Sphingosine-1-phosphate (S1P), a lipid metabolite, has been recently demonstrated to be an important signaling mediator for vital cellular and physiological processes, such as cell motility, invasion, proliferation, angiogenesis and apoptosis. S1P is produced from ceramide and sphingosine via phosphorylation by two isoenzymes, sphingosine kinase-1 (SphK1) and sphingosine kinase-2 (SphK2). The regulation of the levels of these metabolites, a so called “sphingolipid rheostat”, is complex and a number of enzymes have been demonstrated to be important. Upon production, S1P interacts with a family of G protein-coupled receptors (S1PR1–5) on the cell surface and/or intracellular targets, such as histone deacetylase (HDAC) and TRAF2, to play a plethora of roles in diverse pathophysiological conditions such as inflammation, immunity and cancer. Although SphK1 and SphK2 share a high degree of homology, they differ significantly in size, tissue distribution, and subcellular localization.