Increases in cytosolic Ca 2+ induce dynamin- and calcineurin-dependent internalisation of CFTR

The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-regulated, apical anion channel that regulates ion and fluid transport in many epithelia including the airways. We have previously shown that cigarette smoke (CS) exposure to airway epithelia causes a reduction in plasma membrane CFTR expression which correlated with a decrease in airway surface hydration. The effect of CS on CFTR was dependent on an increase in cytosolic Ca . However, the underlying mechanism for this Ca -dependent, internalisation of CFTR is unknown. To gain a better understanding of the effect of Ca on CFTR, we performed whole cell current recordings to study the temporal effect of raising cytosolic Ca on CFTR function. We show that an increase in cytosolic Ca induced a time-dependent reduction in whole cell CFTR conductance, which was paralleled by a loss of cell surface CFTR expression, as measured by confocal and widefield fluorescence microscopy. The decrease in CFTR conductance and cell surface expression were both dynamin-dependent. Single channel reconstitution studies showed that raising cytosolic Ca per se had no direct effect on CFTR. In fact, the loss of CFTR plasma membrane activity correlated with activation of calcineurin, a Ca -dependent phosphatase, suggesting that dephosphorylation of CFTR was linked to the loss of surface expression. In support of this, the calcineurin inhibitor, cyclosporin A, prevented the Ca -induced decrease in cell surface CFTR. These results provide a hitherto unrecognised role for cytosolic Ca in modulating the residency of CFTR at the plasma membrane through a dynamin- and calcineurin-dependent mechanism.