Limits...
The Candida albicans Histone Acetyltransferase Hat1 Regulates Stress Resistance and Virulence via Distinct Chromatin Assembly Pathways.

Tscherner M, Zwolanek F, Je S, Sedlazeck FJ, Petryshyn A, Frohner IE, Mavrianos J, Chauhan N, von Haeseler A, Kuchler K - PLoS Pathog. (2015)

Bottom Line: Hydrogen peroxide resistance in cells lacking Hat1 results from higher induction rates of oxidative stress gene expression, accompanied by reduced histone density as well as subsequent increased RNA polymerase recruitment.Remarkably, the oxidative stress phenotype of hat1Δ/Δ cells is a species-specific trait only found in C. albicans and members of the CTG clade.The reduced azole susceptibility appears to be conserved in a wider range of fungi.

View Article: PubMed Central - PubMed

Affiliation: Department for Medical Biochemistry, Medical University of Vienna, Max F. Perutz Laboratories, Campus Vienna Biocenter, Vienna, Austria.

ABSTRACT
Human fungal pathogens like Candida albicans respond to host immune surveillance by rapidly adapting their transcriptional programs. Chromatin assembly factors are involved in the regulation of stress genes by modulating the histone density at these loci. Here, we report a novel role for the chromatin assembly-associated histone acetyltransferase complex NuB4 in regulating oxidative stress resistance, antifungal drug tolerance and virulence in C. albicans. Strikingly, depletion of the NuB4 catalytic subunit, the histone acetyltransferase Hat1, markedly increases resistance to oxidative stress and tolerance to azole antifungals. Hydrogen peroxide resistance in cells lacking Hat1 results from higher induction rates of oxidative stress gene expression, accompanied by reduced histone density as well as subsequent increased RNA polymerase recruitment. Furthermore, hat1Δ/Δ cells, despite showing growth defects in vitro, display reduced susceptibility to reactive oxygen-mediated killing by innate immune cells. Thus, clearance from infected mice is delayed although cells lacking Hat1 are severely compromised in killing the host. Interestingly, increased oxidative stress resistance and azole tolerance are phenocopied by the loss of histone chaperone complexes CAF-1 and HIR, respectively, suggesting a central role for NuB4 in the delivery of histones destined for chromatin assembly via distinct pathways. Remarkably, the oxidative stress phenotype of hat1Δ/Δ cells is a species-specific trait only found in C. albicans and members of the CTG clade. The reduced azole susceptibility appears to be conserved in a wider range of fungi. Thus, our work demonstrates how highly conserved chromatin assembly pathways can acquire new functions in pathogenic fungi during coevolution with the host.

No MeSH data available.


Loss of Hat1 raises antioxidant enzyme activity and glutathione-mediated H2O2 resistance.(A) Faster CAT1 induction increases catalase activity in hat1Δ/Δ cells. Catalase activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from three independent experiments. (B) Loss of Hat1 leads to increased glutathione peroxidase activity. GPx activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from two independent experiments. (C) Lack of CAT1 does not abolish Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with 1 mM H2O2 for 2 hours, plated and colonies counted after 3 days of incubation on YPD plates at 30°C to determine viability. Data are shown as mean + SD from three independent experiments. (D) Depletion of glutathione biosynthesis abolishes Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with H2O2 for 2 hours and plated on YPD plates containing glutathione. Colonies were counted to determine viability after growth for 3 days at 30°C. Data are shown as mean + SD from three independent experiments. (A-D) n.s.: not significant, *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding control (Student's t-test).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4608838&req=5

ppat.1005218.g007: Loss of Hat1 raises antioxidant enzyme activity and glutathione-mediated H2O2 resistance.(A) Faster CAT1 induction increases catalase activity in hat1Δ/Δ cells. Catalase activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from three independent experiments. (B) Loss of Hat1 leads to increased glutathione peroxidase activity. GPx activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from two independent experiments. (C) Lack of CAT1 does not abolish Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with 1 mM H2O2 for 2 hours, plated and colonies counted after 3 days of incubation on YPD plates at 30°C to determine viability. Data are shown as mean + SD from three independent experiments. (D) Depletion of glutathione biosynthesis abolishes Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with H2O2 for 2 hours and plated on YPD plates containing glutathione. Colonies were counted to determine viability after growth for 3 days at 30°C. Data are shown as mean + SD from three independent experiments. (A-D) n.s.: not significant, *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding control (Student's t-test).

Mentions: Furthermore, we tested whether the increase in transcription of oxidative stress genes in the hat1Δ/Δ mutant is paralleled by elevated activity of the corresponding enzymes. Therefore, we prepared whole cell extracts from cells before and after 60 min exposure to H2O2 and determined catalase activity as well as GPx activity spectrophotometrically. We detected elevated catalase activity in the hat1Δ/Δ mutant already under non-inducing conditions, albeit as expected at a low level (Fig 7A). However, H2O2 treatment of hat1Δ/Δ cells dramatically increased catalase activity when compared to the wild-type, indicating that the mutant is more efficient in degrading hydrogen peroxide (Fig 7A). Likewise, the GPx activity assay detected some increase already under non-inducing conditions (Fig 7B), but H2O2 treatment of the hat1Δ/Δ mutant significantly increased GPx activity (Fig 7B). These data demonstrate that the increased induction rate of oxidative stress genes in the hat1Δ/Δ mutant increases the activities of the enzymes encoded by the target genes.


The Candida albicans Histone Acetyltransferase Hat1 Regulates Stress Resistance and Virulence via Distinct Chromatin Assembly Pathways.

