Src regulates sphingosine-1-phosphate mediated smooth muscle cell migration

Background: Sphingosine-1-phosphate (S-1-P) is a bioactive sphingolipid released from activated platelets at sites of arterial injury that stimulates migration of smooth muscle cells (SMC). The kinase src is a significant focal point in transmembrane signaling. This study examines the role of src during smooth muscle cell migration in response to S-1-P. Methods: Human coronary arterial SMCs were cultured in vitro. Boyden microchemotaxis assays of migration were performed in response to S-1-P in the presence and absence the src inhibitor (PP2, 10 μM) and a dominant negative src construct (DNsrc). siRNA to S-1-P receptors was used to down-regulate the S-1-P receptors. Western blotting was performed for src and MAPK phosphorylation. Results: Inhibition of src with PP2 but not PP3 partially blocked S-1-P-mediated cell migration. S-1-P induced time-dependent activation of src, which was inhibited by PP2 and adenoviral DNsrc. PP3 or an empty vector had no effect. Activation of src by S-1-P was inhibited by siRNA to S-1-PR1 and S-1-PR3 but not by S-1-PR2. When the VSMC were transfected with adenovirus containing βARK _CT, an inhibitor to Gβγ, src activation was significantly attenuated. Src inhibition with PP2 reduced p38 ^MAPK and JNK activation but did not alter ERK1/2 activation. Conclusion: S-1-P mediated VSMC migration is modulated by a G-protein-coupled src pathway partially through src-mediated p38 ^MAPK and JNK signaling and requires S-1-PR1 and S-1-PR3 receptors.