FEBS advanced lecture course on Human Fungal pathogens edition:1 location:La Colle sur Loup, France date:21-28 May 2005
We investigate the nutrient sensing role of the G protein coupled receptor Gpr1 and the Galpha protein Gpa2 in Candida albicans by using the model yeast Saccharomyces cerevisiae. In S. cerevisiae Gpr1 and Gpa2 form a GPCR nutrient sensing system. We have recently shown that in the model yeast this GPCR system acts upstream of the cAMP-PKA pathway and that glucose and sucrose are the ligands of Gpr1 (Lemaire et al., 2004, Mol. Cell). We have now investigated whether the same is true in C. albicans. In a recent paper we already point out that CaGpr1 presumably lies upstream of the cAMP-PKA pathway in this organism as well, and that it interacts with CaGpa2 (Maidan et al, 2005, MBC). The nature of the ligand interacting with CaGpr1, however, has not been found yet. Our current results indicate that this ligand might be a sugar, such as glucose, or amino acids, since formation of hyphae could be induced by simultaneous presence of specific concentrations of glucose and methionine, whereas this could not be achieved in a gpr1 double deletion strain (Maidan et al, 2005, BST). To gain further insights in this matter, we use two approaches. On the one hand, we express the CaGpr1 and CaGpa2 protein, separately or simultaneously, in the corresponding yeast mutants to check possible complementation. To confirm complementation, restoration of the cAMP signal is checked, as well as pseudohyphal growth. On the other hand, we will make use of dominant alleles of Gpr1 or Gpa2. We constructed the dominant GPA2Q355L allele (as described by Sanchez-Martinez and Perez-Martin, 2002) and confirmed that it can suppress the gpr1 deletion phenotype. We will express this allele in several strains containing deletions in the MAP kinase or cAMP-PKA pathways. We will use PCR mutagenesis to identify dominant GPR1 alleles and if obtained, we will perform similar experiments as for the dominant GPA2 allele.