The conductance and organization of the TMC1-containing mechanotransducer channel complex in auditory hair cells.

Abstract

Transmembrane channel-like protein 1 (TMC1) is thought to form the ion-conducting pore of the mechanoelectrical transducer (MET) channel in auditory hair cells. Using single-channel analysis and ionic permeability measurements, we characterized six missense mutations in the purported pore region of mouse TMC1. All mutations reduced the Ca²⁺ permeability of the MET channel, triggering hair cell apoptosis and deafness. In addition, <i>Tmc1</i> p.E520Q and <i>Tmc1</i> p.D528N reduced channel conductance, whereas <i>Tmc1</i> p.W554L and <i>Tmc1</i> p.D569N lowered channel expression without affecting the conductance. <i>Tmc1</i> p.M412K and <i>Tmc1</i> p.T416K reduced only the Ca²⁺ permeability. The consequences of these mutations endorse TMC1 as the pore of the MET channel. The accessory subunits, LHFPL5 and TMIE, are thought to be involved in targeting TMC1 to the tips of the stereocilia. We found sufficient expression of TMC1 in outer hair cells of <i>Lhfpl5</i> and <i>Tmie</i> knockout mice to determine the properties of the channels, which could still be gated by hair bundle displacement. Single-channel conductance was unaffected in <i>Lhfpl5</i>^-/- but was reduced in <i>Tmie^-/-</i>, implying TMIE very likely contributes to the pore. Both the working range and half-saturation point of the residual MET current in <i>Lhfpl5</i>^-/- were substantially increased, suggesting that LHFPL5 is part of the mechanical coupling between the tip-link and the MET channel. Based on counts of numbers of stereocilia per bundle, we estimate that each PCDH15 and LHFPL5 monomer may contact two channels irrespective of location.