Molecular Determinants Underlying the Pathogenic Mechanism of Kid Syndrome Elicited by Cx26G12R Mutation

Abstract

Several mutations in Cx26 promote keratitis ichthyosis deafness (KID), a rare disease in which patients exhibit deafness, skin and corneal disorders. Some KID mutations are located in the amino terminal segment of Cx26, which has been proposed as the putative voltage sensor. Most syndromic mutations elicit a gain of hemichannel function, however the mechanism underlying this phenomenon remains unclear. Here we examined the biophysical properties of the mutant G12R at macroscopic and single channel levels. We perform two-electrode voltage clamp to study the deactivation kinetics and apparent affinity for calcium of hemichannel currents. In contrast to wild-type (wt) Cx26, we found that the activation kinetics of the mutant lack saturation of the current upon depolarization pulses above +40 mV. Estimates of the deactivation time constant yield values of 5s and 10s for G12R and wtCx26 hemichannels, respectively; however the apparent affinity for calcium remains similar to wtCx26. Single channel recordings analysis showed that the mutant virtually eliminates the transitions to the subconductance state. Moreover, although the single channel conductance was similar to wtCx26 hemichannels (320 pS), the open probability and mean open times of G12R hemichannels become larger. Molecular dynamics indicates that the G12R induce an electrostatic effect and ddG calculations support protein destabilization.Supported by Fondecyt-3150634 to IEG; Fondecyt-3140590 to GC; Fondecyt-1130855 to ADM; Anillo-ACT1104 to ADM and CG; Fondecyt-1120802 to CG; Grant RO1-GM099490 to J.E.C; AP has a Conicyt Fellowship. The Centro Interdisciplinario de Neurociencias de Valparaiso is a Chilean Millennium Institute (P09-022-F).