Sli2 (Ypk1), a homologue of mammalian protein kinase SGK, is a downstream kinase in the sphingolipid-mediated signaling pathway of yeast

ISP-1 is a new type of immunosuppressant, the structure of which is homolog ous to that of sphingosine, In a previous study, ISP-1 was found to inhibit mammalian serine palmitoyltransferase, the primary enzyme involved in sphi ngolipid biosynthesis, and to reduce the intracellular pool of sphingolipid s. ISP-1 induces the apoptosis of cytotoxic T cells, which is triggered by decreases in the intracellular levels of sphingolipids. In this study, the inhibition of yeast (Saccharomyces cerevisiae) proliferation by ISP-1 was o bserved, This ISP-1-induced growth inhibition was also triggered by decreas es in the intracellular levels of sphingolipids. In addition, DNA duplicati on without cytokinesis was detected in ISP-1-treated yeast cells on flow cy tometry analysis. We have cloned multicopy suppressor genes of yeast which overcome the lethal sphingolipid depletion induced by ISP-1. One of these g enes, SL12, is synonymous with YPK1, which encodes a serine/threonine kinas e, Kinase-dead mutants of YPK1 did not show any resistance to ISP-1, leadin g us to predict that the kinase activity of the Ypk1 protein should be esse ntial for this resistance to ISP-1, Ypk1 protein overexpression had no effe ct on sphingolipid biosynthesis by the yeast. Furthermore, both the phospho rylation and intracellular localization of the Ypk1 protein were regulated by the intracellular sphingolipid levels. These data suggest that the Ypk1 protein is a downstream kinase in the sphingolipid-mediated signaling pathw ay of yeast. The Ypk1 protein was reported to be a functional homologue of the mammalian protein kinase SGK, which is a downstream kinase of 3-phospho inositide-dependent kinase 1 (PDK1), PDK1 phosphotidylinositol (PI) is regu lated by PT-3,4,5-triphosphate and PI3,4-bisphosphate through the pleckstri n homology (PH) domain. Overexpressisn of mammalian SGK also overcomes the sphingolipid depletion in yeast. Taking both the inability to produce PI-3, 4,5-triphosphate and PI-3,4-bisphosphate and the lack of a PH domain in the yeast homologue of PDK1, the Pkh1 protein, into account, these findings fu rther suggest that yeast may use sphingolipids instead of inositol phosphol ipids as lipid mediators.