Cftr chloride channel activation by genistein: the role of serine/
threonine protein phosphatases.
Reenstra, William W., Karin Yurko-Mauro, Anh Dam, Sasikala Raman, and
Suzanne Shorten.
Department of Clinical Science, Alfred I. duPont Institute, PO Box
269, Wilmington, DE 19899
APStracts 3:0073C, 1996.
We have previously shown (AJP 268:C886) that genistein, a tyrosine
kinase inhibitor, activates the CFTR chloride channel in NIH-3T3
cells that have been stably transfected with an expression vector for
the CFTR (NIH-CFTR cells). In this study, we present evidence
suggesting that both genistein and the serine/threonine protein
phosphatase (PPase) inhibitor calyculin A activate the CFTR by
inhibiting PPase activity. As measured by 125I-efflux, genistein and
calyculin A stimulate the CFTR to 50% of the maximal activity with
forskolin. Neither agonist increases CFTR activity at saturating
forskolin concentrations; but genistein and calyculin A have an
additive effect on CFTR activity. Forskolin, but neither genistein
nor calyculin A, stimulates protein kinase A (PKA) activity. The PKA
inhibitor, H-89, inhibits CFTR activation and in vivo phosphorylation
by all three agonists. Proteolytic digestion of in vivo
phosphorylated CFTR suggests that the CFTR is phosphorylated on the
same sites during stimulation with genistein and forskolin, but on
different sites during stimulation with calyculin A. The data suggest
that genistein and calyculin A inhibit different PPase activities,
allowing CFTR phosphorylation and partial stimulation, by a basal PKA
activity.
Received 21 December 1995; accepted in final form 26 February
1996.
APS Manuscript Number C756-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 20 March 96