Cumulative Mutations Affecting Sterol Biosynthesis in the Yeast Saccharomyces cerevisiae Result in Synthetic Lethality That Is Suppressed by Alterations in Sphingolipid Profiles

Martin Valachovic^*^,1^,2, Bart M. Bareither^*^,1, M. Shah Alam Bhuiyan^*^,1, James Eckstein, Robert Barbuch, Dina Balderes, Lisa Wilcox, Stephen L. Sturley, Robert C. Dickson and Martin Bard^*^,3

^* Department of Biology, Indiana University–Purdue University, Indianapolis, Indiana 46202, Department of Drug Disposition, Eli Lilly, Lilly Corporate Center, Indianapolis, Indiana 46285, Institute of Human Nutrition, Columbia University Medical Center, New York, New York 10032 and Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, Kentucky, 40536

^³ Corresponding author: Department of Biology, Indiana University–Purdue University, 723 W. Michigan St., Indianapolis, IN 46202.
E-mail: mbard{at}iupui.edu

UPC2 and ECM22 belong to a Zn(2)–Cys(6) family of fungaltranscription factors and have been implicated in the regulationof sterol synthesis in Saccharomyces cerevisiae and Candidaalbicans. Previous reports suggest that double deletion of thesegenes in S. cerevisiae is lethal depending on the genetic backgroundof the strain. In this investigation we demonstrate that lethalityof upc2 ${Delta}$ ecm22 ${Delta}$ in the S288c genetic background is attributableto a mutation in the HAP1 transcription factor. In additionwe demonstrate that strains containing upc2 ${Delta}$ ecm22 ${Delta}$ are also inviablewhen carrying deletions of ERG6 and ERG28 but not when carryingdeletions of ERG3, ERG4, or ERG5. It has previously been demonstratedthat UPC2 and ECM22 regulate S. cerevisiae ERG2 and ERG3 andthat the erg2 ${Delta}$ upc2 ${Delta}$ ecm22 ${Delta}$ triple mutant is also syntheticallylethal. We used transposon mutagenesis to isolate viable suppressorsof hap1 ${Delta}$ , erg2 ${Delta}$ , erg6 ${Delta}$ , and erg28 ${Delta}$ in the upc2 ${Delta}$ ecm22 ${Delta}$ genetic background.Mutations in two genes (YND1 and GDA1) encoding apyrases werefound to suppress the synthetic lethality of three of thesetriple mutants but not erg2 ${Delta}$ upc2 ${Delta}$ ecm22 ${Delta}$ . We show that deletionof YND1, like deletion of GDA1, alters the sphingolipid profiles,suggesting that changes in sphingolipids compensate for lethalityproduced by changes in sterol composition and abundance.