Phospholipase A2 (PLA/2) activity results in the synthesis of a number of eicosanoids, potent lipid mediators 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 kinase C (PKC) and annexins (ANX). ANX I is phosphorylated by growth factor receptors, and is a PLA/2 inhibitor. While there has been extensive study of the action of PLA/2 and its modifiers in cell culture, very little is known about their roles in the more complex context of a developing embryo. To investigate PLA/2 function in vivo, I chose a well characterize vertebrate system, the zebrafish Danio rerio, because its embryos are abundant and optically clear. This facilitates both the visualization of gene expression through whole mount in situ hybridization and the development of fluorescently-quenched lipid probes that can directly report PLA/2 activity than non-mitotic cells. I will examine PLA/2's role in cell signaling during development in both wild-type and mutant embryos identified by a biochemical screen for altered PLA/2 activity performed during the first two years of funding. I will examine the effect of over- expression of zebrafish annexins on embryonic PLA/2 activity and the phenotype consequences. I will examine the effect of over expression of zebrafish annexins on embryonic PLA/2 activity and the phenotypic consequences. My long-term objective is to understand the role of PLA/2 and its modifiers during development and thereby provide insight into the pathology associated with their aberrant activation.
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