Accession Number:

ADA439308

Title:

Identification of Two Suppressors of CSG2 Calcium Sensitivity, SCS7 and SUR2, as Genes Encoding Hydroxylases of the Sphingolipid Biosynthetic Pathway of Saccharomyces cerevisiae

Descriptive Note:

Corporate Author:

UNIFORMED SERVICES UNIV OF THE HEALTH SCIENCES BETHESDA MD

Personal Author(s):

Report Date:

1997-01-01

Pagination or Media Count:

127.0

Abstract:

Sphingolipids are abundant constituents of the plasma membrane of many eukaryotic cell types from yeast to human and are known to function in numerous cellular metabolic processes. Sphingolipids have a wide range of structural diversity with respect to the composition of the head group and the hydroxylation state of the ceramide backbone. The biochemical significance of much of this structural variability is not well understood. The sphingolipids of the yeast Saccharomyces cerevisiae undergo the a-hydroxylation of the very long chain fatty acid VLCFA moiety of the cerarnide backbone in the same manner as animal cells. Yeast sphingolipids are additionally hydroxylated on the long chain base LCB moiety and at a second position on the VLCFA. A connection between sphingolipid hydroxylation and calcium homeostasis in yeast is suggested by the phenotypic behavior of the calcium sensitive csg2 mutant and its suppressors. This mutant cannot mannosylate the sphingolipid head groups and thus overaccumulates unmannosylated sphingolipids with hydroxylated cerarnide backbones. Two secondary mutations, sur2 and scs7, suppress csg2 Ca2 sensitivity and the corresponding double mutants overaccumulate unmannosylated sphingolipids with slightly higher thin layer chromatography mobilities than those of csg2 single mutants, consistent with the former species being cerainide unhydroxylated. Additionally, sur2 and scs7 single mutants accumulate species with mobilities predicted for unhydroxylated mannosylated sphingolipids. Sphingolipids and ceramides from sur2 and scs7 mutants in both wild-type and csg2 backgrounds were purified by thin layer chromatography and subjected to acid methanolysis. TLC analysis showed that the acid-hydrolyzed sur2 extracts contam the long chain base, dihydrosphingosine, while the scs7 extracts contain nonhydroxylated VLCFA. This indicates that Sur2p and Scs7p are required for the LCB and VLCFA hydroxylations respectively.

Subject Categories:

  • Genetic Engineering and Molecular Biology
  • Biology
  • Anatomy and Physiology

Distribution Statement:

APPROVED FOR PUBLIC RELEASE