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N-acetylglucosamine induces white to opaque switching, a mating prerequisite in Candida albicans.

Huang G, Yi S, Sahni N, Daniels KJ, Srikantha T, Soll DR - PLoS Pathog. (2010)

Bottom Line: Paradoxically, opaque cells were found to be unstable at physiological temperature, suggesting that mating had little chance of occurring in the host, the main niche of C. albicans.Recently, however, it was demonstrated that high levels of CO(2), equivalent to those found in the host gastrointestinal tract and select tissues, induced the white to opaque switch at physiological temperature, providing a possible resolution to the paradox.Our results therefore suggest that signals produced by bacterial co-members of the gastrointestinal tract microbiota regulate switching and therefore mating of C. albicans.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, The University of Iowa, Iowa City, Iowa, United States of America.

ABSTRACT
To mate, the fungal pathogen Candida albicans must undergo homozygosis at the mating-type locus and then switch from the white to opaque phenotype. Paradoxically, opaque cells were found to be unstable at physiological temperature, suggesting that mating had little chance of occurring in the host, the main niche of C. albicans. Recently, however, it was demonstrated that high levels of CO(2), equivalent to those found in the host gastrointestinal tract and select tissues, induced the white to opaque switch at physiological temperature, providing a possible resolution to the paradox. Here, we demonstrate that a second signal, N-acetylglucosamine (GlcNAc), a monosaccharide produced primarily by gastrointestinal tract bacteria, also serves as a potent inducer of white to opaque switching and functions primarily through the Ras1/cAMP pathway and phosphorylated Wor1, the gene product of the master switch locus. Our results therefore suggest that signals produced by bacterial co-members of the gastrointestinal tract microbiota regulate switching and therefore mating of C. albicans.

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Wor1 localizes to the nucleus in the phosphorylated and unphosphorylated state.A. GFP fluorescence of Wor1 and Wor1TA, in parallel with DAPI staining of nuclei, of strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA, respectively. Cells were grown in glucose liquid medium containing 200 µg per ml of doxycycline at 25°C. B. Western blot analysis of Wor1 and Wor1TA using anti-GFP antibody in strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA. Protein extracts were derived from cells grown at 25°C in glucose liquid medium containing 200 µg per ml of doxycycline.
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ppat-1000806-g005: Wor1 localizes to the nucleus in the phosphorylated and unphosphorylated state.A. GFP fluorescence of Wor1 and Wor1TA, in parallel with DAPI staining of nuclei, of strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA, respectively. Cells were grown in glucose liquid medium containing 200 µg per ml of doxycycline at 25°C. B. Western blot analysis of Wor1 and Wor1TA using anti-GFP antibody in strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA. Protein extracts were derived from cells grown at 25°C in glucose liquid medium containing 200 µg per ml of doxycycline.

Mentions: Because WOR1 and WOR1TA were fused in frame with GFP in the overexpression mutants, we used confocal microscopy to test whether Wor1TA localized normally to the nucleus and was expressed at the same level as Wor1. Both Wor1 and Wor1TA localized to the nucleus of a majority of cells of the overexpression mutants treated with doxycycline, as demonstrated by overlapping GFP fluorescence and staining with DAPI, a DNA indicator (Figure 5A). Moreover, GFP fluorescence of nuclei was qualitatively comparable for Wor1-GFP and Wor1TA-GFP (Figure 5A). These results demonstrated that although the replacement of threonine with alanine caused a dramatic decrease in its capacity to support switching, it did not affect nuclear localization or cause a decrease in the transcript level. The levels of the Wor1 and Wor1TA protein were then compared by western blot analysis using anti-GFP antibody. The levels of Wor1 and Wor1TA expressed in white cells of strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA, respectively, treated with 200 µg per ml of doxycycline were similar (Figure 5B). These results indicate that the decrease in Wor1 function resulting from the replacement of threonine with alanine in the PKA consensus motif of Wor1 was due to a decrease in function, rather than to a decrease in the level of the Wor1 protein or mis-localization.


N-acetylglucosamine induces white to opaque switching, a mating prerequisite in Candida albicans.

