Role of Phospholipase A 2 Isozymes in Agonist-Mediated Signaling in Proximal Tubular Epithelium

• Published in: Hypertension (Dallas, Tex. 1979) Tex. 1979), 1998-03, Vol.31 (3), p.809-814
• Main Authors: Harwalkar, Subash, Chang, Chung-Ho, Dulin, Nickolai O., Douglas, Janice G.
• Format: Article
• Language: English
• ISSN: 0194-911X; 1524-4563
• DOI: 10.1161/01.HYP.31.3.809

Abstract —Angiotensin II in proximal tubule epithelium is known to stimulate the release of arachidonic acid after stimulation of phospholipase A 2 (PLA 2 ) 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 PLA 2 s by comparing the PLA 2 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 PLA 2 activity that was dithiothreitol insensitive, required micromolar concentrations of Ca 2+ for optimal activity, and was inhibited in a dose-dependent manner by an antiserum to a cytosolic PLA 2 with a molecular mass of 85 kD. However, membrane-associated PLA 2 did not demonstrate significant substrate specificity, whereas 1-steroyl-2-[ 14 C]arachidonylphosphatidyl choline was the preferred substrate for cPLA 2 . An antiserum generated against mastoparan, a known PLA 2 activator, inhibited membrane- but not cytosol-associated PLA 2 activity. Membrane fractions showed a broad pH range (7.5 to 8.5) for optimal PLA 2 activity, whereas cytosol was maximum at pH 9.5. Angiotensin II stimulated membrane-associated PLA 2 activity by 88%, whereas bradykinin and epidermal growth factor inhibited activity by 54% and 41%, respectively. The three agonists stimulated cPLA 2 . Moreover, angiotensin II–induced activation of membrane-associated PLA 2 preceded the activation of cPLA 2 . These results demonstrate differential localization and regulation of proximal tubular epithelial PLA 2 isozymes, which may determine the pattern of subsequent arachidonic acid metabolism by the cytochrome P450 system.