Differential stimulation of proline-rich tyrosine kinase 2 and mitogen-activated protein kinase by sphingosine 1-phosphate

Sphingosine l-phosphate (SphP), a metabolite of cellular sphingolipids, has been shown to induce cell proliferation by activating the mitogen-activated protein kinase (MAPK) pathway. Proline-rich tyrosine kinase 2 (Pyk2) is a novel cytosolic tyrosine kinase which mediates activation of the MAPK or c-Jun N-terminal kinase (JNK) signaling pathways in response to a variety of stimuli that elevate intracellular calcium. In this report, we show that SphP stimulates both tyrosine phosphorylation of Pyk2 and MAPK activation in a transient and dose-dependent manner in rat aortic smooth muscle cells. Further studies indicate that Pyk2 phosphorylation, but not MAPK activation, is dependent on a pertussis toxin-sensitive G-protein-coupled receptor as well as partially on actin cytoskeleton. In addition, both intracellular calcium mobilization and protein kinase C (PKC) are required for optimal Pyk2 phosphorylation while either calcium increase or PKC activation is sufficient for MAPK activation in response to SphP. Finally, we show that a tyrosine kinase(s) other than Pyk2 is necessary for MAPK activation by SphP. Together, these results suggest that SphP stimulates tyrosine phosphorylation of Pyk2 through a G-protein coupled receptor, which is dissociated from its activation of the MAPK pathway in these cells.