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The fungal pathogen Candida albicans autoinduces hyphal morphogenesis by raising extracellular pH.

Vylkova S, Carman AJ, Danhof HA, Collette JR, Zhou H, Lorenz MC - MBio (2011)

Bottom Line: Under acidic conditions, this species can raise the pH from 4 to >7 in less than 12 h, resulting in autoinduction of the yeast-hyphal transition, a critical virulence trait.The pH changes are the result of the extrusion of ammonia, as observed in other fungi.This phenomenon is unprecedented in a human pathogen and may substantially impact host physiology by linking morphogenesis, pH adaptation, carbon metabolism, and interactions with host cells, all of which are critical for the ability of C. albicans to cause disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, The University of Texas Health Science Center, Houston, Texas, USA.

ABSTRACT

Unlabelled: pH homeostasis is critical for all organisms; in the fungal pathogen Candida albicans, pH adaptation is critical for virulence in distinct host niches. We demonstrate that beyond adaptation, C. albicans actively neutralizes the environment from either acidic or alkaline pHs. Under acidic conditions, this species can raise the pH from 4 to >7 in less than 12 h, resulting in autoinduction of the yeast-hyphal transition, a critical virulence trait. Extracellular alkalinization has been reported to occur in several fungal species, but under the specific conditions that we describe, the phenomenon is more rapid than previously observed. Alkalinization is linked to carbon deprivation, as it occurs in glucose-poor media and requires exogenous amino acids. These conditions are similar to those predicted to exist inside phagocytic cells, and we find a strong correlation between the use of amino acids as a cellular carbon source and the degree of alkalinization. Genetic and genomic approaches indicate an emphasis on amino acid uptake and catabolism in alkalinizing cells. Mutations in four genes, STP2, a transcription factor regulating amino acid permeases, ACH1 (acetyl-coenzyme A [acetyl-CoA] hydrolase), DUR1,2 (urea amidolyase), and ATO5, a putative ammonia transporter, abolish or delay neutralization. The pH changes are the result of the extrusion of ammonia, as observed in other fungi. We propose that nutrient-deprived C. albicans cells catabolize amino acids as a carbon source, excreting the amino nitrogen as ammonia to raise environmental pH and stimulate morphogenesis, thus directly contributing to pathogenesis.

Importance: Candida albicans is the most important fungal pathogen of humans, causing disease at multiple body sites. The ability to switch between multiple morphologies, including a rounded yeast cell and an elongated hyphal cell, is a key virulence trait in this species, as this reversible switch is thought to promote dissemination and tissue invasion in the host. We report here that C. albicans can actively alter the pH of its environment and induce its switch to the hyphal form. The change in pH is caused by the release of ammonia from the cells produced during the breakdown of amino acids. This phenomenon is unprecedented in a human pathogen and may substantially impact host physiology by linking morphogenesis, pH adaptation, carbon metabolism, and interactions with host cells, all of which are critical for the ability of C. albicans to cause disease.

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Related in: MedlinePlus

Ammonia extrusion by alkalinizing C. albicans colonies. (A) The wild-type strain, the stp2∆ mutant, and its complement were prepared as described in the legend to Fig. 4 and spotted to GM-BCP, pH 4.0 (plus 2% glucose where indicated). Plates were photographed at the indicated time points. At 4 days, a stereomicroscope was used to photograph the colony borders at ×20 magnification. Hyphal growth is indicated in the alkalinizing samples by the brush border at the colony edge. (B) Ammonia release from the same strains was measured as described in Materials and Methods at the indicated time points. A strong correlation between medium color (indicating alkalinization) and ammonia concentration was observed. The measurements in panel B are not from the same plates shown in panel A, as the arrangement of the acid trap to collect ammonia does not facilitate manipulation of the plates, but the time course in panel A is representative.
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f6: Ammonia extrusion by alkalinizing C. albicans colonies. (A) The wild-type strain, the stp2∆ mutant, and its complement were prepared as described in the legend to Fig. 4 and spotted to GM-BCP, pH 4.0 (plus 2% glucose where indicated). Plates were photographed at the indicated time points. At 4 days, a stereomicroscope was used to photograph the colony borders at ×20 magnification. Hyphal growth is indicated in the alkalinizing samples by the brush border at the colony edge. (B) Ammonia release from the same strains was measured as described in Materials and Methods at the indicated time points. A strong correlation between medium color (indicating alkalinization) and ammonia concentration was observed. The measurements in panel B are not from the same plates shown in panel A, as the arrangement of the acid trap to collect ammonia does not facilitate manipulation of the plates, but the time course in panel A is representative.

