Homologs of
the GPI-anchored beta(1,3)-Glucan transferase Gas1p of Saccharomyces
cerevisiae
have specific roles during sporulation.
Enrico Ragni (1),
Cristina Carotti (1), Marina Vai (2), Laura Popolo (1)
(1) Fisiologia e Biochimica Gen, Università di Milano, Via Celoria 26, Milano,
20133, Italy (Enrico.Ragni@unimi.it); (2) Università di Milano - Bicocca, P.le
delle Scienze 2 - Milano Italy
Gas family of proteins comprehends five members: Gas1-2-3-4- and Gas5p, Gas1p being the best-characterized. Gas1p is expressed during vegetative growth and is endowed of endo-beta(1,3) glucanase/beta(1,3)-Glucan transferase activity. This activity in vivo is required for the assembly of the cell wall. Gas2, Gas3, Gas4 and Gas5 proteins show different degrees of amino acid identity with Gas1p, ranging from 45% (Gas2p) to 33% (Gas3p). Gas proteins show also a different modular organization of domains with respect to Gas1p. All have the catalytic domain but the cys-box is present only in Gas1p and Gas2p. GAS2 and GAS4 are middle-meiotic induced genes whereas GAS3 appears to occur earlier during meiosis. GAS5 expression profile is restricted to vegetative growth. The ectopic expression of GAS2, GAS3 and GAS4 genes driven by the GAS1 promoter does not complement gas1 mutant phenotype whereas GAS5 does. Diploid null mutants were obtained. gas2 gas4 single or double null mutants showed specific defects in the maintenance of spore and ascus cell wall integrity. The proteins Gas1, Gas2, Gas3, Gas4 and Gas5 were expressed in a soluble, secreted HA-tagged form in Pichia pastoris. Gas2, Gas3 and Gas4 proteins were purified and found to be inactive in the enzymatic assays of beta-endoglucanase activity used for Gas1p. Thus Gas homologs, Gas2, Gas3 and Gas4, play specific roles in sporulation and have unique substrate specificity or requirements for optimal enzymatic activity.