Conductance and permeability of the residual state of connexin43 gap junction channels.
We used cell lines expressing wild-type connexin43 and connexin43 fused with the enhanced green fluorescent protein (Cx43-EGFP) to examine conductance and perm-selectivity of the residual state of Cx43 homotypic and Cx43/Cx43-EGFP heterotypic gap junction channels. Each hemichannel in Cx43 cell-cell channel possesses two gates: a fast gate that closes channels to the residual state and a slow gate that fully closes channels; the transjunctional voltage (V(j)) closes the fast gate in the hemichannel that is on the relatively negative side. Here, we demonstrate macroscopically and at the single-channel level that the I-V relationship of the residual state rectifies, exhibiting higher conductance at higher V(j)s that are negative on the side of gated hemichannel. The degree of rectification increases when Cl(-) is replaced by Asp(-) and decreases when K(+) is replaced by TEA(+). These data are consistent with an increased anionic selectivity of the residual state. The V(j)-gated channel is not permeable to monovalent positively and negatively charged dyes, which are readily permeable through the fully open channel. These data indicate that a narrowing of the channel pore accompanies gating to the residual state. We suggest that the fast gate operates through a conformational change that introduces positive charge at the cytoplasmic vestibule of the gated hemichannel, thereby producing current rectification, increased anionic selectivity, and a narrowing of channel pore that is largely responsible for reducing channel conductance and restricting dye transfer. Consequently, the fast V(j)-sensitive gating mechanism can serve as a selectivity filter, which allows electrical coupling but limits metabolic communication.
Bukauskas
Feliksas F
FF
Department of Neuroscience, Albert Einstein College of Medicine, Bronx, NY 10461, USA. fbukausk@aecom.yu.edu
Bukauskiene
Angele
A
Verselis
Vytas K
VK
D000818:Animals; D018031:Connexin 43 / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D004553:Electric Conductivity; D005456:Fluorescent Dyes / Q000378:metabolism; D017629:Gap Junctions / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D015640:Ion Channel Gating / Q000235:genetics / Q000502:physiology; D008564:Membrane Potentials / Q000235:genetics / Q000502:physiology; D010539:Permeability; D051381:Rats; D014162:Transfection; D014407:Tumor Cells, Cultured
D018031:Connexin 43; D005456:Fluorescent Dyes