From the Department of Medicine, Division of Hypertension, Case Western
Reserve University School of Medicine and University Hospitals of Cleveland,
Cleveland, Ohio.
Correspondence to Dr Subash Harwalkar, Division of Hypertension, 10900 Euclid Ave, W-161, Case Western Reserve University, School of Medicine, Cleveland, OH 44106.
Abstract—Angiotensin II
in proximal tubule epithelium is known to stimulate the release of
arachidonic acid after stimulation of phospholipase
A2 (PLA2) independent of phospholipase
C–mediated signaling. Furthermore, an angiotensin II type
2 receptor subtype has been linked to this signaling cascade. We
investigated the regulation and differential stimulation of
PLA2s by comparing the PLA2 activities
associated with the membranes and cytosol of rabbit renal proximal
tubular epithelial cells after stimulation with angiotensin
II, epidermal growth factor, and bradykinin. Both fractions
demonstrated PLA2 activity that was dithiothreitol
insensitive, required micromolar concentrations of Ca2+ for
optimal activity, and was inhibited in a dose-dependent manner by an
antiserum to a cytosolic PLA2 with a molecular mass of 85
kD. However, membrane-associated PLA2 did not demonstrate
significant substrate specificity, whereas
1-steroyl-2-[14C]arachidonylphosphatidyl choline was the
preferred substrate for cPLA2. An antiserum generated
against mastoparan, a known PLA2 activator,
inhibited membrane- but not cytosol-associated PLA2
activity. Membrane fractions showed a broad pH range (7.5 to 8.5) for
optimal PLA2 activity, whereas cytosol was maximum at pH
9.5. Angiotensin II stimulated membrane-associated
PLA2 activity by 88%, whereas bradykinin and epidermal
growth factor inhibited activity by 54% and 41%, respectively. The
three agonists stimulated cPLA2. Moreover,
angiotensin II–induced activation of membrane-associated
PLA2 preceded the activation of cPLA2. These
results demonstrate differential localization and regulation of
proximal tubular epithelial PLA2 isozymes, which may
determine the pattern of subsequent arachidonic acid
metabolism by the cytochrome P450 system.
© 1998 American Heart Association, Inc.
Scientific Contributions
Role of Phospholipase A2 Isozymes in Agonist-Mediated Signaling in Proximal Tubular Epithelium
Key Words: phospholipase A2 • mastoparan • angiotensin II • mitogen-activated protein kinase • bradykinin • isozymes • endothelium
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