PAPULACANDIN-B RESISTANCE IN BUDDING AND FISSION YEASTS - ISOLATION AND CHARACTERIZATION OF A GENE INVOLVED IN (1,3)BETA-D-GLUCAN SYNTHESISIN SACCHAROMYCES-CEREVISIAE

Papulacandin B, an antifungal agent that interferes with the synthesis of yeast cell wall (1,3)beta-D-glucan, was used to isolate resistant mutants in Schizosaccharomyces pombe and Saccharomyces cerevisiae. The resistance to papulacandin B always segregated as a recessive charact er that defines a single complementation group in both yeasts (pbr1(+) and PBR1, respectively). Determination of several kinetic parameters of (1,3)beta-D-glucan synthase activity revealed no differences betwee n S. pombe wild-type and pbr1 mutant strains except in the 50% inhibit ory concentration for papulacandin B of the synthases (about a 50 fold increase in mutant activity). Inactivation of the synthase activity o f both yeasts after in vivo treatment with the antifungal agent showed that mutant synthases were more resistant than the corresponding wild -type ones. Detergent dissociation of the S. pombe synthase into solub le and particulate fractions and subsequent reconstitution indicated t hat the resistance character of pbr1 mutants resides in the particulat e fraction of the enzyme. Cloning and sequencing of PBR1 from S. cerev isiae revealed a gene identical to others recently reported (FKS1, ETG 1, CWH53, and CND1). Its disruption leads to reduced levels of both (1 ,3)beta-D-glucan synthase activity and the alkali-insoluble cell wall fraction. Transformants containing the PBR1 gene reverse the defect in (1,3)beta-D-glucan synthase. It is concluded that Pbr1p is probably p art of the (1,3)beta-D-glucan synthase complex.