The physiological catecholamines, epinephrine and norepinephrine, and many clinical useful pharmacological agonists and antagonists act on beta-adrenergic receptors. Although much new knowledge has recently been provided regarding beta-adrenergic receptor structure, much remains unknown regarding regulation of receptor expression and function in target cells. In this project we propose to continue studies that emphasize cellular and molecular mechanisms that regulate signal transduction by beta-adrenergic receptors in intact cells. My colleagues and I will continue to study wild-type S49 lymphoma cells and S49 variants in the pathway for beta receptor signalling via the guanine nucleotide regulatory protein, G(s) and in turn, adenylyl cyclase and cAMP-dependent protein kinase. We propose three specific aims designed to characterize the life cycles of beta-adrenergic receptors, Galpha(s), and the catalyst (C) of adenylyl cyclase. Antibodies to these components will be used to characterize biosynthesis and turnover and to assess the role of phosphorylation of beta-receptors and C. Other studies are designed to use molecular biological techniques (such as antisense technology and transfection with cDNAs) to help define the role of the beta-adrenergic receptor kinase (beta-ARK) in S49 cells and to evaluate stoichiometry between alpha(s) and C. Additional experiments will use biochemical and micrologic techniques to assess the subcellular localization of Galpha(s). Taken together, experimental approaches should provide new information regarding the key components involved in beta-adrenergic receptor signal transduction. As such, the results should be relevant to a variety of clinical settings, in particular cardiovascular disorders, in which betaadrenergic receptor function may be altered. In addition, these studies should help advance understanding of the molecular basis of pharmacological action at beta-adrenergic receptors and at other receptors that activate G(s) and adenylyl cyclase.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM040781-09S1
Application #
2806872
Study Section
Pharmacology A Study Section (PHRA)
Project Start
1988-08-01
Project End
1998-11-30
Budget Start
1996-12-01
Budget End
1998-11-30
Support Year
9
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of California San Diego
Department
Pharmacology
Type
Schools of Medicine
DUNS #
077758407
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Buscher, R; Herrmann, V; Insel, P A (1999) Human adrenoceptor polymorphisms: evolving recognition of clinical importance. Trends Pharmacol Sci 20:94-9
Buscher, R; Herrmann, V; Insel, P A (1998) PCR-based methods for identifying genetic variations in human alpha1B- and beta2-adrenergic receptors. Mol Genet Metab 64:266-70
Post, S R; Rump, L C; Zambon, A et al. (1998) ATP activates cAMP production via multiple purinergic receptors in MDCK-D1 epithelial cells. Blockade of an autocrine/paracrine pathway to define receptor preference of an agonist. J Biol Chem 273:23093-7
Post, S R; Aguila-Buhain, O; Insel, P A (1996) A key role for protein kinase A in homologous desensitization of the beta 2-adrenergic receptor pathway in S49 lymphoma cells. J Biol Chem 271:895-900
Denker, S P; McCaffery, J M; Palade, G E et al. (1996) Differential distribution of alpha subunits and beta gamma subunits of heterotrimeric G proteins on Golgi membranes of the exocrine pancreas. J Cell Biol 133:1027-40
Mochizuki, N; Cho, G; Wen, B et al. (1996) Identification and cDNA cloning of a novel human mosaic protein, LGN, based on interaction with G alpha i2. Gene 181:39-43
Post, S R; Jacobson, J P; Insel, P A (1996) P2 purinergic receptor agonists enhance cAMP production in Madin-Darby canine kidney epithelial cells via an autocrine/paracrine mechanism. J Biol Chem 271:2029-32
Insel, P A (1996) Seminars in medicine of the Beth Israel Hospital, Boston. Adrenergic receptors--evolving concepts and clinical implications. N Engl J Med 334:580-5
Hughes, R J; Anderson, K L; Kiel, D et al. (1996) Cloning of GRK2 cDNA from S49 murine lymphoma cells. Am J Physiol 270:C885-91
Post, S R; Hilal-Dandan, R; Urasawa, K et al. (1995) Quantification of signalling components and amplification in the beta-adrenergic-receptor-adenylate cyclase pathway in isolated adult rat ventricular myocytes. Biochem J 311 ( Pt 1):75-80

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