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Flavodoxin-Like Proteins Protect Candida albicans from Oxidative Stress and Promote Virulence.

Li L, Naseem S, Sharma S, Konopka JB - PLoS Pathog. (2015)

Bottom Line: Quinone reductase activity confers important antioxidant effects because resistance to oxidation was restored in the quadruple mutant by expressing either Escherichia coli wrbA or mammalian NQO1, two distinct types of quinone reductases.These observations suggested that FLPs reduce ubiquinone (coenzyme Q), enabling it to serve as an antioxidant in the membrane.The potential of FLPs as novel targets for antifungal therapy is further underscored by their absence in mammalian cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, United States of America.

ABSTRACT
The fungal pathogen Candida albicans causes lethal systemic infections in humans. To better define how pathogens resist oxidative attack by the immune system, we examined a family of four Flavodoxin-Like Proteins (FLPs) in C. albicans. In agreement with previous studies showing that FLPs in bacteria and plants act as NAD(P)H quinone oxidoreductases, a C. albicans quadruple mutant lacking all four FLPs (pst1Δ, pst2Δ, pst3Δ, ycp4Δ) was more sensitive to benzoquinone. Interestingly, the quadruple mutant was also more sensitive to a variety of oxidants. Quinone reductase activity confers important antioxidant effects because resistance to oxidation was restored in the quadruple mutant by expressing either Escherichia coli wrbA or mammalian NQO1, two distinct types of quinone reductases. FLPs were detected at the plasma membrane in C. albicans, and the quadruple mutant was more sensitive to linolenic acid, a polyunsaturated fatty acid that can auto-oxidize and promote lipid peroxidation. These observations suggested that FLPs reduce ubiquinone (coenzyme Q), enabling it to serve as an antioxidant in the membrane. In support of this, a C. albicans coq3Δ mutant that fails to synthesize ubiquinone was also highly sensitive to oxidative stress. FLPs are critical for survival in the host, as the quadruple mutant was avirulent in a mouse model of systemic candidiasis under conditions where infection with wild type C. albicans was lethal. The quadruple mutant cells initially grew well in kidneys, the major site of C. albicans growth in mice, but then declined after the influx of neutrophils and by day 4 post-infection 33% of the mice cleared the infection. Thus, FLPs and ubiquinone are important new antioxidant mechanisms that are critical for fungal virulence. The potential of FLPs as novel targets for antifungal therapy is further underscored by their absence in mammalian cells.

No MeSH data available.


Related in: MedlinePlus

Plasma membrane localization of the C. albicans FLP proteins.Fluorescence microscopy of C. albicans cells producing (A) Pst1-GFP and Pst3-GFP fusion proteins in which GFP was added to the C terminus. (B) Cells producing GFP-Pst2 and GFP-Ycp4 fusions in which GFP was added to the N terminus. The GFP fusion genes in panel A were regulated by their endogenous promoters, whereas the fusion genes in panel B were expressed using the ADH1 promoter.
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ppat.1005147.g007: Plasma membrane localization of the C. albicans FLP proteins.Fluorescence microscopy of C. albicans cells producing (A) Pst1-GFP and Pst3-GFP fusion proteins in which GFP was added to the C terminus. (B) Cells producing GFP-Pst2 and GFP-Ycp4 fusions in which GFP was added to the N terminus. The GFP fusion genes in panel A were regulated by their endogenous promoters, whereas the fusion genes in panel B were expressed using the ADH1 promoter.

Mentions: FLPs were fused to GFP to examine their subcellular localization. Pst1-GFP and Pst3-GFP were detected at the plasma membrane by fluorescence microscopy (Fig 7A). To improve detection for the other two FLPs, the strong ADH1 promoter was used to express GFP fusions to the PST2 and YCP4 genes. These GFP-Pst2 and GFP-Ycp4 fusion proteins gave a strong plasma membrane signal (Fig 7B). The GFP-tagged FLPs all showed a slightly patchy distribution in the plasma membrane, suggesting that they localize in part to the eisosome subdomains, as do their S. cerevisiae orthologs [18, 42]. Cytoplasmic GFP signal was also detected in cells. However, this could be due to proteolytic cleavage of the FLP proteins resulting in the presence of free cytoplasmic GFP, as Western blot analysis detected a strong signal at the expected size of GFP (~30 kD) (S3 Fig).


