Odorant mixtures which simultaneously activate both the adenylate cyclase and phospholipase C transduction pathways in individual lobster olfactory receptor neurons cause an integrated cellular response. Co-activation of interacting transduction pathways may increase the quality coding capacity of the receptor neuron population by increasing the diversity of receptor neuron response types. The objective of this proposal is test this broad hypothesis by fully characterizing these two transduction pathways and their interactions. Experiments using molecular cloning, biochemical, and electrophysiological techniques are proposed. The hypotheses to be tested are: 1) that all receptor neurons possess both transduction pathways, 2) that cDNA clones from G protein alpha subunits, effector enzymes and second messenger-gated ion channels participate in the series of molecular interactions necessary to mediate olfactory transduction and desensitization, and 3) that any of several biochemical mechanisms mediate interactions between the transduction pathways. Lobster olfactory cDNA sequences for all or part of two G protein alpha subunits and a phospholipase C have been cloned using homology cloning techniques, which will also be used to clone an olfactory adenylate cyclase and a cyclic nucleotide-gated potassium channel. An IP3-gated channel cDNA clone will be isolated by immunoscreening of an expression library. In situ hybridizations will determine the distribution of each clone (hypothesis 1). Antibodies raised to each sequence will be used to test the presence of each clone in olfactory dendrites (hypothesis 2). The interaction pathway from G-protein to enzyme to channel will be studied in heterologous expression studies in eukaryotic cells and by co-immunoprecipitation from olfactory cilia (hypothesis 2). The effects of calcium, calmodulin, and phosphorylation upon the cloned enzymes and ion channels will be determined in functional expression studies (hypotheses 2 and 3).
| Xu, F; Hollins, B; Gress, A M et al. (1997) Molecular cloning and characterization of a lobster G alphaS protein expressed in neurons of olfactory organ and brain. J Neurochem 69:1793-800 |
| McClintock, T S; Xu, F; Quintero, J et al. (1997) Molecular cloning of a lobster G alpha(q) protein expressed in neurons of olfactory organ and brain. J Neurochem 68:2248-54 |
