Collections > Electronic Theses and Dissertations > Cloning and Characterization of a Novel Phospholipase C Enzyme, Human Phospholipase C Eta2
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G-protein-coupled receptors are cell surface macromolecules that regulate numerous cell physiologies through activation of the heterotrimeric G proteins. One very important signaling pathway downstream of heterotrimeric G protein is the hormone-activated phospholipase C pathway. Proteins of the phosphoinositide-specific phospholipase C (PLC) family play important roles in cell signaling. In response to numerous extracellular stimuli, such as neurotransmitters, hormones and growth factors, PLC catalyzes the hydrolysis of phosphatidylinositol (4,5)bisphosphate (PtdIns(4,5)P2) to generate two well-established second messengers, inositol(1,4,5)trisphosphate and diacylglycerol, and thereby increasing intracellular Ca2+ concentration and activating protein kinase C. These cellular signaling events eventually lead to a number of cellular responses such as neurotransmission, cardiac- and smooth muscle-contraction and blood pressure regulation. In addition to the hormone receptor activation of PLC, receptor tyrosine kinases, non-receptor tyrosine kinases, and members of the Ras superfamily GTPases also utilize PLC to regulate important cell physiologies. Moreover, PtdIns(4,5)P2 has been increasingly realized to be an important molecule that interacts with a broad range of signaling proteins. By altering ratio of PtdIns(4,5)P2, PLC potentially affects membrane association and activity of many signaling proteins. Therefore, PLC is a key enzyme in normal cell physiology. The work described herein reports the indentification, molecular cloning and characterization of a novel PLC isozyme, PLC-η2. First, I cloned PLC-η2 from human retina cDNA, and I examined possible existence of splice variants of the gene. Next I studied potential regulation of PLC-η2 by heterotrimeric G proteins and several members of the Ras family. Finally, I purified enzymes to homogeneity and demonstrated enzyme activity and direct activation in phospholipid vesicls. The work presented here expands further complexity and interplay between different PLC isozymes and possibly their regulatory networks.