ACTIVATION OF A HIGH-AFFINITY G(I) PROTEIN-COUPLED PLASMA-MEMBRANE RECEPTOR BY SPHINGOSINE-1-PHOSPHATE

Sphingosine-1-phosphate (SPP) has attracted much attention as a possib le second messenger controlling cell proliferation and motility and as an intracellular Ca2+-releasing agent. Here, we present evidence that SPP activates a G protein-coupled receptor in the plasma membrane of various cells, leading to increase in cytoplasmic Ca2+ concentration ( [Ca2+](i)), inhibition of adenylyl cyclase, and opening of G protein-r egulated potassium channels, In human embryonic kidney (HEK) cells, SP P potently (EC(50), 2 nM) and rapidly increased [Ca2+](i), in a pertus sis toxin-sensitive manner. Pertussis toxin-sensitive increase in [Ca2 +](i) was also observed with sphingosylphosphorylcholine (EC(50), 460 nM), whereas other sphingolipids, including ceramide-1-phosphate, N-pa lmitoyl-sphingosine, psychosine, and D-erythro-sphingosine at micromol ar concentrations did not or only marginally increased [Ca2+](i). Furt hermore, SPP inhibited forskolin-stimulated cAMP accumulation in HEK c ells and increased binding of guanosine 5'-3-O(thio)triphosphate to HE K cell membranes. Rapid [Ca2+](i) responses were also observed in huma n transitional bladder carcinoma (J82) cells, monkey COS-1 cells, mous e NIH 3T3 cells. Chinese hamster ovary (CHO-K1) cells, and rat C6 glio ma cells, whereas human HL-60 leukemia cells and human erythroleukemia cells failed to respond to SPP. In guinea pig atrial myocytes, SPP ac tivated G(i) protein-regulated inwardly rectifying potassium channels, Activation of these channel occurred strictly when SPP was applied at the extracellular face of atrial myocyte plasma membrane as measured in cell-attached and inside-out patch clamp current recordings, We con clude that SPP, in addition to its proposed direct action on intracell ular Ca2+ stores, interacts with a high affinity G(1) protein-coupled receptor in the plasma membrane of apparently many different cell type s.