GENISTEIN POTENTIATES WILD-TYPE AND DELTA-F508-CFTR CHANNEL ACTIVITY

Effects of genistein on wildtype (wt) and Delta F50S-cystic fibrosis t ransmembrane conductance regulator (CFTR) were studied in NIH/3T3 cell s stably transfected with wt or mutant CFTR cDNA. As measured by I- ef flux, half-maximal concentration of agonist (K-1/2) for forskolin-depe ndent activation was greater for Delta F508-CFTR than wt-CFTR. Geniste in decreased the K-1/2 for both forms of the channel and increased the maximal activity of Delta F508-CFTR by 3.7-fold. In cell-attached pat ches, 10 mu M forskolin induced minimal Delta F508-CFTR, activity with characteristic prolonged closed times (estimated time constant, >30 s ). Genistein increased the forskolin-induced macroscopic currents of w t-CFTR and Delta F508-CFTR by 3- and 19-fold, respectively. Variance a nalysis suggested that in the presence of forskolin and genistein the open probabilities (P-o) of Wt- and Delta F508-CFTR were identical. In single-channel studies, at maximal adenosine 3',5'-cyclic monophospha te (cAMP) stimulation, genistein increased the P-o of wt-CFTR by prolo nging the open time, but, at submaximal cAMP stimulation, the P-o was increased by prolonging the open time and shortening the closed time. In excised patches with CFTR channels preactivated in the cell-attache d mode, genistein increased ATP-dependent wt-and Delta F508-CFTR curre nt about twofold hg prolonging the open time. Our results thus suggest that phosphorylation-dependent activation of Delta F508-CFTR is defec tive and that genistein corrects this defect at least in part by bindi ng to the CFTR protein.