Prolonged treatment of cells with genistein modulates the expression and function of the cystic fibrosis transmembrane conductance regulator

Background and purpose: Cystic fibrosis (CF) is caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl¯ channel. In the search for new CF therapies, small molecules have been identified that rescue the defective channel gating of CF mutants (termed CFTR potentiators). Here, we investigate the long-term effects of genistein, the best-studied CFTR potentiator, on the expression and function of CFTR. Experimental approach: We pre-treated baby hamster kidney (BHK) cells expressing wild-type or F508del-CFTR (the most common CF mutant) with concentrations of genistein that potentiate (30 μM) or inhibit (100 μM) CFTR function for 2 or 24 h at 37 ºC before examining CFTR maturation, expression and single-channel activity. Key results: Using the iodide efflux technique, we found that genistein pre-treatment failed to restore function to F508del-CFTR, but altered that of wild-type CFTR. Pre-treatment of cells with genistein for 2 h had little effect on CFTR processing, whereas pre-treatment for 24 h either augmented (30 μM genistein) or impaired (100 μM genistein) CFTR maturation. Using immunocytochemistry, we found that all genistein pre-treatments increased the localization of CFTR protein to the cell surface. However, following the incubation of cells with genistein (100 μM) for 2 h, individual CFTR Cl¯ channels exhibited characteristics of channel block upon channel activation. Conclusions and implications: Genistein pre-treatment alters the maturation, cell surface expression and single-channel function of CFTR in ways distinct from its acute effects. Thus, CFTR potentiators have the potential to influence CFTR by mechanisms distinct from their effects on channel gating.