Crj. Kennedy et al., BRADYKININ-STIMULATED ARACHIDONIC-ACID RELEASE FROM MDCK CELLS IS NOTPROTEIN-KINASE-C DEPENDENT, American journal of physiology. Cell physiology, 42(5), 1997, pp. 1605-1612
Bradykinin (BK)-induced release of arachidonic acid (AA) from Madin-Da
rby canine kidney (MDCK) D1 cells was investigated. Phorbol 12-myrista
te 13-acetate (PMA) caused a synergistic increase in BK- and A-23187-i
nduced release of AA but alone had no effect on this release. Inhibiti
on of protein kinase C (PKC) with bisindolmaleimide I (BIS) abolished
the synergistic effects of PMA but did not affect AA release caused by
BK or A-23187 alone. Downregulation of PKC with 100 nM PMA resulted i
n a reduction of AA release induced by BK or A-23187 addition, which c
orresponded to a decrease in cytoplasmic phospholipase A(2) (cPLA(2))
activity as measured in cell extracts. Although Western blotting revea
led no differences in cPLA(2) expression as a result of PMA treatment,
phosphorylation of the enzyme, as assessed by phosphoserine content,
was significantly reduced in PKC-depleted cells. These results imply t
hat, with PKC downregulation, subsequent BK stimulation results in a C
a2+-dependent translocation of a less phosphorylated, less active form
of cPLA(2). Any stimulation of PKC by BK addition did not appear as a
significant event in onset reponses leading to AA release. On the oth
er hand, inhibition of the mitogen-activated protein kinase (MAPK) cas
cade with the MAPK kinase inhibitor, PD-98059, significantly decreased
BK-induced release of AA, a finding that, with our other results, poi
nts to the existence of a PKC-independent route for stimulation of MAP
K and the propagation of onset responses.