Is the chemical gate of connexins voltage sensitive? Behavior of Cx32 wild-type and mutant channels

Connexin channels are gated by transjunctional voltage (V-j) or CO2 via dis tinct mechanisms. The cytoplasmic loop (CL) and arginines of a COOH-termina l domain (CT1) of connexin32 (Cx32) were shown to determine CO2 sensitivity , and a gating mechanism involving CL-CT1 association-dissociation was prop osed. This study reports that Cx32 mutants, tandem, 5R/E, and 5R/N, designe d to weaken CL-CT1 interactions, display atypical V-j and CO2 sensitivities when tested heterotypically with Cx32 wild-type channels in Xenopus oocyte s. In tandems, two Cx32 monomers are linked NH2-to-COOH terminus. In 5R/E a nd 5R/N mutants, glutamates or asparagines replace CT1 arginines. On the ba sis of the intriguing sensitivity of the mutant-32 channel to V-j polarity, the existence of a "slow gate" distinct from the conventional V-j gate is proposed. To a lesser extent the slow gate manifests itself also in homotyp ic Cx32 channels. Mutant-32 channels are more CO2 sensitive than homotypic Cx32 channels, and CO2-induced chemical gating is reversed with relative de polarization of the mutant oocyte, suggesting V-j sensitivity of chemical g ating. A hypothetical pore-plugging model involving an acidic cytosolic pro tein (possibly calmodulin) is discussed.