Activation of map kinases in airway smooth muscle.
Gerthoffer, William T., Ilia A. Yamboliev, Jennifer Pohl, Robert
Haynes, Steve Dang, and Jeannette McHugh.
Department of Pharmacology, University of Nevada School of
Medicine, Reno, NV 89557
APStracts 3:0186L, 1996.
To test the hypothesis that MAP kinases are activated by contractile
agonists in intact, nonproliferating airway smooth muscle, kinase
activities were compared in resting and stimulated canine tracheal
smooth muscle. Kinase activities in SDS extracts were assayed by a
gel renaturation method. MBP kinase activities corresponding to ERK1
and ERK2 immunoreactive proteins were activated two-fold above basal
level within 5 min by 10 M carbachol. MAP kinase activity assayed in
crude homogenates using a synthetic peptide substrate (APRTPGGRR)
also increased two-fold above basal in muscles stimulated with 10 M
carbachol. Two protein kinases separated by Mono-Q chromatography
were identified on Western blots as ERK1 and ERK2 MAP kinases.
Carbachol stimulation increased caldesmon phosphorylation in intact
muscle, and purified caldesmon was a substrate for activated murine
ERK2 MAP kinase. Activated ERK2 MAP kinase added to Triton-X 100
permeabilized fibers potentiated Ca2+-induced contraction. The
results show that ERK MAP kinases are activated following stimulation
of muscarinic receptors in airway smooth muscle which is consistent
with coupling of MAP kinases to phosphorylation of caldesmon in vivo.
Received 8 November 1996; accepted in final form 8 October 1996.
APS Manuscript Number L319-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 5 November 1996