Sphingosine-1-phosphate receptor subtype-specific positive and negative regulation of Rac and haematogenous metastasis of melanoma cells

We have recently reported that S1P (sphingosine-1-phosphate) differentially regulates cellular Rac activity and cell migration in either a positive or a negative direction via distinct G-protein-coupled receptor subtypes, i.e. S1P(1)/Edg1 (endothelial differentiation gene) and S1P(1)/Edg5 respectively, when each of the S1P receptor subtypes is expressed in CHO (Chinese-hamster ovary) cells. In B16F10 mouse melanoma cells, in which S1P(2), but not the other S1P-receptor subtypes, is endogenously expressed, S1P inhibited cell migration with concomitant inhibition of Rac and stimulation of RhoA in dose-dependent manners. Overexpression of S1P, in the melanoma cells resulted in potentiation of S1P inhibition of both Rac and cell migration. In contrast, overexpression of S1P(1) led to stimulation of cell migration, particularly at the lower S1P concentrations. Treatment of B16F10 cells with S1P inhibited lung metastasis 3 weeks after injection into mouse tail veins. Intriguingly, overexpression of S1P(2) greatly potentiated the inhibition of metastasis by S1P, whereas that of S1P(1) resulted in aggravation of metastasis. Suppression of cellular Rac activity by adenovirus-transduced expression of N(17)Rac, but not N(19)RhoA, strongly inhibited cell migration in vitro and lung metastasis in vivo. These results provide the first evidence that G-protein-coupled receptors could participate in the regulation of metastasis, in which ligand-dependent, subtype-specific regulation of the cellular Rac activity is probably critically involved as a mechanism.