Heterotypic docking of Cx43 and Cx45 connexons blocks fast voltage gating of Cx43

Immunohistochemical co-localization of distinct connexins (Cxs) in junction al areas suggests the formation of heteromultimeric channels. To determine the docking effects of the heterotypic combination of Cx43 and Cx45 on the voltage-gating properties of their channels, we transfected DNA encoding Cx 43 or Cx45 into N2A neuroblastoma or HeLa cells. Using a double whole-cell voltage-clamp technique, we determined macroscopic and single-channel gatin g properties of the intercellular channels formed. Cx43-Cx45 heterotypic ch annels had rectifying properties where Cx45 connexons inactivated rapidly u pon hyperpolarizing voltage pulses applied to the Cx45-expressing cell. Dur ing depolarizing pulses to the Cx45-expressing cell, Cx43 connexons inactiv ated with substantially reduced kinetics as compared with homotypic Cx43 ch annels. Similar slow kinetics was observed for homotypic Cx43M257 (truncati on mutant). Heterotypic channels had a main conductance whose value was pre dicted by the sum of corresponding homomeric connexon conductances; it was not voltage dependent and had no detectable residual conductance. The volta ge-gating kinetics of heterotypic channels and their single-channel behavio r implicate a role for the Cx43 carboxyl-terminal domain in the fast gating mechanism and in the establishment of residual conductance. Our results al so suggest that heterotypic docking may lead to conformational changes that inhibit this action of the Cx43 carboxyl-terminal domain.