Submitted on July 19, 2004
Accepted on January 23, 2005
Inversion of both gating polarity and CO2 sensitivity of voltage gating with D3N mutation of connexin50
Camillo Peracchia and Lillian L. Peracchia*
* To whom correspondence should be addressed. E-mail: lillian_peracchia{at}urmc.rochester.edu.
The effect of CO2-induced acidification on transjunctional voltage (Vj) gating was studied by dual voltage-clamp in oocytes expressing mouse Cx50 or a Cx50 mutant (Cx50-D3N) in which the third residue, aspartate (D), was mutated to asparagine (N). This mutation inverted the gating polarity of Cx50 from positive to negative. CO2 application greatly decreased the Vj sensitivity of Cx50 channels, and increased that of Cx50-D3N channels. CO2 also affected the kinetics of Vj dependent inactivation of junctional current (Ij), decreasing the gating speed of Cx50 channels and increasing that of Cx50-D3N channels. In addition, the D3N mutation increased the CO2 sensitivity of chemical gating such that even CO2 concentrations as low as 2.5% significantly lowered junctional conductance (Gj). With Cx50 channels Gj dropped by 78% with a drop in pHi to 6.83, whereas with Cx50-D3N channels Gj dropped by 95% with a drop in pHi to just 7.19. We have previously hypothesized that the way in which Vj gating reacts to CO2 may be related to the gating polarity of the connexin. This hypothesis is confirmed here by evidence that the D3N mutation inverts the gating polarity as well as the effect of CO2 on Vj gating sensitivity and speed.
