Mutant analysis reveals complex regulation of sphingolipid long chain base phosphates and long chain bases during heat stress in yeast.

Sphingolipid long chain bases (LCBs) in Saccharomyces cerevisiae, dihydrosphingosine (DHS) and phytosphingosine (PHS) and their phosphates (DHS-P and PHS-P) are thought to play roles in heat stress. However, quantitative studies of LCBs and LCBPs have been limited by analytical methods. A new analytical procedure allowed us to measure changes in all known LCBPs and LCBs in wild-type and mutant cells during heat shock and to correlate the changes with heat stress resistance. All five molecular species of LCBPs increased rapidly but transiently when log and stationary phase cells were heat-stressed and when log-phase cells were induced for thermotolerance, suggesting that LCBPs play a role in heat stress. In support of this hypothesis, cells lacking the minor LCB kinase, Lcb5p, but not the major kinase, Lcb4p, were two-fold less resistant to killing when log-phase cells were induced for thermotolerance. Thus, LCBPs seem to play a minor role in heat-stress resistance. However, their role may be masked by LCBs, which are elevated in mutant strains, such as one lacking Lcb4p. This elevation demonstrates that one function of Lcb4p is to regulate LCB levels. Two new compounds, C(16) DHS and C(16) DHS-P, were identified, with the latter being degraded by the Dpl1p lyase. Our data provide a basis for determining how the basal and heat-induced levels of individual species of LCBs and LCBPs are governed by the Lcb4p and Lcb5p kinases, the Dpl1p lyase and the Lcb3p phosphatase.