Sphingosine-1-phosphate (S1P) lyase catalyzes the cleavage of the bioactive lipid molecule S1P to phosphoethanolamine and hexadecenal, both of which are utilized as glycerophospholipid precursors. Until now, only one gene, SPL, has been identified as encoding a S1P lyase. In the present study, SPL-null F9 cells were able to convert radiolabeled dihydrosphingosine to glycerophospholipids, albeit at much lower efficiency than parent cells. Lysates prepared front the SPL-null cells exhibited weak but significant dihydrosphingosine-1-phosphate lyase activity in vitro. These results provide evidence of the existence of an alternative S1P lyase. (c) 2005 Elsevier Inc. All rights reserved.