The goals of this proposal relate to deepening understanding of the consequences of activation of alpha1 adrenergic receptors, with emphasis on signal transduction mechanisms in vascular smooth muscle cells. Preliminary data indicate that the contribution of alpha1 receptors to vascular cell growth has been underestimated in comparison to peptide growth factors. The proposal serves to continue exploration of biologically interesting alpha1 receptor mechanisms that could also have clinical significance for atherosclerosis and vascular growth in hypertension. The proposal has two major aims: 1 Signal transduction mechanisms of al receptors in vascular smooth muscle and transfected NIH3T3 cells. al receptors activate a variety of signaling pathways including MAP kinases, PI 3-kinase, and p70S6 kinase. These pathways have importance for receptor-activated increases in protein and DNA synthesis. The primary purpose of this aim is to develop deeper insight into the mechanisms used by alpha1 receptors to activate these signaling pathways and to contrast them with the actions of angiotensin II and other growth factors such as platelet derived growth factor. 1A. Investigate the mechanism for the essential role of Ca2+ in alpha1 receptor- mediated activation of MAP kinase and p70S6 kinase and tyrosine protein phosphorylation, especially of phospholipase Cgamma. 1B. Determine the role of alpha1 receptors in the activation of PI-3 kinase isoforms and p70S6 kinase. 1C. alpha1 and angiotensin II receptors stimulate PI 3-kinase activity in vascular smooth muscle cells yet do not stimulate PKB which is generally activated down-stream of PI 3-kinase. What is the mechanism responsible for this inability to activate PKB? 2. Regulation of gene expression by alpha1 receptors toys Preliminary results suggest that alpha1 receptors increase expression of a range of genes, including nerve growth factor and various tyrosine kinases and transcription factors. We propose to characterize using microarray gene chip technology the pattern of gene expression induced by alpha1 receptors in vascular smooth muscle and by specific alpha1 receptor subtypes in transfected HEK-293 cells. We will then characterize in detail the effects of alpha1 receptors on expression of identified genes of particular biological interest, both at the mRNA and protein level, as well as investigating possible biological implications of the change in expression of these proteins.
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