Role of extracellular signal-regulated kinase and protein kinase ca
in cytosolic pla2 activation by bradykinin in madin-darby canine
kidney-d1 cells.
Xing, Mingzhao, Ling Tao, and Paul A. Insel.
Departments of Pharmacology and Medicine 0636, University of
California at San Diego, La Jolla, CA 92093-0636
APStracts 3:0342C, 1996.
The actions of bradykinin (BK) in Madin-Darby Canine Kidney (MDCK) and
other cell types involve formation of arachidonic acid (AA) and AA
products by as-yet undefined mechanisms. We found that BK promoted AA
release and an increase in phospholipase A2 (PLA2) activity in
subsequently prepared MDCK-D1 cell lysates, both of which were Ca2+
-dependent and were inhibited by the 85-kDa cytosolic PLA2 (cPLA2)
inhibitor AACOCF3 . In addition, BK treatment of cells led to
increased PLA2 activity of cPLA2 immunoprecipitated from lysates.
Thus, BK receptors mediate AA release via cPLA2 in MDCK-D1 cells. The
BK-promoted increase of cPLA2 activity was reversed by treatment of
cell lysates with potato acid phosphatase, implying that
phosphorylation underlies the activation of cPLA2. However,
extracellular signal-regulated kinase (ERK) appeared not to be
responsible for this phosphorylation, because treatment of cells with
BK (in contrast with the results obtained with epinephrine and
phorbol ester) caused neither enzyme activation nor phosphorylation
(as judged by molecular weight shift) of this kinase. Although the
[alpha] isoform of protein kinase C (PKC) is responsible for AA
release promoted by phorbol ester treatment of MDCK-D1 cells (Godson
et al, 1993, J. Biol. Chem. 268, 11946-11950), neither treatment of
cells with the PKCa-selective inhibitor GF109203X nor transfection of
cells with PKCa antisense cDNA altered BK-mediated AA release. We
conclude that PKCa is unlikely to play an important role in the
regulation of cPLA2 by BK receptors in MDCK-D1 cells. The tyrosine
kinase inhibitor herbimycin A, on the other hand, inhibited both BK
-promoted AA release in intact cells and cPLA2 activation in cell
lysates, suggesting the involvement of tyrosine kinase in the
regulation of this lipase by BK receptors. Taken together, these data
suggest that BK receptors in MDCK-D1 cells regulate cPLA2 via
phosphorylation mediated by kinases other than ERK and PKCa.
Received 30 July 1996; accepted in final form 18 October 1996.
APS Manuscript Number C427-6.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 13 November 1996