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Cryptococcus neoformans STE12alpha regulates virulence but is not essential for mating.

Chang YC, Wickes BL, Miller GF, Penoyer LA, Kwon-Chung KJ - J. Exp. Med. (2000)

Bottom Line: Furthermore, ste12alpha disruptants were fertile when mated with MATa strains, albeit with reduced mating frequency.Using reporter gene constructs, we found that STE12alpha controls the expression of several phenotypes known to be involved in virulence, such as capsule and melanin production.These results demonstrate a clear molecular link between mating type and virulence in C. neoformans.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Clinical Investigation, National Institute of Allergy and Infectious Diseases, Office of the Director, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The Cryptococcus neoformans STE12alpha gene, a homologue of Saccharomyces cerevisiae STE12, exists only in mating type (MAT)alpha cells. In S. cerevisiae, STE12 was required for mating and filament formation. In C. neoformans, haploid fruiting on filament agar required STE12alpha. The ability to form hyphae, however, was not affected by deletion of STE12alpha when convergently growing MATa strains were present. Furthermore, ste12alpha disruptants were fertile when mated with MATa strains, albeit with reduced mating frequency. Most importantly, the virulence of a ste12alpha disruptant of serotype D strain was significantly reduced in a mouse model. When the ste12alpha locus was reconstituted with the wild-type allele by cotransformation, virulence was restored. Histopathological analysis demonstrated a reduction in capsular size of yeast cells, less severe cystic lesions, and stronger immune responses in meninges of mice infected with ste12alpha cells than those of mice infected with STE12alpha cells. Using reporter gene constructs, we found that STE12alpha controls the expression of several phenotypes known to be involved in virulence, such as capsule and melanin production. These results demonstrate a clear molecular link between mating type and virulence in C. neoformans.

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Hyphal formation on SLAD medium. (A) B-4500 × B-4476. (B) TYCC245F1 × TYCC245F1. (C) TYCC245F1 × B-4476. Cells of each isolate were streaked in parallel on SLAD medium and incubated at room temperature for 24 h (bar = 15 μm).
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Figure 5: Hyphal formation on SLAD medium. (A) B-4500 × B-4476. (B) TYCC245F1 × TYCC245F1. (C) TYCC245F1 × B-4476. Cells of each isolate were streaked in parallel on SLAD medium and incubated at room temperature for 24 h (bar = 15 μm).

Mentions: We have previously observed that MATα cells produce hyphae within 24 h when streaked in close proximity (<500 μm) with MATa cells on SLAD medium (our unpublished observation). SLAD medium has a composition similar to filament agar and has been shown to induce pseudohyphal formation in S. cerevisiae 12. When MATα cells of C. neoformans were grown alone on SLAD medium, no hyphae were produced. It was of interest to investigate whether ste12α disruptants lost the ability to produce hyphae on SLAD medium in response to the presence of MATa cells. Fig. 5 A shows hyphal production by B-4500 (STE12α, MATα) when streaked in parallel to a streak of B-4476 (MATa) within a 500-μm distance, whereas MATa cells failed to respond in a similar manner (no hyphal filaments were produced). Similarly, no hyphal structures were detected when two streaks of ste12α (TYCC245F1) cells were paired on SLAD agar (Fig. 5 B). TYCC245F1 produced hyphae only when it was streaked in parallel with MATa cells in close proximity (<500 μm distance; Fig. 5 C). The hyphae produced by TYCC245F1 (Fig. 5 C), however, were morphologically different, tending to be more sinuous compared with hyphae produced by B-4500 (Fig. 5 A). When plates containing parallel streaks of opposite mating type strains were incubated for 1 wk, hyphae produced by the MATα strain reached MATa cells, and both basidia and spore chains were produced sporadically at the margin of MATα streaks (data not shown). Similar results were observed on SLAD agar without ammonium sulfate, as well as on minimal medium (SLAD agar with 37.8 mM ammonium sulfate). It is clear, therefore, that there are at least two different signaling pathways leading to hyphal formation in C. neoformans. These observations suggest that although the ste12α disruptant lost its ability to undergo haploid fruiting in response to nitrogen starvation, it retained the ability to form hyphae in response to the presence of MATa cells on SLAD agar.


