The overall aim of this research is to understand how a simple eukaryote distinguishes itself from many other nonselves in regulating mating and development. A specific objective is to elucidate the molecular mechanism by which the B mating-type genes trigger sexual development in the wood rotting mushroom Schizophyllum commune. The B genes are in two linked loci, Ba and Bb, each of which has nine different versions, or specificities, in the world-wide population. A mating pair must be different at either Ba or Bb to be compatible. Analyses of two specificities of each locus has revealed genes for pheromones and pheromone receptors similar to those found in other fungi. Each locus contains a single pheromone receptor gene and several pheromone genes. This multiplicity of pheromone and receptor genes appears to be unique to the higher basidiomycete fungi. Molecular analyses to date indicate that as few as one or two amino acid changes in either a pheromone or receptor molecule can be correlated with a switch in mate recognition and signaling of sexual. The proposed research is divided into three parts: 1) A comprehensive comparison of sequence and function for a variety of natural and previously generated mutant variants of pheromones and receptors to identify amino acid residues in positions relevant to pheromone/receptor interactions. 2) Test the effects of substituting various amino acids at specific sites that could be relevant to pheromone/receptor recognition for each type of molecule, using the methods of site-directed mutagenesis and DNA-mediated transformation in S. commune. 3) Exploit a recently assembled heterologous pheromone-response reporter system in the yeast Saccharomyces cerevisiae to generate new mutant variants of S. commune pheromones and receptors and efficiently characterize their effects on ligand/receptor interaction.
