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Elsevier, Cellular Signalling, 1(28), p. 120-129, 2016

DOI: 10.1016/j.cellsig.2015.10.011

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Comprehensive analysis of chemokine-induced cAMP-inhibitory responses using a real-time luminescent biosensor

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This paper is available in a repository.

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Abstract

Chemokine receptors are members of the G-protein-coupled receptor (GPCR) family coupled to members of the Gi class, whose primary function is to inhibit the cellular adenylate cyclases. We used a cAMP-related and PKA-based luminescent biosensor (GloSensor(TM) F-22) to monitor the real-time downstream response of chemokine receptors, especially CX3CR1 and CXCR4, after activation with their cognate ligands CX3CL1 and CXCL12. We found that the amplitudes and kinetic profiles of the chemokine responses were conserved in various cell types and were independent of the nature and concentration of the molecules used for cAMP prestimulation, including either the adenylate cyclase activator forskolin or ligands mediating Gs-mediated responses like prostaglandin E2 or beta-adrenergic agonist. This tool allowed showing that the cAMP-inhibitory responses to multiple chemokines exhibited no cross-desensitization. Moreover the response kinetics appeared to be governed by the chemokine receptor internalization. Finally, we show that this conserved chemokine response can be accounted for by a simple model only combining Gi inhibitory action on adenylate cyclase and the associated phosphodiesterase activity. We conclude that the cAMP chemokine response is robustly conserved in various inflammatory conditions. Moreover, the cAMP-related luminescent biosensor appear as a valuable tool to analyze the details of Gi-mediated cAMP-inhibitory cellular responses, even in native conditions and could help to decipher their precise role in cell function.