Long-term balancing selection in highly dynamic mating loci generates trans-species polymorphisms in fungi

Balancing selection, an evolutionary force that retains genetic diversity, has been detected in multiple genes and organisms, such as the sexual mating loci in fungi. In tetrapolar basidiomycete fungi, sexual type is determined by two unlinked loci, MATA and MATB. These loci are usually highly diverse, but with conserved domains. Previous studies have revealed that species of the genus Trichaptum (Hymenochaetales, Basidiomycota) possess a tetrapolar mating system, with multiple inferred alleles for MATA and MATB. Here, we sequenced a total of a hundred and eighty specimens of three Trichaptum species. We characterized the chromosomal location of MATA (chromosome 2) and MATB (chromosome 9), the molecular structure of MAT regions and their allelic richness. We found multiple MAT alleles segregating in both multiple Trichaptum specimens, and the non-Trichaptum species included for comparison. Phylogenetic analyses and various nucleotide statistics suggested that long-term balancing selection has generated trans-species polymorphisms. Mating sequences were classified in different allelic classes based on an identity threshold of higher than 86%. The observed allelic classes could potentially generate 14,560 different mating types. The inferred allelic information mirrored the outcome of in vitro crosses, thus allowing us to support the degree of allelic divergence needed for successful mating. Even with the high amount of divergence, key amino acids in functional domains are conserved. We conclude that the genetic diversity of mating in Trichaptum loci is due to long-term balancing selection that likely promote sexual outcrossing, with limited recombination and duplication activity. Our large number of sequenced specimens highlighted the importance of sequencing multiple individuals from different species to detect the mating-related genes, the mechanisms generating diversity and the evolutionary forces maintaining them.