Inhibitors of granule fusion block cl- secretion in a mucin
-secreting cell line. evidence for recruitment of purinergically
stimulated cl- channels to the surface membrane.
Merlin, Didier, Xiaowei Guo, Kathleen Martin, Christian Laboisse,
Dennis Landis, George Dubyak1 and Ulrich Hopfer.
Departments of Physiology and Biophysics & Department of
Neurology, School of Medicine, Case Western Reserve University,
Cleveland, OH, 44106 and, Groupe de Recherche Fonctions
S[acute]ecr[acute]etoires Des Epitheliums Digestifs (C.J.F.
I.N.S.E.R.M 94-04)2, Universit[acute]e de Nantes, Facult[acute]e de
M[acute]edecine, F-44035 Nantes (France)
APStracts 3:0086C, 1996.
HT29-Cl.16E and HT29-Cl.19A are two different subcloned cell lines
derived from the human adenocarcinoma cell line HT29. They are
similar in their ability to grow and differentiate to polarized
epithelial cells, but differ in that Cl.16E is goblet cell-like with
many mucin granules, while Cl.19A lacks mucin granules. Extracellular
ATP stimulates Cl- secretion in both cell lines through luminal P2u
receptors and, in Cl.16E, also mucin secretion release. To evaluate
whether fusion of mucin granules is associated with an increase in
Cl- conductance of the plasma membrane, the effects of two fusion
inhibitors on luminal Cl- conductance were measured. Blockage of
actin depolymerization with phalloidin (1 [mu]M) inhibited
purinergically, but not cAMP stimulated luminaL Cl- efflux by 50% in
Cl.16E. The same treatment was without effect in Cl.19A. The fungal
metabolite wortmannin, which is an inhibitor of regulated exocytosis
in leukocytes, at 100 nM inhibited Cl- secretion by 70% in Cl.16E.
This inhibition was not a direct effect on purinergically stimulated
Cl- channels as wortmannin up to 1 [mu]M did not affect the secretory
response in Cl.19A. The wortmannin inhibition of Cl- secretion is
associated with an inhibition of granule fusion as judged by electron
microscopy. The differential inhibition of Cl- secretion in the
related HT29 clones that differ with respect to the presence of mucin
granules indicates that i) the granule fusion inhibitors, phalloidin
and wortmannin have no direct inhibitory effects on purinergically
and cAMP activated Cl- channels; ii) a major portion of
purinergically, but not cAMP activated Cl- channels is associated
with granule fusion in Cl.16E; and iii) the signaling pathways for
Cl- secretion and granule fusion are not completely identical.
Received 19 July 1995; accepted in final form 6 March 1996.
APS Manuscript Number C436-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 27 March 96