Mentions: Localized alkalinization in several fungal species has been associated with the release of volatile ammonia, a strong base, from the cells. To track the release of ammonia during this pH change, we grew strains on GM-BCP plates under alkalinizing and nonalkalinizing conditions (with or without 2% glucose) and placed an “acid trap” containing 10% citric acid below the cells in the beginning of this process (see Materials and Methods). Over 24 to 72 h, the pH rose (Fig. 6A), and we analyzed the acid trap for the presence of ammonium, released as volatile ammonia from the cells, via a colorimetric assay based on Nessler’s reagent, as previously described (48). Detectible ammonia increased significantly (Fig. 6B) over the 72-h period, correlating with the pH change observed on the plates (Fig. 6A). Very little volatile ammonia was released by nonalkalinizing cultures with the use of either the stp2∆ mutant or a glucose-repressed wild-type strain. We attempted to measure ammonia release from liquid cultures, aerated or not, and were unsuccessful using several approaches, possibly as a result of the volatile nature of this compound. On solid media, however, extrusion of ammonia correlates with the observed pH changes, providing a probable explanation for extracellular alkalinization, consistent with reports of ammonia release from Colletotrichum during its alkalinization in planta (23).


The fungal pathogen Candida albicans autoinduces hyphal morphogenesis by raising extracellular pH.

Vylkova S, Carman AJ, Danhof HA, Collette JR, Zhou H, Lorenz MC - MBio (2011)

Ammonia extrusion by alkalinizing C. albicans colonies. (A) The wild-type strain, the stp2∆ mutant, and its complement were prepared as described in the legend to Fig. 4 and spotted to GM-BCP, pH 4.0 (plus 2% glucose where indicated). Plates were photographed at the indicated time points. At 4 days, a stereomicroscope was used to photograph the colony borders at ×20 magnification. Hyphal growth is indicated in the alkalinizing samples by the brush border at the colony edge. (B) Ammonia release from the same strains was measured as described in Materials and Methods at the indicated time points. A strong correlation between medium color (indicating alkalinization) and ammonia concentration was observed. The measurements in panel B are not from the same plates shown in panel A, as the arrangement of the acid trap to collect ammonia does not facilitate manipulation of the plates, but the time course in panel A is representative.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101780&req=5

