Expression and Characterization of Wildtype and Mutant Forms of Human Connexin31 using Xenopus Oocytes

Abstract

Connexin31 is a gap junction protein encoded by the GJB3 gene in humans. Mutations in the gene lead to a rare skin disorder known as erythrokeratodermia variabilis (EKV) a disease characterized by figurate red patches and general hyperkeratosis. While a few mutations have also been linked to deafness and neuropathy, over twenty mutations, mostly point mutations within the coding region, have been associated with EKV. Oocytes from Xenopus laevis were used for expression and analysis of Cx31 and mutations. As far as we know this is the first time Cx31 has been expressed and characterized in oocytes. Wildtype Cx31 displayed gating properties similar to those reported after expression in mammalian cells but with some interesting differences. Several skin disease mutations induced leaky membranes associated with cell death, an effect that was both time- and concentration-dependent. It was hypothesized that aberrant hemichannel behavior was responsible for leaky membranes, which was supported by the observation that addition of calcium or cobalt to the extracellular media enhanced viability. Other mutations induced behavior ranging from failure to form functional channels to expression and gating similar to wtCx31. As well as providing insight into the mechanisms underlying skin disease, deafness and neuropathy, we will discuss the role of Cx31 mutants as tools to assess connexin interactions in skin.