (cdh23) is a key component of hair cell tip links, fine filaments that directly convey mechanical force to mechanotransduction channels mediating inner-ear sensory perception. The extracellular domain of cdh23 is made of 27 extracellular cadherin (EC) repeats of similar fold and sequence with ∼100 amino acids each. There are over 50 missense mutations that target these EC repeats and that cause inherited deafness, yet the molecular mechanisms underlying dysfunction for many of them are unknown. We systematically analyzed and classified deafness mutations according to their location within cdh23 EC repeats, mapped them to cdh23 EC1+2, and then produced selected cdh23 EC1+2 mutants for biophysical studies. Here, we present the crystal structure of cdh23 EC1+2 carrying the R72W mutation, the most common among those observed in cdh23 EC repeats. The structure shows a fully occupied, canonical calcium-binding site despite local re-arrangement of the linker region. In addition, thermostability and calcium-binding assays for R72W and several other mutations reveal variable shifts in melting temperatures and changes in affinity for calcium. Our data suggest a hierarchy of biochemical effects that might correlate with the severity of the associated deafness phenotype.