S1P has previously been demonstrated to control several endothelial cell (EC) responses, including cell movement, through regulating different signaling proteins like PI3Ks, Akt and Rac. In the present study, we showed that in HUVEC and MLEC the PI3K isoform PI3Kβ is important for S1P-induces Akt phosphorylation, whereas both PI3Kβ and PI3Kγ are essential for S1P-stimulated Rac activation. Moreover, PI3Kγ is only responsible for S1P-induced EC chemotaxis, while PI3Kβ is required for both chemotaxis and chemokinesis.