Discovery of a Modified Tetrapolar Sexual Cycle in Cryptococcus amylolentus and the Evolution of MAT in the Cryptococcus Species Complex

Abstract

Sexual reproduction in fungi is governed by a specialized genomic region called the mating-type locus (MAT). The human fungal pathogenic and basidiomycetous yeast Cryptococcus neoformans has evolved a bipolar mating system (a, α) in which the MAT locus is unusually large (>100 kb) and encodes >20 genes including homeodomain (HD) and pheromone/receptor (P/R) genes. To understand how this unique bipolar mating system evolved, we investigated MAT in the closely related species Tsuchiyaea wingfieldii and Cryptococcus amylolentus and discovered two physically unlinked loci encoding the HD and P/R genes. Interestingly, the HD (B) locus sex-specific region is restricted (∼2 kb) and encodes two linked and divergently oriented homeodomain genes in contrast to the solo HD genes (SXI1α, SXI2 a) of C. neoformans and Cryptococcus gattii. The P/R (A) locus contains the pheromone and pheromone receptor genes but has expanded considerably compared to other outgroup species (Cryptococcus heveanensis) and is linked to many of the genes also found in the MAT locus of the pathogenic Cryptococcus species. Our discovery of a heterothallic sexual cycle for C. amylolentus allowed us to establish the biological roles of the sex-determining regions. Matings between two strains of opposite mating-types (A1B1×A2B2) produced dikaryotic hyphae with fused clamp connections, basidia, and basidiospores. Genotyping progeny using markers linked and unlinked to MAT revealed that meiosis and uniparental mitochondrial inheritance occur during the sexual cycle of C. amylolentus. The sexual cycle is tetrapolar and produces fertile progeny of four mating-types (A1B1, A1B2, A2B1, and A2B2), but a high proportion of progeny are infertile, and fertility is biased towards one parental mating-type (A1B1). Our studies reveal insights into the plasticity and transitions in both mechanisms of sex determination (bipolar versus tetrapolar) and sexual reproduction (outcrossing versus inbreeding) with implications for similar evolutionary transitions and processes in fungi, plants, and animals.

Molecular and Genetic Evidence for a Tetrapolar Mating System in the Basidiomycetous Yeast Kwoniella mangrovensis and Two Novel Sibling Species

“…The expansion of this nonrecombining sex-determining genomic region mirrors the models proposed for the genesis of sex chromosomes in multicellular eukaryotes (2). This study has since been expanded to include several other closely aligned sap-robic species in the order Tremellales (class Tremellomycetes, subphylum Agaricomycotina), namely, Cryptococcus amylolentus, Cryptococcus heveanensis, Filobasidiella depauperata, and Tsuchiyaea wingfieldii (7)(8)(9)(10). The latter studies disclosed sexual states in two species previously classified as asexual, viz.…”

Evolution of Mating Systems in Basidiomycetes and the Genetic Architecture Underlying Mating-Type Determination in the YeastLeucosporidium scottii

“…Since initial studies suggested that L. scottii is a tetrapolar species, we subsequently looked into the distribution and diversity of the HD1/HD2 alleles (B alleles) and their association with the two pheromone receptors (A alleles). If both loci are located on different chromosomes, as typically observed in tetrapolar species (Bakkeren et al 2006;Findley et al 2012), we should expect a random association between P/R and HD alleles in natural populations as a result of independent assortment of these alleles during meiosis. To investigate this, we amplified and sequenced a fragment spanning the 59 end and intergenic regions of the HD1 and HD2 genes in 43 strains of the L. scottii species complex.…”

“…This can be attained either by (i) loss of function of the P/R locus as an incompatibility factor (as, e.g., in Coprinus disseminatus, Pholiota microspore, and Phanerochaete chrysosporium) (Raper 1966;James et al 2006James et al , 2011Yi et al 2009) or (ii) coalescence of unlinked P/R and HD loci into a contiguous inheritable unit, with both MAT genes remaining functional in mating-type determination. This last scenario was shown to be at the basis of extant bipolar species belonging to two of the three major Basidiomycota lineages, namely Cryptococcus neoformans (subphylum Agaricomycotina) Findley et al 2012) and Ustilago hordei (subphylum Ustilaginomycotina) (Bakkeren et al 2006). In these cases, the MAT locus has expanded into a large, nonrecombining region of a chromosome that is usually highly rearranged between the two mating types and rich in repetitive elements Bakkeren et al 2006).…”

“…Interestingly, most of the known tetrapolar yeast species exhibit a biallelic P/R locus and a multiallelic HD locus (viz. Ustilago maydis, Kwoniella heveanensis, Kwoniella mangrovensis, Cryptococcus amylolentus) (Bölker et al 1992;Kämper et al 1995;Metin et al 2010;Findley et al 2012;Guerreiro et al 2013), a configuration originally designated as "modified tetrapolar" by Robert J. Bandoni to denote that outcrossing in such a system cannot exceed 50% (Bandoni 1963;Kües et al 2011). Indeed, only by evolving multiple alleles at both MAT loci can a tetrapolar system maximize the outcrossing efficiency to .50%, as observed in mushroomforming species (viz.…”

Cryptococcus gattii Infections

“…These species are distinct from the Ͼ80 species in the polyphyletic genus Cryptococcus, based on recent phylogenetic analysis of the D1/D2 region of the large-subunit (LSU) ribosomal DNA (rDNA) gene (3,4).…”