Functional molecular-genetic analysis of sexual reproduction in the fungus Botrytis cinerea

Botrytis cinerea, a typical necrotrophic plant pathogenic fungus that causes severe loss in crop production, performs sexual reproduction. This fungus displays a heterothallic (self-incompatible) lifestyle, which means it requires a compatible partner with opposite mating type for sexual reproduction to occur. The MAT (mating type) locus, containing MAT genes, determines the mating types and the sexual lifestyle of B. cinerea. However, little is known on the function of mating type genes MAT1-1-5 and MAT1-2-10 in sexual reproduction. I propose that MAT1-1-5 and MAT1-2-10 play important roles in sexual reproduction by regulating signal transduction cascades and/or controlling the expression of genes. It was reported that the heterothallic fungus C. heterostrophus can be converted to homothallism by introducing the MAT1-1-1 and MAT1-2-1 genes from the homothallic species C. luttrelli into a mat-deleted strain of C. heterostrophus. I will first transfer the B. cinerea MAT1-2 idiomorph to the sequenced reference strain B05.10 (containing the MAT1-1 locus) to generate a homothallic strain which is self-compatible without requiring a mating partner. Previous research projects in WU Phytopathology showed that MAT1-1-5 and MAT1-2-10 jointly control the developmental transition from an apothecial stipe to a disk and it was suggested that they act as (possibly dimeric) regulators that directly or indirectly control the formation of crozier cells. The absence of either of these proteins presumably results in failure to proceed to karyogamy, which blocks further downstream processes such as apothecial disk expansion. I will study the physical interaction between MAT1-1-5 and MAT1-2-10 proteins, and determine whether the interaction contributes to sexual reproduction. Subsequently, downstream target proteins and genes will be explored by proximity labelling and CHIP-seq assays. It was reported that pheromone receptors are regulated by MAT genes in heterothallic fungi, while in homothallic fungi the pheromone receptors seem to regulate the MAPK signalling pathway, the master regulator of conjugation and meiosis, acting upstream of mating-type genes. However, the function of pheromone receptors in B. cinerea is unclear. I will investigate whether the pheromone receptors are dispensable in homothallic B. cinerea, whether the mating type genes can regulate pheromone receptors or vice versa, and screen the target proteins of pheromone receptors in sexual reproduction. The proposed project will help to clarify the function of mating type genes and pheromone receptors in sexual reproduction, and bring new insight to the mechanism underlying sexual reproduction in B. cinerea.