Huang G, Yi S, Sahni N, Daniels KJ, Srikantha T, Soll DR - PLoS Pathog. (2010)

Wor1 localizes to the nucleus in the phosphorylated and unphosphorylated state.A. GFP fluorescence of Wor1 and Wor1TA, in parallel with DAPI staining of nuclei, of strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA, respectively. Cells were grown in glucose liquid medium containing 200 µg per ml of doxycycline at 25°C. B. Western blot analysis of Wor1 and Wor1TA using anti-GFP antibody in strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA. Protein extracts were derived from cells grown at 25°C in glucose liquid medium containing 200 µg per ml of doxycycline.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2837409&req=5

ppat-1000806-g005: Wor1 localizes to the nucleus in the phosphorylated and unphosphorylated state.A. GFP fluorescence of Wor1 and Wor1TA, in parallel with DAPI staining of nuclei, of strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA, respectively. Cells were grown in glucose liquid medium containing 200 µg per ml of doxycycline at 25°C. B. Western blot analysis of Wor1 and Wor1TA using anti-GFP antibody in strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA. Protein extracts were derived from cells grown at 25°C in glucose liquid medium containing 200 µg per ml of doxycycline.
Mentions: Because WOR1 and WOR1TA were fused in frame with GFP in the overexpression mutants, we used confocal microscopy to test whether Wor1TA localized normally to the nucleus and was expressed at the same level as Wor1. Both Wor1 and Wor1TA localized to the nucleus of a majority of cells of the overexpression mutants treated with doxycycline, as demonstrated by overlapping GFP fluorescence and staining with DAPI, a DNA indicator (Figure 5A). Moreover, GFP fluorescence of nuclei was qualitatively comparable for Wor1-GFP and Wor1TA-GFP (Figure 5A). These results demonstrated that although the replacement of threonine with alanine caused a dramatic decrease in its capacity to support switching, it did not affect nuclear localization or cause a decrease in the transcript level. The levels of the Wor1 and Wor1TA protein were then compared by western blot analysis using anti-GFP antibody. The levels of Wor1 and Wor1TA expressed in white cells of strains wor1/wor1+TETp-WOR1 and wor1/wor1+TETp-WOR1TA, respectively, treated with 200 µg per ml of doxycycline were similar (Figure 5B). These results indicate that the decrease in Wor1 function resulting from the replacement of threonine with alanine in the PKA consensus motif of Wor1 was due to a decrease in function, rather than to a decrease in the level of the Wor1 protein or mis-localization.

Bottom Line: Paradoxically, opaque cells were found to be unstable at physiological temperature, suggesting that mating had little chance of occurring in the host, the main niche of C. albicans.Recently, however, it was demonstrated that high levels of CO(2), equivalent to those found in the host gastrointestinal tract and select tissues, induced the white to opaque switch at physiological temperature, providing a possible resolution to the paradox.Our results therefore suggest that signals produced by bacterial co-members of the gastrointestinal tract microbiota regulate switching and therefore mating of C. albicans.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, The University of Iowa, Iowa City, Iowa, United States of America.

ABSTRACT
To mate, the fungal pathogen Candida albicans must undergo homozygosis at the mating-type locus and then switch from the white to opaque phenotype. Paradoxically, opaque cells were found to be unstable at physiological temperature, suggesting that mating had little chance of occurring in the host, the main niche of C. albicans. Recently, however, it was demonstrated that high levels of CO(2), equivalent to those found in the host gastrointestinal tract and select tissues, induced the white to opaque switch at physiological temperature, providing a possible resolution to the paradox. Here, we demonstrate that a second signal, N-acetylglucosamine (GlcNAc), a monosaccharide produced primarily by gastrointestinal tract bacteria, also serves as a potent inducer of white to opaque switching and functions primarily through the Ras1/cAMP pathway and phosphorylated Wor1, the gene product of the master switch locus. Our results therefore suggest that signals produced by bacterial co-members of the gastrointestinal tract microbiota regulate switching and therefore mating of C. albicans.

Show MeSH
Related in: MedlinePlus