Functional analysis of genes in the mating type locus of Botrytis cinerea

Botrytis cinerea is a heterothallic ascomycete with two mating types, MAT1-1 and MAT1-2, each containing two genes. Besides the archetypal genes encoding the MAT1-1-1 (alpha-domain) protein and the MAT1-2-1 (HMG-box) protein, each idiomorph contains one additional gene, designated MAT1-1-5 and MAT1-2-4, respectively. Homologs of these genes are only found in closely related taxa, and their function is as yet unknown. Knockout mutants were generated in all four genes in the B. cinerea MAT locus, either in the MAT1-1 strain SAS56 or in the MAT1-2 strain SAS405. Mutants were crossed with a strain of the opposite mating type, either the wild type or a knockout mutant, in all possible combinations. Knockout mutants in the MAT1-1-1 gene and the MAT1-2-1 gene fail to show any sign of primordial outgrowth and are entirely sterile. This confirms the essential role of the alpha-domain protein and the HMG-box protein in the mating process. By contrast, mutants in the MAT1-1-5 gene and the MAT1-2-4 gene do produce stipes, but these fail to develop further into an apothecial disk. The MAT1-1-5 and MAT1-2-4 mutants show identical phenotypes, suggesting that these two genes jointly control the transition from stipe to disk development. RNA-seq data were obtained from a cross between two wild type strains and from a cross involving a MAT1-1-5 knockout mutant, from tissue at the stage of transition from stipe to disk. Differential gene expression analysis was performed to identify genes that are possibly involved in development of the apothecial disk.