Abstract
Previous studies have shown that gap junctional channels formed from the lens connexins Cx50 (or its chicken orthologue, Cx45.6) and Cx43 exhibit marked differences in transjunctional voltage gating and unitary conductance. In the present study, we used the negatively charged dye, Lucifer Yellow (LY), to examine and compare quantitative differences in dye transfer between pairs of HeLa cells stably transfected with Cx45.6 or Cx43. Our results show that Cx45.6 gap junctional channels are three times less permeable to LY than Cx43 channels. Replacement of the N-terminus of Cx45.6 with the corresponding domain of Cx43 increased LY permeability, reduced the transjunctional voltage (V(j)) gating sensitivity, and reduced the unitary conductance of Cx45.6-43N gap junctional channels. Further experiments, using a series of Alexa probes that had similar net charge but varied in size showed that the Cx45.6-43N had a significantly higher permeability for the two largest Alexa dyes than Cx45.6. These data suggest that the N-terminus plays a critical role in determining many of biophysical properties of Cx45.6 gap junctional channels, including molecular permeability and voltage gating.