Flavodoxin-Like Proteins Protect Candida albicans from Oxidative Stress and Promote Virulence.

Li L, Naseem S, Sharma S, Konopka JB - PLoS Pathog. (2015)

Plasma membrane localization of the C. albicans FLP proteins.Fluorescence microscopy of C. albicans cells producing (A) Pst1-GFP and Pst3-GFP fusion proteins in which GFP was added to the C terminus. (B) Cells producing GFP-Pst2 and GFP-Ycp4 fusions in which GFP was added to the N terminus. The GFP fusion genes in panel A were regulated by their endogenous promoters, whereas the fusion genes in panel B were expressed using the ADH1 promoter.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556627&req=5

ppat.1005147.g007: Plasma membrane localization of the C. albicans FLP proteins.Fluorescence microscopy of C. albicans cells producing (A) Pst1-GFP and Pst3-GFP fusion proteins in which GFP was added to the C terminus. (B) Cells producing GFP-Pst2 and GFP-Ycp4 fusions in which GFP was added to the N terminus. The GFP fusion genes in panel A were regulated by their endogenous promoters, whereas the fusion genes in panel B were expressed using the ADH1 promoter.
Mentions: FLPs were fused to GFP to examine their subcellular localization. Pst1-GFP and Pst3-GFP were detected at the plasma membrane by fluorescence microscopy (Fig 7A). To improve detection for the other two FLPs, the strong ADH1 promoter was used to express GFP fusions to the PST2 and YCP4 genes. These GFP-Pst2 and GFP-Ycp4 fusion proteins gave a strong plasma membrane signal (Fig 7B). The GFP-tagged FLPs all showed a slightly patchy distribution in the plasma membrane, suggesting that they localize in part to the eisosome subdomains, as do their S. cerevisiae orthologs [18, 42]. Cytoplasmic GFP signal was also detected in cells. However, this could be due to proteolytic cleavage of the FLP proteins resulting in the presence of free cytoplasmic GFP, as Western blot analysis detected a strong signal at the expected size of GFP (~30 kD) (S3 Fig).

Bottom Line: Quinone reductase activity confers important antioxidant effects because resistance to oxidation was restored in the quadruple mutant by expressing either Escherichia coli wrbA or mammalian NQO1, two distinct types of quinone reductases.These observations suggested that FLPs reduce ubiquinone (coenzyme Q), enabling it to serve as an antioxidant in the membrane.The potential of FLPs as novel targets for antifungal therapy is further underscored by their absence in mammalian cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, United States of America.

ABSTRACT
The fungal pathogen Candida albicans causes lethal systemic infections in humans. To better define how pathogens resist oxidative attack by the immune system, we examined a family of four Flavodoxin-Like Proteins (FLPs) in C. albicans. In agreement with previous studies showing that FLPs in bacteria and plants act as NAD(P)H quinone oxidoreductases, a C. albicans quadruple mutant lacking all four FLPs (pst1Δ, pst2Δ, pst3Δ, ycp4Δ) was more sensitive to benzoquinone. Interestingly, the quadruple mutant was also more sensitive to a variety of oxidants. Quinone reductase activity confers important antioxidant effects because resistance to oxidation was restored in the quadruple mutant by expressing either Escherichia coli wrbA or mammalian NQO1, two distinct types of quinone reductases. FLPs were detected at the plasma membrane in C. albicans, and the quadruple mutant was more sensitive to linolenic acid, a polyunsaturated fatty acid that can auto-oxidize and promote lipid peroxidation. These observations suggested that FLPs reduce ubiquinone (coenzyme Q), enabling it to serve as an antioxidant in the membrane. In support of this, a C. albicans coq3Δ mutant that fails to synthesize ubiquinone was also highly sensitive to oxidative stress. FLPs are critical for survival in the host, as the quadruple mutant was avirulent in a mouse model of systemic candidiasis under conditions where infection with wild type C. albicans was lethal. The quadruple mutant cells initially grew well in kidneys, the major site of C. albicans growth in mice, but then declined after the influx of neutrophils and by day 4 post-infection 33% of the mice cleared the infection. Thus, FLPs and ubiquinone are important new antioxidant mechanisms that are critical for fungal virulence. The potential of FLPs as novel targets for antifungal therapy is further underscored by their absence in mammalian cells.

No MeSH data available.


Related in: MedlinePlus