The long-term objective of our research program is to understand how the vertebrate gustatory system transduces signals and encodes information. Of particular importance are taste cell guanine nucleotide binding regulatory proteins (G proteins) and the seven transmembrane helix receptors that couple to them. G proteins regulate effector enzymes such as phosphodiesterase (PDE) and phospholipase C (PLC) to effect taste cell changes in intracellular second messengers. We have used molecular cloning to identify a number of the key components of taste transduction pathways, including Alpha-gustducin, a taste specific G protein Alpha -subunit closely related to the Alpha-transducins. The scope of this Competing Continuation encompasses the roles in taste transduction and coding of heterotrimeric gustducin?s component subunits and the downstream effector enzymes to which they couple. To elucidate the specific roles in taste transduction and coding of Alpha-gustducin, BetaGamma-gustducin, Ggamma13, taste-expressed PDEs and PLCBeta2 we will use a multidisciplinary approach applying biochemical, transgenic, behavioral, and electrophysiological techniques. The Experimental Goals of the proposal are the following. 1. To determine which PDEs present in taste tissue can be activated by alpha-gustducin. 2. To determine if these PDEs are co-expressed with alpha-gustducin in taste cells. 3. To determine if these PDEs can be activated by any other G protein alpha-subunits co-expressed with them in taste cells. 4. To generate alpha-gustducin mutants selectively deficient in the ability to activate the taste PDEs. 5. To generate transgenic mice expressing such an alpha-gustducin mutant. 6. To biochemically characterize taste transduction responses of the alpha-gustducin mutant mice. 7. To behaviorally characterize the alpha-gustducin mutant mice. 8. To electrophysiologically characterize the alpha-gustducin mutant mice. 9. To generate transgenic mice lacking signals mediated by gustducin?s beta, gamma-subunits. 10. To biochemically characterize taste transduction responses of the beta gamma-gustducin deficient mice. 11. To behaviorally characterize the beta, gamma-gustducin deficient mice. 12. To electrophysiologically characterize the beta, gamma-gustducin deficient mice. The results of these studies will provide significant new insights into the molecular mechanisms underlying taste transduction and coding. Gustatory and metabolic disorders such as malgeusia, dysgeusia and cachexia frequently occur in conjunction with several types of cancer. The knowledge gained from this proposal should further our understanding of the molecular bases of the taste disorders and may lead to effective intervention.
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