Tscherner M, Zwolanek F, Je S, Sedlazeck FJ, Petryshyn A, Frohner IE, Mavrianos J, Chauhan N, von Haeseler A, Kuchler K - PLoS Pathog. (2015)

Loss of Hat1 raises antioxidant enzyme activity and glutathione-mediated H2O2 resistance.(A) Faster CAT1 induction increases catalase activity in hat1Δ/Δ cells. Catalase activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from three independent experiments. (B) Loss of Hat1 leads to increased glutathione peroxidase activity. GPx activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from two independent experiments. (C) Lack of CAT1 does not abolish Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with 1 mM H2O2 for 2 hours, plated and colonies counted after 3 days of incubation on YPD plates at 30°C to determine viability. Data are shown as mean + SD from three independent experiments. (D) Depletion of glutathione biosynthesis abolishes Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with H2O2 for 2 hours and plated on YPD plates containing glutathione. Colonies were counted to determine viability after growth for 3 days at 30°C. Data are shown as mean + SD from three independent experiments. (A-D) n.s.: not significant, *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding control (Student's t-test).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608838&req=5

ppat.1005218.g007: Loss of Hat1 raises antioxidant enzyme activity and glutathione-mediated H2O2 resistance.(A) Faster CAT1 induction increases catalase activity in hat1Δ/Δ cells. Catalase activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from three independent experiments. (B) Loss of Hat1 leads to increased glutathione peroxidase activity. GPx activity was determined in whole cell extracts isolated from cells before and after H2O2 treatment. Data are shown as mean + SD from two independent experiments. (C) Lack of CAT1 does not abolish Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with 1 mM H2O2 for 2 hours, plated and colonies counted after 3 days of incubation on YPD plates at 30°C to determine viability. Data are shown as mean + SD from three independent experiments. (D) Depletion of glutathione biosynthesis abolishes Hat1-mediated H2O2 resistance. Cells of the indicated strains were treated with H2O2 for 2 hours and plated on YPD plates containing glutathione. Colonies were counted to determine viability after growth for 3 days at 30°C. Data are shown as mean + SD from three independent experiments. (A-D) n.s.: not significant, *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding control (Student's t-test).
Mentions: Furthermore, we tested whether the increase in transcription of oxidative stress genes in the hat1Δ/Δ mutant is paralleled by elevated activity of the corresponding enzymes. Therefore, we prepared whole cell extracts from cells before and after 60 min exposure to H2O2 and determined catalase activity as well as GPx activity spectrophotometrically. We detected elevated catalase activity in the hat1Δ/Δ mutant already under non-inducing conditions, albeit as expected at a low level (Fig 7A). However, H2O2 treatment of hat1Δ/Δ cells dramatically increased catalase activity when compared to the wild-type, indicating that the mutant is more efficient in degrading hydrogen peroxide (Fig 7A). Likewise, the GPx activity assay detected some increase already under non-inducing conditions (Fig 7B), but H2O2 treatment of the hat1Δ/Δ mutant significantly increased GPx activity (Fig 7B). These data demonstrate that the increased induction rate of oxidative stress genes in the hat1Δ/Δ mutant increases the activities of the enzymes encoded by the target genes.

Bottom Line: Hydrogen peroxide resistance in cells lacking Hat1 results from higher induction rates of oxidative stress gene expression, accompanied by reduced histone density as well as subsequent increased RNA polymerase recruitment.Remarkably, the oxidative stress phenotype of hat1Δ/Δ cells is a species-specific trait only found in C. albicans and members of the CTG clade.The reduced azole susceptibility appears to be conserved in a wider range of fungi.

View Article: PubMed Central - PubMed

Affiliation: Department for Medical Biochemistry, Medical University of Vienna, Max F. Perutz Laboratories, Campus Vienna Biocenter, Vienna, Austria.

ABSTRACT
Human fungal pathogens like Candida albicans respond to host immune surveillance by rapidly adapting their transcriptional programs. Chromatin assembly factors are involved in the regulation of stress genes by modulating the histone density at these loci. Here, we report a novel role for the chromatin assembly-associated histone acetyltransferase complex NuB4 in regulating oxidative stress resistance, antifungal drug tolerance and virulence in C. albicans. Strikingly, depletion of the NuB4 catalytic subunit, the histone acetyltransferase Hat1, markedly increases resistance to oxidative stress and tolerance to azole antifungals. Hydrogen peroxide resistance in cells lacking Hat1 results from higher induction rates of oxidative stress gene expression, accompanied by reduced histone density as well as subsequent increased RNA polymerase recruitment. Furthermore, hat1Δ/Δ cells, despite showing growth defects in vitro, display reduced susceptibility to reactive oxygen-mediated killing by innate immune cells. Thus, clearance from infected mice is delayed although cells lacking Hat1 are severely compromised in killing the host. Interestingly, increased oxidative stress resistance and azole tolerance are phenocopied by the loss of histone chaperone complexes CAF-1 and HIR, respectively, suggesting a central role for NuB4 in the delivery of histones destined for chromatin assembly via distinct pathways. Remarkably, the oxidative stress phenotype of hat1Δ/Δ cells is a species-specific trait only found in C. albicans and members of the CTG clade. The reduced azole susceptibility appears to be conserved in a wider range of fungi. Thus, our work demonstrates how highly conserved chromatin assembly pathways can acquire new functions in pathogenic fungi during coevolution with the host.

No MeSH data available.