Nrg-1 belongs to the endothelial differentiation gene family of G protein-coupled sphingosime-1-phosphate receptors

The previously cloned rat nerve growth factor-regulated G protein-coupled receptor NRG-1 (Glickman, M., Malek, R. L., Kwitek-Black, A. E., Jacob, H. J., and Lee N. H. (1999) Mot Cell. Neurosci. 14, 141-52), also known as EDG-8, binds sphingosine-l-phosphate (S1P) with high affinity and specificity, In this paper we examined the signal transduction pathways regulated by the binding of SIP to EDG-8. In Chinese hamster ovary cells heterologously expressing EDG-8, SIP inhibited forskolin-induced cAMP accumulation and activated c-Jun NH2-terminal kinase. Surprisingly, S1P inhibited serum-induced activation of extracellular regulated protein kinase 1 and 2 (ERK1/2). Treatment with pertussis toxin, which ADP-ribosylates and inactivates G(i), blacked S1P-mediated inhibition of cAMP accumulation, but had no effect on c-Jun NH2-terminal kinase activation or inhibition of ERK1/2. The inhibitory effect of SIP on ERK1/2 activity was abolished by treatment with orthovanadate, suggesting the involvement of a tyrosine phosphatase. A subunit selective [S-35] guanosine 5'-3-O-(thio)triphosphate binding assay demonstrates that; EDG-8 activated G(i/o) and G(12) but not G(s) and G(q/11) in response to S1P. In agreement, EDG-8 did not stimulate phosphoinositide turnover or cAMP accumulation. The ability of S1P to induce mitogenesis in cells expressing the EDG-1 subfamily of G protein-coupled receptors is well characterized. In contrast, SIP inhibited proliferation in Chinese hamster ovary cells expressing EDG-8 but not empty vector. The antiproliferative effect, like SIP-mediated ERK1/2 inhibition, was orthovanadate-sensitive and pertussis toxin-insensitive. Our results indicate that EDG-8, a member of the EDG-1 subfamily, couples to unique signaling pathways.