Genetic, biochemical, and morphological evidence for the involvement of N-glycosylation in biosynthesis of the cell wall beta1,6-glucan of Saccharomyces cerevisiae.
Recent evidence indicates that Stt3p plays a central role in the recognition and/or catalytic step in N-glycosylation (asparagine-linked glycosylation) in the lumen of the endoplasmic reticulum. It is known that stt3 mutants exhibit certain phenotypic features that are suggestive of a cell wall defect. To understand the basis of these phenotypes, we devised a genetic screen to isolate strains bearing mutations that lead to synthetic lethality in combination with the stt3-1 mutation. Using this screen, we were surprised to identify two KRE genes (KRE5 and KRE9) that are involved in the biosynthesis of the cell wall beta1,6-glucan. This finding led us to propose that the N-glycosylation process is essential in the biosynthesis of cell wall beta1,6-glucan. This proposal was supported by the observation that several stt3 mutants exhibited a 60-70% reduction in the content of cell wall beta1,6-glucan as compared with WT cells. Transmission electron microscopy revealed that the stt3 mutant strains exhibit a diffused cell wall with loss of the outer mannoprotein layer as compared with the WT cells. Thus, we provide genetic, morphological, and biochemical evidence for the critical involvement of N-glycosylation in some step in assembly of the cell wall beta1,6-glucan in Saccharomyces cerevisiae.
Chavan
Manasi
M
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY 11794, USA.
Suzuki
Tadashi
T
Rekowicz
Magdalena
M
Lennarz
William
W
D001483:Base Sequence; D002473:Cell Wall / Q000378:metabolism*; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D005804:Genes, Lethal; D005936:Glucans / Q000096:biosynthesis* / Q000235:genetics; D006023:Glycoproteins / Q000235:genetics; D006031:Glycosylation; D009154:Mutation; D010641:Phenotype; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics* / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D047071:beta-Glucans
D017931:DNA Primers; D005936:Glucans; D006023:Glycoproteins; C064198:KRE5 protein, S cerevisiae; C083328:KRE9 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D047071:beta-Glucans; C064197:beta-1,6-glucan