Differential modulation of unapposed connexin 43 hemichannel electrical conductance by protein kinase C isoforms.
Opening of unapposed connexin 43 hemichannels (Cx43Hc) in the plasma membrane results in altered ionic homeostasis leading to cell damage. Although it is generally acknowledged that Cx43Hc function is regulated by protein kinase C (PKC), information regarding the functional role of PKC in the modulation of Cx43Hc electrical conductance is lacking. In this work, we used the patch-clamp technique to study the effect of phorbol 12-myristate 13-acetate (PMA), a general PKC activator, on the electrical conductance of exogenous Cx43Hc expressed in tsA201 cells. Subsequently, a matrix of synthetic PKC isoform-specific inhibitor peptides was used to dissect the functional role of individual PKC isoforms in Cx43Hc regulation. Superfusion with 10 nM PMA abolished Cx43Hc currents by 74%, an effect that was prevented by pretreatment with a general PKC inhibitor, GF109203X. It is interesting to note that intracellular diffusion of epsilon V1-2 (0.1 microM), an epsilon PKC-specific inhibitor peptide, completely antagonized PMA-induced current inhibition. Cell dialysis with either beta II- or delta PKC inhibitor peptides partially decreased PMA effect. Neither alpha- nor beta I PKC inhibition altered PMA-induced current reduction. This study shows for the first time that Cx43Hc electrical conductance is inhibited after PKC activation. Moreover, this inhibition is predominantly mediated by the "novel" epsilon PKC isoform, whereas partial inhibition may be provided by the "conventional" beta II PKC as well as the "novel" delta PKC isoforms.
Hawat
G
G
Centre de Biomédecine, Hôpital du Sacré-Coeur de Montréal, Montréal, Québec, Canada.
Baroudi
G
G
D000818:Animals; D002461:Cell Line, Transformed; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism*; D003956:Dialysis; D004553:Electric Conductivity; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D004847:Epithelial Cells / Q000201:enzymology*; D006801:Humans; D007527:Isoenzymes; D008564:Membrane Potentials; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D064546:Protein Kinase C beta; D051745:Protein Kinase C-delta / Q000378:metabolism; D051744:Protein Kinase C-epsilon / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection
D018031:Connexin 43; D020536:Enzyme Activators; D007527:Isoenzymes; D047428:Protein Kinase Inhibitors; C497258:PRKCD protein, human; C497255:PRKCE protein, human; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D013755:Tetradecanoylphorbol Acetate