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Co-transfection with protein kinase D confers phorbol-ester-mediated inhibition on glucagon-stimulated cAMP accumulation in COS cells transfected to overexpress glucagon receptors

Tobias, E.S., Rozengurt, E., Connell, J., and Houslay, M.D. (1997) Co-transfection with protein kinase D confers phorbol-ester-mediated inhibition on glucagon-stimulated cAMP accumulation in COS cells transfected to overexpress glucagon receptors. Biochemical Journal, 326 (2). pp. 545-551. ISSN 0264-6021

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Abstract

Glucagon elicited a profound increase in the intracellular cAMP concentration of COS-7 cells which had been transiently transfected with a cDNA encoding the rat glucagon receptor and under conditions where cAMP phosphodiesterase activity was fully inhibited. This was achieved in a dose-dependent fashion with an EC50 of 1.8+/-0.4 nM glucagon. In contrast with previous observations made using hepatocytes [Heyworth, Whetton, Kinsella and Houslay (1984) FEBS Lett. 170, 38-42], treatment of transfected COS-7 cells with PMA did not inhibit the ability of glucagon to increase intracellular cAMP levels. PMA-mediated inhibition was not conferred by treatment with okadaic acid, nor by co-transfecting cells with cDNAs encoding various protein kinase C isoforms (PKC-alpha, PKC-betaII and PKC-epsilon) or with the PMA-activated G-protein-receptor kinases GRK2 and GRK3. In contrast, PMA induced the marked inhibition of glucagon-stimulated cAMP production in COS-7 cells that had been co-transfected with a cDNA encoding protein kinase D (PKD). Such inhibition was not due to an action on the catalytic unit of adenylate cyclase, as forskolin-stimulated cAMP production was unchanged by PMA treatment of COS cells that had been co-transfected with both the glucagon receptor and PKD. PKD transcripts were detected in RNA isolated from hepatocytes but not from COS-7 cells. Transcripts for GRK2 were present in hepatocytes but not in COS cells, whereas transcripts for GRK3 were not found in either cell type. It is suggested that PKD may play a role in the regulation of glucagon-stimulated adenylate cyclase.

Item Type:Article
Status:Published
Refereed:Yes
Glasgow Author(s):Houslay, Prof Miles and Tobias, Prof Edward and Connell, Prof John
Authors: Tobias, E.S., Rozengurt, E., Connell, J., and Houslay, M.D.
College/School:College of Medical Veterinary and Life Sciences > Institute of Neuroscience and Psychology > Cognitive Neuroimaging and Neuroengineering Technologies
College of Medical Veterinary and Life Sciences > School of Medicine
College of Medical Veterinary and Life Sciences > School of Medicine > Clinical Specialities
Journal Name:Biochemical Journal
Publisher:Portland Press Ltd.
ISSN:0264-6021
ISSN (Online):1470-8728
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