Evidence for and Subcellular Localization of a Ca-Stimulated Phospholipase D from Maize Roots

Autolytic lipid changes in corn (Zea mays L.) root crude homogenates and isolated membranes were examined by the use of high performance thin-layer chromatography. In the absence of added CaCl(2), losses in phosphatidylcholine and other phospholipids corresponds to increase in fatty acids without the accumulation of either phosphatidic acid or lyso-phosphatidylcholine. However, in the presence of 1 millimolar CaCl(2), phosphatidylcholine concentrations declined more rapidly with an immediate increase in phoshatidic acid, and slower rate of fatty acid accumulation. Autolytic phospholipid degradation yielded primarily free fatty acids in the absence of Ca and phosphatidic acid in the presence of 1 millimolar CaCl(2), suggesting the presence of an acyl hydrolase and phospholipase D activities. Differential centrifugation studies indicate that 50 to 80% of the crude homogenate's phospholipase D activity is membrane-bound. Density centrifugation experiments suggest that the membrane-bound phospholipase D activity is localized primarily on mitochondrial membranes.