GAS3, a developmentally regulated gene, encodes a highly mannosylated and inactive protein of the Gas family of Saccharomyces cerevisiae.
The multigene GAS family of Saccharomyces cerevisiae is constituted by five genes encoding GPI-anchored proteins required for cell wall or spore wall assembly. GAS1 and GAS5 are expressed in vegetative growth and repressed during sporulation, whereas GAS2 and GAS4 exhibit the opposite expression pattern. This study focuses on GAS3, a still poorly characterized member of the family. To date, attempts to reveal the glucan elongase activity typical of Gas proteins have been unsuccessful, suggesting that Gas3p is the only inactive member of the family. Here, we compared the mRNA levels of GAS1, GAS3 and GAS5 and demonstrate that GAS3 is the weakest-expressed paralogue in vegetative growth. Moreover, GAS3 mRNA increased during sporulation, showing a bimodal profile typical of the early-middle meiotic genes. GAS3 product was identified as a low-abundance, polydisperse mannoprotein. Loss of Gas3p did not affect growth and sporulation. The overexpression of GAS3, driven by the GAS1 promoter, slightly reduced growth rate in a wild-type strain and led to hyperaccumulation of Gas3p in the membranes and in the cell wall. To determine whether GAS3 could replace GAS1 function in vivo, GAS3 was also overexpressed in a gas1Delta mutant. Increased amounts of Gas3p were not only unable to complement the defects of the gas1Delta cells but exacerbated them. A mutated Gas3p-E283Q, where one of the catalytic glutamate residues essential for GH72 enzyme activity was replaced by glutamine, was also noxious to gas1Delta cells, indicating that the increased expression of Gas3p, rather than a potential activity, is deleterious for gas1Delta cells.
Rolli
Eleonora
E
Università degli Studi di Milano, Dipartimento di Scienze Biomolecolari e Biotecnologie, Via Celoria 26, 20133 Milano, Italy.
Ragni
Enrico
E
Rodriguez-Peña
José Manuel
JM
Arroyo
Javier
J
Popolo
Laura
L
D019943:Amino Acid Substitution; D017353:Gene Deletion; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D005964:Glucosyltransferases / Q000096:biosynthesis* / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics* / Q000254:growth & development*; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis* / Q000235:genetics*; D013172:Spores, Fungal / Q000737:chemistry / Q000235:genetics / Q000254:growth & development
C066709:GAS1 protein, S cerevisiae; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; C553583:GAS3 protein, S cerevisiae; C553582:GAS5 protein, S cerevisiae; D005964:Glucosyltransferases