The mechanisms by which extracellular signals are transduced to intracellular messenger systems is a major focus of attention in a wide variety of biological systems. A number of different signaling mechanisms have been described, and a major form of hormonal and sensory communication among eukaryotic cells involves the stimulation of specific cell surface receptors which interact with heterotrimeric GTP-binding proteins, G proteins. A large number of G protein-linked receptors have been characterized, from organisms as diverse as yeast, Dictyostelium, Drosophila and humans, and include multi gene families for opsins, and the adrenergic, muscarinic acetylcholine, serotonin, and dopamine receptors. Not surprisingly, defects in (3-protein linked systems have now been associated with several congenital disease states, and certain cancers are associated with aberrant 6 protein-mediated signaling. The family of G protein-coupled, cAMP receptors (cARs) in Dictyostelium has been used as a model of signal transduction, and much is known about cAMP- mediated signaling during the early hours of Dictyostelium development. It is clear that the cAMP receptor, cAR I, is essential for transducing most of the early cAMP signals, and our studies lead us to suggest that cAR2 plays an equally important role in later development. The question is how does cAR2 provide that unique function? Is cAR2's low affinity for cAMP the sole basis for its function? Do cARI and cAR2 activate the same or different effector systems? Is cAR2 regulated by the same intracellular components that regulate cAR I? To address these questions, the details of cAR2-mediated signaling will be determined.We will establish which pathways are normally activated by cAR 2, and determine the relationship between its structure and function(s). We will also begin to identify the intracellular regulators of cAR2-mediated signaling. Both biochemical and genetic methods will be employed to identify functionally important regions of cAR2 and various methods, including insertional mutagenesis will be used to identify intracellular components coupled to cAR2-mediated signaling. Completion of the aims of this grant will provide details for one relatively simple system, cAMP signaling in late Dictyostelium development. Answers to questions addressed here should bear on broader issues of how multiple receptors, for specific signals, are used to regulate cell behavior and development.
