Lysophospholipids are lipids generated by the action of phospholipase. They cause lysis of cellular membranes, play a role in signal transduction, serve as precursors of platelet activating factor (PAF), and are implicated in ischemia and cardiovascular disease. Lysophospholipases (Lyso PLA) hydrolyze to produce free fatty acids and simple phosphodiesters, thereby controlling the levels of lysophospholipid in the cell. While phospholipase, such as phospholipase A2 and phospholipase C, have been extensively studied, little is known about the lysoPLA's. Several years ago, we discovered that lysoPLA's were present in surprisingly high levels in some cells and tissues. This led us to purify three different lysoPLA's and to study the properties of their lysophospholipid substrates. We now propose to carry out a comprehensive study of lyso PLA's and lysophospholipids. We will focus on four different enzymes which represent two of the three different classes of lysoPLA. The first class is represented by the 28 kDa lysoPLA I and 27 kDa lysoPLA II from the murine macrophage-like cell line, P388D1. These enzymes are specific lysoPLAs and do not have any other lipase activities. The second type of lysoPLA catalyzes both phospholipase A2 and lysoPLA activity. The 85 kDa human cytosolic phospholipase A2 belongs to this class. The fourth enzyme we will study is the 17 kDa human eosinophil lysoPLA whose activities have not been defined. This is the smallest lysoPLA identified to date, has a remarkably low specific activity, and has a propensity to crystalize in vivo to form Charcot-Leyden crystals. The proposed studies on the specificity, kinetics, chemical modification, inhibition, and mutagenesis of these enzymes should lead to an enhanced understanding of the cellular function, regulation, mechanism of action, and structure/function correlates of lysoPLA's.
