Modulation of cftr chloride channels by calyculin a and
genistein.
Yang, Iris C-H., Tzu-Hurng Cheng, Fei Wang, Elmer M. Price, and Tzyh
-Chang Hwang.
Departments of Physiology and Veterinary Biomedical Sciences#,
Dalton Cardiovascular Research Center, University of Missouri
-Columbia, Columbia, MO 65211; Department of Pharmacology, National
Defense Medical Center, Taipei, Taiwan
APStracts 3:0266C, 1996.
Regulation mechanisms of the CFTR Cl- channel were studied in Hi-5
insect cells infected with baculovirus containing the wild-type CFTR.
No CFTR-like Cl- channels were present in noninfected cells. In more
than 90% of infected cells that show CFTR activity, channel currents
were not observed until stimulated by forskolin presumably because of
a low level of basal CFTR phosphorylation in these cells. CFTR
channels expressed in insect cells show similar biophysical
properties as native channels. These include 11 pS single-channel
conductance and ATP-dependent gating. Calyculin A increased
forskolin-induced CFTR activity by 17.2 fold suggesting that
phosphatase 1 and/or 2A is involved in dephosphorylation of the CFTR.
Partial activity of CFTR channels was observed after forskolin was
withdrawn in the continuous presence of calyculin A. Genistein
enhanced forskolin-induced CFTR activity by 44.9 fold, but could
neither activate the CFTR by itself nor prevent complete deactivation
upon removal of forskolin. Noise analysis of the macroscopic CFTR
currents revealed significant differences in the mean
current/variance relationship and the corner frequency (fc) of the
noise spectra between currents activated by forskolin plus genistein
and those activated by forskolin plus calyculin A. Furthermore,
genistein enhanced CFTR activity induced by saturating concentrations
of forskolin and calyculin A; similarly, calyculin A further
increased CFTR activity induced by saturating concentrations of
forskolin and genistein. Genistein together with calyculin A could
adequately prevent deactivation of CFTR channels. Potentiation of
forskolin-induced CFTR activity by genistein was also observed in
NIH3T3 cells stably expressing the wild-type CFTR. Our results
suggest that genistein and calyculin A modulate the CFTR by different
mechanisms, and that genistein might inhibit calyculin A-insensitive
dephosphorylation of the CFTR.
Received 17 May 1996; accepted in final form 23 July 1996.
APS Manuscript Number C270-6.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 21 August 1996