DIFFERENTIAL EXPRESSION AND FUNCTION OF 2 HOMOLOGOUS SUBUNITS OF YEAST 1,3-BETA-D-GLUCAN SYNTHASE

1,3-beta-D-Glucan is a major structural polymer of yeast and fungal ce ll walls and is synthesized from UDP-glucose by the multisubunit enzym e 1,3-beta-D-glucan synthase. Previous work has shown that the FKS1 ge ne encodes a 215-kDa integral membrane protein (Fks1p) which mediates sensitivity to the echinocandin class of antifungal glucan synthase in hibitors and is a subunit of this enzyme, We have cloned and sequenced FKS2, a homolog of FKS1 encoding a 217-kDa integral membrane protein (Fks2p) which is 88% identical to Fks1p, The residual glucan synthase activity present in strains with deletions of fks1 is (i) immunodeplet ed by antibodies prepared against FKS2 peptides, demonstrating that Fk s2p is also a component of the enzyme, and (ii) more sensitive to the echinocandin L-733,560, explaining the increased sensitivity of fks1 n ull mutants to this drug, Simultaneous disruption of FKS1 and FKS2 is lethal, suggesting that Fks1p and Fks2p are alternative subunits with essential overlapping function, Analysis of FKS1 and FKS2 expression r eveals that transcription of FKS1 is regulated in the cell cycle and p redominates during growth on glucose, while FKS2 is expressed in the a bsence of glucose. FKS2 is essential for sporulation, a process which occurs during nutritional starvation, FKS2 is induced by the addition of Ca2+ to the growth medium, and this induction is completely depende nt on the Ca2+/calmodulin-dependent phosphoprotein phosphatase calcine urin, We have previously shown that growth of fks1 null mutants is hig hly sensitive to the calcineurin inhibitors FK506 and cyclosporin A, E xpression of FKS2 from the heterologous ADH1 promoter results in FK506 -resistant growth, Thus, the sensitivity of fks1 mutants to these drug s can be explained by the calcineurin-dependent transcription of FKS2, Moreover, FKS2 is also highly induced in response to pheromone in a c alcineurin-dependent manner, suggesting that FKS2 may also play a role in the remodeling of the cell wall during the mating process.