Cytosolic pH regulates GCl through control of phosphorylation states of CFTR

Our objective inthis study was to determine the effect of changes in luminal andcytoplasmic pH on cystic fibrosis transmembrane regulator (CFTR)Cl conductance(G_Cl). Wemonitored CFTRG_Cl in the apicalmembranes of sweat ducts as reflected byCl diffusion potentials(V_Cl) andtransepithelial conductance(G_Cl). We foundthat luminal pH (5.0-8.5) had little effect on thecAMP/ATP-activated CFTRG_Cl, showing thatCFTR G_Cl ismaintained over a broad range of extracellular pH in which it functionsphysiologically. However, we found that phosphorylation activation ofCFTR G_Cl issensitive to intracellular pH. That is, in the presence of cAMP and ATP [adenosine5'-O-(3-thiotriphosphate)],CFTR could be phosphorylated at physiological pH (6.8) but not at lowpH (~5.5). On the other hand, basic pH prevented endogenousphosphatase(s) from dephosphorylating CFTR.After phosphorylationof CFTR with cAMP and ATP, CFTRG_Cl is normallydeactivated within 1 min after cAMP is removed, even in the presence of5 mM ATP. This deactivation was due to an increase in endogenousphosphatase activity relative to kinase activity, since it was reversedby the reapplication of ATP and cAMP. However, increasing cytoplasmicpH significantly delayed the deactivation of CFTRG_Cl in adose-dependent manner, indicating inhibition of dephosphorylation. Weconclude that CFTRG_Cl may beregulated via shifts in cytoplasmic pH that mediate reciprocal controlof endogenous kinase and phosphatase activities. Luminal pH probably has little direct effect on these mechanisms. This regulation of CFTRmay be important in shifting electrolyte transport in the duct fromconductive to nonconductive modes.