Phospholipase A2 (PLA2) activation results in the synthesis of a number or potent lipid mediators (eicosanoids) that play a role in inflammation, mitogenesis and metastasis of cancer cells, and are likely to be important during development. The substrate for the synthesis of all eicosanoids is arachidonic acid (AA), a molecule generated by PLA2 activity. Modifiers of PLA2 activity include cytokines, growth factors, protein kinase C (PKC) and lipocortin 1 (LC1). LC1 is both phosphorylated by growth factor receptors, and is a PLA2 inhibitor. While there has been extensive study of the action of PLA2 and its modifiers in cell culture, very little is known about their role in normal vertebrate development. To investigate PLA2 function in vivo, I chose a well characterized vertebrate system, the developing zebrafish, Danio rerio because its embryos are abundant and optically clear. This facilitates both the visualization of gene expression through whole embryo in situ hybridization and the development of lipid probes that can directly report PLA2 activity in living embryos. Through direct observation, I plan to test the hypothesis that cells undergoing proliferation exhibit higher PLA2 activity than non-mitotic cells. I will also examine PLA2's role in cell signaling during development. I will concentrate on an easily visualized row of cells in the ventral midline of the embryonic central nervous system, the floor plate, where LC1 is selectively expressed in both mouse and rat embryos. By cloning zebrafish LC1 and examining its expression during floor plate formation, I can determine whether LC1 and PLA2 are candidate mediators of the signaling events that lead to floor plate development. In addition, there are well characterized zebrafish mutants that lack floor plate and/or notochord (a structure that is thought to release signals necessary for floor plate development) which will be useful to test the actions of LC1. My long- term objective is to understand the role of phospholipases during development and thereby provide insight into the pathology associated with their aberrant activation.
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| Hendrickson, H S; Hendrickson, E K; Johnson, I D et al. (1999) Intramolecularly quenched BODIPY-labeled phospholipid analogs in phospholipase A(2) and platelet-activating factor acetylhydrolase assays and in vivo fluorescence imaging. Anal Biochem 276:27-35 |
