The principal goal of this project is to develop a panel of immunologic reagents directed against inositol phospholipid (InsPL)-specific phospholipase C (PLC). This goal includes the characterization of their reactivity, specificity, and their effect in functional assays. A secondary objective is the utilization of these reagents in investigating the mechanism of T lymphocyte activation. Signal transduction by several receptor, including muscarinic receptors, certain growth factor receptors, and the antigen receptors of B and T lymphocytes, is mediated, at least in part, by the products of hydrolysis of InsPL by PLC. The regulation of this enzymatic activity is crucial to controlling a cell's response. To better characterize the role of this enzymes in immune cells and attempt in depth analysis of PLC regulation, including reconstitution studies, a new generation of highly specific reagents need to be produced. Based on the deduced amino acid sequences of the various PLC isozymes and a careful analysis of their homology, we propose the synthesis of a set of peptides which will be used as Ag for the generation of antisera and MoAb. In addition, the synthetic peptides, which are tailored to regions with a high degree of probability of being involved in regulatory functions, may also be used as probes to investigate PLC regulation.
