WdStuAp, an APSES transcription factor, is a regulator of yeast-hyphal transitions in Wangiella (Exophiala) dermatitidis

Abstract

APSES transcription factors are well-known regulators of fungal cellular development and differentiation. To study the function of an APSES protein in the fungus Wangiella dermatitidis, a conidiogenous and polymorphic agent of human phaeohyphomycosis with yeast predominance, the APSES transcription factor gene WdSTUA was cloned, sequenced, disrupted, and overexpressed. Analysis showed that its derived protein was most similar to the APSES proteins of other conidiogenous molds and had its APSES DNA-binding domain located in the amino-terminal half. Deletion of WdSTUA in W. dermatitidis induced convoluted instead of normal smooth colony surface growth on the rich yeast maintenance agar medium yeast extract-peptone-dextrose agar (YPDA) at 37 degrees C. Additionally, deletion of WdSTUA repressed aerial hyphal growth, conidiation, and invasive hyphal growth on the nitrogen-poor, hypha-inducing agar medium potato dextrose agar (PDA) at 25 degrees C. Ectopic overexpression of WdSTUA repressed the convoluted colony surface growth on YPDA at 37 degrees C, and also strongly repressed hyphal growth on PDA at 25 degrees C and 37 degrees C. These new results provide additional insights into the diverse roles played by APSES factors in fungi. They also suggest that the transcription factor encoded by WdSTUA is both a positive and negative morphotype regulator in W. dermatitidis and possibly other of the numerous human pathogenic, conidiogenous fungi capable of yeast growth.

FIG. 1.

Effects of WdSTUA disruption on growth and colony morphology on YPDA at 25°C and 37°C. (A) Yeast cells were spotted (5 μl) at 10⁵, 10⁴, 10³, and 10² concentrations on YPDA and incubated at 25°C and 37°C. After 4 days, the growth at the spots was photographed without magnification, whereas the surface morphologies at the spots inoculated with the 10² yeast concentration were visualized with a dissecting microscope and also photographed. WT, wild type. (B) Wild-type and WdstuaΔ1A strains cultured with shaking in YPDB at 37°C to late log phase were visualized with a compound light microscope and then photographed. Note that the WdstuaΔ1A strain produced more hyphae that tended to be aggregated. Scale bar, 10 μm (applicable to the growth in both photomicrographs).

FIG. 2.

The WdSTUA deletion reduced W. dermatitidis filamentous growth on PDA at 25°C. The wild-type (WT) and WdstuaΔ1A strains were streaked on PDA and incubated at 25°C (A) and 37°C (B). The filamentous growth at the colony edges was visualized with a compound light microscope, and the photomicrographs were taken after 4 days of incubation. Scale bar, 0.2 mm (applicable to all colonies in the figure).

FIG. 3.

The WdSTUA deletion repressed aerial hyphal growth and consequently conidiogenesis. The wild-type (WT) and WdstuaΔ1A strains were inoculated on PDA in slide cultures and incubated at 25°C. Growth at the edge of the medium and protruding into the air space between the slide and coverslip was visualized with a compound light microscope and photographed after 2 weeks of incubation. Scale bar, 10 μm (applicable to all the growth in both photomicrographs). The arrow points to a cluster of conidia produced at the terminus of a conidiogenous hypha.

FIG. 4.

The WdstuaΔ mutant was defective in invasive growth. (A) Wild-type (WT) and WdstuaΔ1A cells were spotted on PDA and incubated at 25°C. After incubation for 8 days, colonies were visualized with a dissecting microscope and photographed. (B) The biomass above the agar was then rinsed away and the remaining, invasive growth photographed again. (C) Cross sections of the growth that invaded the agar. (D) The morphotypes in the invasive growth visualized with a light microscope. Scale bar, 10 μm (applicable to the growth in both photomicrographs).

FIG. 5.

WdSTUA deletion in the Hf1 strain inhibited filamentous growth on PDA. Wild-type (WT), WdstuaΔ1A, Hf1, and Hf1 WdstuaΔ1 strains were grown on YPDA at 37°C for 6 days (A) and on PDA at 25°C for 6 days (B). The Hf1 and Hf1 WdstuaΔ1 strains were also inoculated on PDA for slide culture at 25°C. After 2 weeks, conidiophores and conidia were visualized with a light microscope and photographed (C). Scale bars, 10 μm (applicable to the growth in both photomicrographs). The arrows point to clusters of conidia produced at the termini of conidiogenous hyphae.

FIG. 6.

Ectopic overexpression of WdSTUA in W. dermatitidis. Strains were grown on YPDA at 37°C (A) and PDA at 37°C (B) and 25°C (C). After 5 days of incubation, colonies were photographed with the aid of a dissecting microscope.