The long-term goal of this proposal is to determine the molecular mechanisms involved in the regulation of expression of GTP-binding regulatory proteins (G-protein), the signal tranduction proteins necessary for neurotransmitters and hormone receptor action, in neural cells. This proposal uses a combination of biochemical, immunological, and molecular biological approaches to study the molecular mechanisms regulating G-protein expression. We have identified a novel form of Gs alpha mRNA in astrocytoma cells which is also found in rat and mouse brain, but not other tissues. This proposal will isolate cDNA and genomic clones to determine its origin and relationship to previously identified forms of Gs alpha. This proposal will also use expression vectors to determine the effects of overexpression of specific G-protein subunits on the synthesis, assembly, and function of the G-protein proteins in situ in intact cells. In addition, we have obtained preliminary evidence that treatment of rat PC12 pheochromocytoma cells with nerve growth factor increases the levels of G-protein subunits. This proposal will use antibody and cDNA probes to determine the mechanisms involved in the regulation of G-protein expression by nerve growth factor. The polypeptide Go alpha is expressed in particularly high levels in brain and neuronal cells but expressed at low levels or unexpressed in astrocytoma, glioma, and other non- neuronal neural cell types. This proposal will localize the regulatory elements responsible for the high level of expression of Go alpha in neuronal cells. This proposal should provide new insights into the regulation of the number and function of G- proteins in neural cells.
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