Academic journal article Genetics

A Tetrad Analysis of the Basidiomycete Fungus Cryptococcus Neoformans

Academic journal article Genetics

A Tetrad Analysis of the Basidiomycete Fungus Cryptococcus Neoformans

Article excerpt

ABSTRACT

Cryptococcus neoformans is a basidiomycete fungus that is found worldwide and causes disease in humans and animal species. The fungus grows asexually as a budding yeast. Under laboratory conditions it is capable of sexual reproduction between two mating types. After cell fusion a dikaryotic filament develops, at the tip of which a basidium gives rise to four chains of basidiospores. Because the chains each comprise 10-30 spores, rather than single spores, the analysis of individual meiotic events has not been attempted in C. neoformans in the style of tetrad analyses performed in other fungal species. Here, the basidiospores from >100 basidia were micromanipulated and the resultant >2500 progeny analyzed for three genetic markers to understand the sexual process in this fungus, leading to four observations: (i) Marker segregation provides genetic evidence for a single meiotic event within the basidium followed by multiple rounds of mitosis. (ii) Using each basidium as an unordered tetrad, the ADE2 and URA5 genes are linked to their centromeres, consistent with adjacent genomic regions rich in repetitive elements predicted to comprise Cryptococcus centromeres. (iii) Lack of germination of basidiospores is attributed to aneuploidy, rather than dormancy. (iv) Analysis of basidiospores derived from single chains demonstrates that each chain can contain different genotypes. This mechanism of sexual spore production would benefit the species with a high rate of dispersal and at the same time aid in simultaneous dissemination of both mating types to new locations in the environment.

CRYPTOCOCCUS neoformans is a fungus distributed worldwide and the cause of cryptococcosis disease in humans and animal species (Mitchell and Perfect 1995; Casadevall and Perfect 1998). The fungus grows as a budding yeast, which, as a morphological trait, hampered an understanding of its phylogenetic position within the fungal kingdom. In the mid-1970s a sexual cycle was characterized to demonstrate that the species belongs in the phylum Basidiomycota, because it generates a filamentous sexual state that results in spore production from a basidium structure (Kwon-Chung 1975). C. neoformans is divided into two serotypes and is closely related to C. gattii that also causes cryptococcosis. Unlikemany other basidiomycetes that undergo meiosis to produce four spores,Cryptococcus andits relatives are distinguishedby basidia with four long chains of attached spores. Cytological analysis of the nuclear behavior in mating reactions demonstrated that two nuclei are present in the filaments and are as such dikaryotic, while in the basidium either a larger diploid nucleus or multiple nuclei expected to be the products of meiosis are present (Kwon-Chung 1976). Mating occurs between cells of two mating types (MATα and MATα) in a bipolar system (Kwon-Chung 1976), although a monokaryotic fruiting process of self-self mating can also occur under nutrient poor conditions (Wickes et al. 1996; Lin et al. 2005). Initial analysis of two genetic markers and subsequent Mendelian genetic analyses, leading to a full genetic map, demonstrated that meiosis occurs in the basidium (Kwon-Chung 1976;Marra et al. 2004). It is presumed that in each basidium there is a single meiotic event followed by rounds of mitosis to give rise to the chains of spores (Hull and Heitman 2002; McClelland et al. 2004; Lin 2009), although this process in the basidium has not been shown genetically.

In addition to its use as an experimental tool, mating in C. neoformans is potentially of medical relevance. First, it is hypothesized that the basidiospores are the infectious particles because they are sufficiently smaller in size than yeast cells, such that they would penetrate the lung alveoli to establish disease (Cohen et al. 1982), and recent studies have shown that spores are equally virulent as yeast cells (Giles et al. 2009; Velagapudi et al. 2009). Mating is supported by population genetic evidence of recombination (Litvintseva et al. …

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