MODULATION OF CFTR CHLORIDE CHANNELS BY CALYCULIN-A AND GENISTEIN

Modulation of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel by calyculin A and genistein was studied in Hi-5 in sect cells infected with baculovirus containing the wild-type CFTR cDN A. In cell-attached patches, CFTR channel activity was not observed un til stimulated by forskolin in 90% of the cells, suggesting a low leve l of basal adenosine 3',5'-cyclic monophosphate activity. Calyculin A, a specific inhibitor of phosphatases 1 and 2A, increased forskolin-in duced CFTR activity by 17.2-fold. CFTR channel currents did not deacti vate completely after forskolin was withdrawn in the continued presenc e of calyculin A. Genistein enhanced forskolin-induced CFTR activity b y 44.9-fold but could neither activate the CFTR by itself nor prevent complete deactivation on removal of forskolin. Genistein together with calyculin A could adequately prevent deactivation of CFTR currents. N oise analysis of the macroscopic CFTR currents revealed significant di fferences in the mean current-variance relationship and the corner fre quency of the noise spectra between currents activated by forskolin pl us genistein and those activated by forskolin plus calyculin A. Furthe rmore, genistein enhanced CFTR activity induced by saturating concentr ations of forskolin and calyculin A. Our results suggest that genistei n and calyculin A modulate the CFTR by different mechanisms and that g enistein might inhibit calyculin A-insensitive dephosphorylation of th e CFTR.