f6: Ammonia extrusion by alkalinizing C. albicans colonies. (A) The wild-type strain, the stp2∆ mutant, and its complement were prepared as described in the legend to Fig. 4 and spotted to GM-BCP, pH 4.0 (plus 2% glucose where indicated). Plates were photographed at the indicated time points. At 4 days, a stereomicroscope was used to photograph the colony borders at ×20 magnification. Hyphal growth is indicated in the alkalinizing samples by the brush border at the colony edge. (B) Ammonia release from the same strains was measured as described in Materials and Methods at the indicated time points. A strong correlation between medium color (indicating alkalinization) and ammonia concentration was observed. The measurements in panel B are not from the same plates shown in panel A, as the arrangement of the acid trap to collect ammonia does not facilitate manipulation of the plates, but the time course in panel A is representative.
Mentions: Localized alkalinization in several fungal species has been associated with the release of volatile ammonia, a strong base, from the cells. To track the release of ammonia during this pH change, we grew strains on GM-BCP plates under alkalinizing and nonalkalinizing conditions (with or without 2% glucose) and placed an “acid trap” containing 10% citric acid below the cells in the beginning of this process (see Materials and Methods). Over 24 to 72 h, the pH rose (Fig. 6A), and we analyzed the acid trap for the presence of ammonium, released as volatile ammonia from the cells, via a colorimetric assay based on Nessler’s reagent, as previously described (48). Detectible ammonia increased significantly (Fig. 6B) over the 72-h period, correlating with the pH change observed on the plates (Fig. 6A). Very little volatile ammonia was released by nonalkalinizing cultures with the use of either the stp2∆ mutant or a glucose-repressed wild-type strain. We attempted to measure ammonia release from liquid cultures, aerated or not, and were unsuccessful using several approaches, possibly as a result of the volatile nature of this compound. On solid media, however, extrusion of ammonia correlates with the observed pH changes, providing a probable explanation for extracellular alkalinization, consistent with reports of ammonia release from Colletotrichum during its alkalinization in planta (23).

Bottom Line: Under acidic conditions, this species can raise the pH from 4 to >7 in less than 12 h, resulting in autoinduction of the yeast-hyphal transition, a critical virulence trait.The pH changes are the result of the extrusion of ammonia, as observed in other fungi.This phenomenon is unprecedented in a human pathogen and may substantially impact host physiology by linking morphogenesis, pH adaptation, carbon metabolism, and interactions with host cells, all of which are critical for the ability of C. albicans to cause disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, The University of Texas Health Science Center, Houston, Texas, USA.

ABSTRACT

Unlabelled: pH homeostasis is critical for all organisms; in the fungal pathogen Candida albicans, pH adaptation is critical for virulence in distinct host niches. We demonstrate that beyond adaptation, C. albicans actively neutralizes the environment from either acidic or alkaline pHs. Under acidic conditions, this species can raise the pH from 4 to >7 in less than 12 h, resulting in autoinduction of the yeast-hyphal transition, a critical virulence trait. Extracellular alkalinization has been reported to occur in several fungal species, but under the specific conditions that we describe, the phenomenon is more rapid than previously observed. Alkalinization is linked to carbon deprivation, as it occurs in glucose-poor media and requires exogenous amino acids. These conditions are similar to those predicted to exist inside phagocytic cells, and we find a strong correlation between the use of amino acids as a cellular carbon source and the degree of alkalinization. Genetic and genomic approaches indicate an emphasis on amino acid uptake and catabolism in alkalinizing cells. Mutations in four genes, STP2, a transcription factor regulating amino acid permeases, ACH1 (acetyl-coenzyme A [acetyl-CoA] hydrolase), DUR1,2 (urea amidolyase), and ATO5, a putative ammonia transporter, abolish or delay neutralization. The pH changes are the result of the extrusion of ammonia, as observed in other fungi. We propose that nutrient-deprived C. albicans cells catabolize amino acids as a carbon source, excreting the amino nitrogen as ammonia to raise environmental pH and stimulate morphogenesis, thus directly contributing to pathogenesis.

Importance: Candida albicans is the most important fungal pathogen of humans, causing disease at multiple body sites. The ability to switch between multiple morphologies, including a rounded yeast cell and an elongated hyphal cell, is a key virulence trait in this species, as this reversible switch is thought to promote dissemination and tissue invasion in the host. We report here that C. albicans can actively alter the pH of its environment and induce its switch to the hyphal form. The change in pH is caused by the release of ammonia from the cells produced during the breakdown of amino acids. This phenomenon is unprecedented in a human pathogen and may substantially impact host physiology by linking morphogenesis, pH adaptation, carbon metabolism, and interactions with host cells, all of which are critical for the ability of C. albicans to cause disease.

Show MeSH
Related in: MedlinePlus