The Predicted G-Protein-Coupled Receptor GPR-1 Is Required for Female Sexual Development inthe Multicellular Fungus Neurospora crassa

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Abstract
G-protein-coupled receptors (GPCRs) control important aspects of asexual and sexual development in eukaryotic organisms. We have identified a predicted GPCR in the filamentous fungus Neurospora crassa with similarity to cyclic AMP-receptor like GPCRs from Dictyostelium discoideum and GCR1 from Arabidopsis thaliana . Expression of gpr-1 is highest in female reproductive structures, and deletion of gpr-1 leads to defects during sexual development. Unfertilized female structures (protoperithecia) fromΔ gpr-1 strains are weakly pigmented, small, and submerged in the agar. The perithecia produced after fertilization have deformed beaks that lack ostioles, the openings through which ascospores are discharged. Localization studies using a GPR-1-green fluorescent protein fusion protein showed that GPR-1 is targeted to female reproductive structures. Genetic epistasis experiments with the three Gα genes were inconclusive due to the early block in mating exhibited by Δ gna-1 strains. Phenotypic analysis of mutants from a high-throughput N. crassa knockout project allowed identification of BEK-1, a homeodomain transcription factor that is a potential target of GPR-1. The perithecial defects ofΔ bek-1 strains are similar to those of theΔ gpr-1 strain, and epistasis analysis indicates that bek-1 could function downstream of gpr-1 during postfertilization events. The effect must be posttranscriptional, as bek-1 transcript levels are not affected inΔ gpr-1 strains. The lack of ostioles inΔ gpr-1 and Δ bek-1 mutants has an undesirable effect on the ability to spread progeny (ascospores) by the normal ejection mechanism and would severely compromise the fitness of these strains in nature.