Activation of map kinase kinase (mek) and ras by cholecystokinin in
rat pancreatic acini.
Duan, Rui-Dong, Chao-Feng Zheng, Kun-Liang Guan, and John A. Williams.
Departments of Physiology, Biological Chemistry and Internal
Medicine, University of Michigan, Ann Arbor, MI 48109
APStracts 2:0029G, 1995.
Cholecystokinin (CCK) has recently been shown to activate mitogen
-activated protein (MAP) kinase in rat pancreatic acini (Duan and
Williams, Am J Physiology 267:G401, 1994). To evalute the mechanism
of MAP kinase activation, we studied the effects of CCK on MAP kinase
kinase (MEK) in rat pancreatic acini. Two forms of MEK were
identified by immunoblotting using antibodies specific to MEK1 and
MEK2. MEK activity in acinar extracts and following
immunoprecipitation with anti MEK was detected using a recombinant
fusion protein GST-MAP kinase as a substrate. MEK activity rapidly
increased following stimulation of acini by CCK with significant
stimulation at 1 min, a maximal effect at 5 min, followed by a slow
decline to slightly above control levels after 30 min. The threshold
concentration of CCK was about 10 pM and the maximum effect was
induced by 1 nM CCK, which increased MEK activity by 120%. In
addition to CCK, bombesin and carbachol, but not secretin or VIP,
enhanced MEK activity. Phorbol ester mimicked the effect of CCK,
whereas ionomycin and thapsigargin failed to activate MEK. We further
studied the activation of Ras, an important component leading to
activation of MEK by growth factors. Ras in acini was
immunoprecipitated and identified by Western blotting. CCK and TPA
stimulated the incorporation of GTP into Ras, a requirement for its
activation and reaching maximum at 10 min of about 120% over control.
In conclusion, the activation of MAP kinase by CCK can be explained
by activation of MEK and may involve the activation of Ras by a PKC
-dependent mechanism.
Received 11 October 1994; accepted in final form 20 January 1995.
APS Manuscript Number G409-4.
Article publication pending Am. J. Physiol. (Gastrointest. Liver
Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 7 March 1995.