Cryptococcus neoformans STE12alpha regulates virulence but is not essential for mating.

Chang YC, Wickes BL, Miller GF, Penoyer LA, Kwon-Chung KJ - J. Exp. Med. (2000)

Hyphal formation on SLAD medium. (A) B-4500 × B-4476. (B) TYCC245F1 × TYCC245F1. (C) TYCC245F1 × B-4476. Cells of each isolate were streaked in parallel on SLAD medium and incubated at room temperature for 24 h (bar = 15 μm).
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Related In: Results  -  Collection

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Figure 5: Hyphal formation on SLAD medium. (A) B-4500 × B-4476. (B) TYCC245F1 × TYCC245F1. (C) TYCC245F1 × B-4476. Cells of each isolate were streaked in parallel on SLAD medium and incubated at room temperature for 24 h (bar = 15 μm).
Mentions: We have previously observed that MATα cells produce hyphae within 24 h when streaked in close proximity (<500 μm) with MATa cells on SLAD medium (our unpublished observation). SLAD medium has a composition similar to filament agar and has been shown to induce pseudohyphal formation in S. cerevisiae 12. When MATα cells of C. neoformans were grown alone on SLAD medium, no hyphae were produced. It was of interest to investigate whether ste12α disruptants lost the ability to produce hyphae on SLAD medium in response to the presence of MATa cells. Fig. 5 A shows hyphal production by B-4500 (STE12α, MATα) when streaked in parallel to a streak of B-4476 (MATa) within a 500-μm distance, whereas MATa cells failed to respond in a similar manner (no hyphal filaments were produced). Similarly, no hyphal structures were detected when two streaks of ste12α (TYCC245F1) cells were paired on SLAD agar (Fig. 5 B). TYCC245F1 produced hyphae only when it was streaked in parallel with MATa cells in close proximity (<500 μm distance; Fig. 5 C). The hyphae produced by TYCC245F1 (Fig. 5 C), however, were morphologically different, tending to be more sinuous compared with hyphae produced by B-4500 (Fig. 5 A). When plates containing parallel streaks of opposite mating type strains were incubated for 1 wk, hyphae produced by the MATα strain reached MATa cells, and both basidia and spore chains were produced sporadically at the margin of MATα streaks (data not shown). Similar results were observed on SLAD agar without ammonium sulfate, as well as on minimal medium (SLAD agar with 37.8 mM ammonium sulfate). It is clear, therefore, that there are at least two different signaling pathways leading to hyphal formation in C. neoformans. These observations suggest that although the ste12α disruptant lost its ability to undergo haploid fruiting in response to nitrogen starvation, it retained the ability to form hyphae in response to the presence of MATa cells on SLAD agar.

Bottom Line: Furthermore, ste12alpha disruptants were fertile when mated with MATa strains, albeit with reduced mating frequency.Using reporter gene constructs, we found that STE12alpha controls the expression of several phenotypes known to be involved in virulence, such as capsule and melanin production.These results demonstrate a clear molecular link between mating type and virulence in C. neoformans.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Clinical Investigation, National Institute of Allergy and Infectious Diseases, Office of the Director, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The Cryptococcus neoformans STE12alpha gene, a homologue of Saccharomyces cerevisiae STE12, exists only in mating type (MAT)alpha cells. In S. cerevisiae, STE12 was required for mating and filament formation. In C. neoformans, haploid fruiting on filament agar required STE12alpha. The ability to form hyphae, however, was not affected by deletion of STE12alpha when convergently growing MATa strains were present. Furthermore, ste12alpha disruptants were fertile when mated with MATa strains, albeit with reduced mating frequency. Most importantly, the virulence of a ste12alpha disruptant of serotype D strain was significantly reduced in a mouse model. When the ste12alpha locus was reconstituted with the wild-type allele by cotransformation, virulence was restored. Histopathological analysis demonstrated a reduction in capsular size of yeast cells, less severe cystic lesions, and stronger immune responses in meninges of mice infected with ste12alpha cells than those of mice infected with STE12alpha cells. Using reporter gene constructs, we found that STE12alpha controls the expression of several phenotypes known to be involved in virulence, such as capsule and melanin production. These results demonstrate a clear molecular link between mating type and virulence in C. neoformans.

Show MeSH
Related in: MedlinePlus