Characterization of phospholipase A2 activity enriched in the nerve growth cone.

Nerve growth cones isolated from fetal rat brain exhibit in their cytosol a robust level of phospholipase A 2 activity hydrolyzing phosphatidylinositol (PI) and phosphatidylethanolamine (PE) but not phosphatidylcholine (PC). Western blot analysis with an antibody to the well-characterized cytosolic phospholipase A 2 (mol wt 85,000) reveals only trace amounts of this PC- and PE-selective enzyme in growth cones. By gel filtration on Superose 12, growth cone phospholipase A 2 activity elutes essentially as two peaks of high molecular mass, at ∼65 kDa and at well over 100 kDa. Anion exchange chromatography completely separates a PI-selective from a PE-selective activity, indicating the presence of two different, apparently monoselective phospholipase A 2 species. The PI-selective enzyme, the predominant phospholipase A 2 activity in whole growth cones, is enriched greatly in these structures relative to their parent fractions from fetal brain. This phospholipase A 2 is resistant to reducing agents and is found in the cytosol as well as membrane-associated in the presence of Ca 2+ . However, its catalytic activity is Ca 2+ -independent regardless of whether the enzyme is associated with pure substrate or mixed-lipid growth cone vesicles. The PE-selective phospholipase A 2 in growth cones was studied in less detail but shares with the PI-selective enzyme several properties, including intracellular localization, the existence of cytosolic and membrane-associated forms, and Ca 2+ independence. Our data indicate growth cones contain two high-molecular-weight forms of phospholipase A 2 that share many properties with known, Ca 2+ -independent cytosolic phospholipase A 2 species but that appear to be monoselective for PI and PE, respectively. In particular, the PI-selective enzyme may represent a new member of the growing family of cytoplasmic phospholipases A 2 . The enrichment of the PI-selective phospholipase A 2 in growth cones suggests it plays a major role in the regulation of growth cone function.