REGULATION OF MURINE CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR CL- CHANNELS EXPRESSED IN CHINESE-HAMSTER OVARY CELLS

1. We investigated the effect of protein kinases and phosphatases on m urine cystic fibrosis transmembrane conductance regulator (CFTR) Cl- c hannels, expressed in Chinese hamster ovary (CHO) cells, using iodide efflux and the excised inside-out configuration of the patch-clamp tec hnique. 2. The protein kinase C (PKC) activator, phorbol dibutyrate, e nhanced cAMP-stimulated iodide efflux. However, PKC did not augment th e single-channel activity of either human or murine CFTR Cl- channels that had previously been activated by protein kinase A. 3. Fluoride, a non-specific inhibitor of protein phosphatases, stimulated both human and murine CFTR Cl- channels. However, calyculin A, a potent inhibito r of protein phosphatases 1. and 2A, did not enhance cAMP-stimulated i odide efflux. 4. The alkaline phosphatase inhibitor, (-)-bromotetramis ole augmented cAMP-stimulated iodide efflux and, by itself, stimulated a larger efflux than that evoked by cAMP agonists. However, (+)-bromo tetramisole, the inactive enantiomer, had the same effect.. For murine CFTR, neither enantiomer enhanced single-channel activity. In contras t, both enantiomers increased the open probability (P-o) of human CPTR , suggesting that bromotetramisole may promote the opening of human CF TR. 5. As murine CFTR had a low P-o and was refractory to stimulation by activators of human CFTR, we investigated whether murine CFTR may o pen to a subconductance state. When single-channel records were filter ed at 50 Hz, a very small subconductance state of murine CFTR was obse rved that had a P-o greater than that of human CFTR. The occupancy of this subconductance state may explain the differences in channel regul ation observed between human and murine CFTR.