Cloning and characterization of GNS1: a Saccharomyces cerevisiae gene involved in synthesis of 1,3-beta-glucan in vitro

Abstract

The GNS1 gene product is required for the synthesis of 1,3-␤-glucan in vitro, since mutations in this gene result in exhibit an 80 to 90% reduction in 1,3-␤-glucan synthase specific activity. gns1 mutant strains display a pleiotropic phenotype including resistance to a pneumocandin B 0 analog (L-733,560), slow growth, and mating and sporulation defects. The gns1-1 mutation was genetically mapped to within 1.35 centimorgans from the MAT locus on chromosome III. The wild-type GNS1 gene was isolated by complementing the pneumocandin resistance phenotype of the gns1-1 mutation and by hybridization with a chromosome III-derived sequence being used as a probe. The nucleotide sequence of GNS1 was determined and compared with the homologous region of the chromosome. The genetic and nucleotide sequence analyses revealed that GNS1 and the open reading frame, YCR34 [S. Oliver, Q. van der Aart, M. Agostoni-Carbone, and the Chromosome III Sequencing Group, Nature (London) 357:38-46, 1992], represent identical loci in the genome. Cells deleted for GNS1 are viable but exhibit slow growth as well as the pleiotropic phenotype of the gns1 mutants. The putative protein product is predicted to be an integral membrane protein with five transmembrane helices displaying an exoplasmic orientation for the N terminus and a cytoplasmic orientation for the C terminus. This protein may be a subunit of 1,3-␤-glucan synthase.

ELO2 and ELO3, Homologues of theSaccharomyces cerevisiae ELO1 Gene, Function in Fatty Acid Elongation and Are Required for Sphingolipid Formation

“…Mutations or disruptions of ELO2 and ELO3 were previously reported as affecting ␤-glucan synthase activity (12), the plasma membrane, (H ϩ )-ATPase (14), bud localization defects (13), and resistance to sterol synthesis inhibitors (11,13). All of these functions are apparently produced by the activities of intrinsic membrane proteins, which could be affected by changes in the composition of their lipid environment or by the absence of an essential sphingolipid-membrane protein interaction.…”

“…ELO2 was initially cloned by complementation of mutants of GNS1 (12). Those mutants confer resistance to echinocandins and have defects in ␤-glucan synthase activities.…”

Yeast 1,3-β-Glucan Synthase Activity Is Inhibited by Phytosphingosine Localized to the Endoplasmic Reticulum

“…Another gene, GNS1, was originally isolated as a positive component required for GS activity in vitro (10). GS activity is severely reduced in the membrane fraction of a ⌬gns1 mutant (10). GNS1 interacts genetically with FKS1: a ⌬gns1 ⌬fks1 double mutant grows more slowly and exhibits more reduced GS activity in the membrane fraction than single mutants (10).…”

“…Since GTP-bound Rho1p is required not only for cell wall synthesis but also for intracellular actin organization (9), signal transduction leading to Rho1p plays a key role in cell morphogenesis. Another gene, GNS1, was originally isolated as a positive component required for GS activity in vitro (10). GS activity is severely reduced in the membrane fraction of a ⌬gns1 mutant (10).…”

Characterization of Enzymes Involved in Fatty Acid Elongation

“…ELO2 was initially cloned by complementation of GNS1 mutants, which conferred resistance to echinocandins and had defects in "-glucan synthase activities [38].…”