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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.


Related in: MedlinePlus

Lack of histone chaperones mimics deletion of HAT1.(A) Loss of Cac2 increases H2O2 resistance. Deletion of RTT106 or HIR1 does not affect susceptibility to hydrogen peroxide. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 3 days. (B) Deletion of HAT1 or CAC2 increases survival to transient hydrogen peroxide treatment. Exponentially growing cells were treated with the indicated concentrations of H2O2 for 2 hours. Cells were 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. (C) Deletion of HIR1 reduces voriconazole (Voric.) susceptibility. The hat1hir1Δ/Δ double deletion strain mimics lack of Hat1. Loss of Cac2 has only a minor effect and deletion of RTT106 does not alter azole susceptibility. Experiment was performed as described in (A). (D) Increased azole tolerance of hat1Δ/Δ, hir1Δ/Δ and hat1hir1Δ/Δ was confirmed using a liquid growth inhibition assay. Logarithmically growing cells were diluted into medium containing the indicated concentrations of voriconazole (Voric.) and incubated at 30°C for 18 hours. OD600 was determined and growth inhibition relative to untreated samples was calculated. Data are shown as mean + SD from three independent experiments. (E) Lack of Spt6 reduces H2O2 susceptibility. Experiment was performed as described in (B). Cells were treated with 10 mM H2O2. Data are shown as mean + SD from two independent experiments. (F) Deletion of SPT6 increases H2O2 resistance and azole tolerance. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 5 days. (G) Reduction of histone gene dosage decreases H2O2 and azole susceptibility. Experiment was performed as described in (A). (B, D, E) *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding wild-type (Student's t-test).
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ppat.1005218.g002: Lack of histone chaperones mimics deletion of HAT1.(A) Loss of Cac2 increases H2O2 resistance. Deletion of RTT106 or HIR1 does not affect susceptibility to hydrogen peroxide. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 3 days. (B) Deletion of HAT1 or CAC2 increases survival to transient hydrogen peroxide treatment. Exponentially growing cells were treated with the indicated concentrations of H2O2 for 2 hours. Cells were 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. (C) Deletion of HIR1 reduces voriconazole (Voric.) susceptibility. The hat1hir1Δ/Δ double deletion strain mimics lack of Hat1. Loss of Cac2 has only a minor effect and deletion of RTT106 does not alter azole susceptibility. Experiment was performed as described in (A). (D) Increased azole tolerance of hat1Δ/Δ, hir1Δ/Δ and hat1hir1Δ/Δ was confirmed using a liquid growth inhibition assay. Logarithmically growing cells were diluted into medium containing the indicated concentrations of voriconazole (Voric.) and incubated at 30°C for 18 hours. OD600 was determined and growth inhibition relative to untreated samples was calculated. Data are shown as mean + SD from three independent experiments. (E) Lack of Spt6 reduces H2O2 susceptibility. Experiment was performed as described in (B). Cells were treated with 10 mM H2O2. Data are shown as mean + SD from two independent experiments. (F) Deletion of SPT6 increases H2O2 resistance and azole tolerance. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 5 days. (G) Reduction of histone gene dosage decreases H2O2 and azole susceptibility. Experiment was performed as described in (A). (B, D, E) *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding wild-type (Student's t-test).

Mentions: Strikingly, lack of CAC2, a subunit of the CAF-1 histone chaperone complex, also strongly increased resistance to H2O2 and tBOOH (Fig 2A and S1D Fig). Furthermore, a quantification of H2O2 resistance by determination of the survival rate in liquid culture confirmed this result (Fig 2B). Interestingly however, spot dilution assays suggested a minor effect on azole susceptibility of cac2Δ/Δ cells (Fig 2C). By contrast, deletion of HIR1, a component of the HIR histone chaperone complex, dramatically increased tolerance to voriconazole, but did not alter H2O2 susceptibility (Fig 2A and 2C). Importantly, HAT1 and HIR1 are epistatic, since a double deletion strain failed to show increased azole resistance when compared to the corresponding single deletions based on spot dilution assays and growth inhibition in liquid culture (Fig 2C and 2D).


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)

Lack of histone chaperones mimics deletion of HAT1.(A) Loss of Cac2 increases H2O2 resistance. Deletion of RTT106 or HIR1 does not affect susceptibility to hydrogen peroxide. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 3 days. (B) Deletion of HAT1 or CAC2 increases survival to transient hydrogen peroxide treatment. Exponentially growing cells were treated with the indicated concentrations of H2O2 for 2 hours. Cells were 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. (C) Deletion of HIR1 reduces voriconazole (Voric.) susceptibility. The hat1hir1Δ/Δ double deletion strain mimics lack of Hat1. Loss of Cac2 has only a minor effect and deletion of RTT106 does not alter azole susceptibility. Experiment was performed as described in (A). (D) Increased azole tolerance of hat1Δ/Δ, hir1Δ/Δ and hat1hir1Δ/Δ was confirmed using a liquid growth inhibition assay. Logarithmically growing cells were diluted into medium containing the indicated concentrations of voriconazole (Voric.) and incubated at 30°C for 18 hours. OD600 was determined and growth inhibition relative to untreated samples was calculated. Data are shown as mean + SD from three independent experiments. (E) Lack of Spt6 reduces H2O2 susceptibility. Experiment was performed as described in (B). Cells were treated with 10 mM H2O2. Data are shown as mean + SD from two independent experiments. (F) Deletion of SPT6 increases H2O2 resistance and azole tolerance. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 5 days. (G) Reduction of histone gene dosage decreases H2O2 and azole susceptibility. Experiment was performed as described in (A). (B, D, E) *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding wild-type (Student's t-test).
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ppat.1005218.g002: Lack of histone chaperones mimics deletion of HAT1.(A) Loss of Cac2 increases H2O2 resistance. Deletion of RTT106 or HIR1 does not affect susceptibility to hydrogen peroxide. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 3 days. (B) Deletion of HAT1 or CAC2 increases survival to transient hydrogen peroxide treatment. Exponentially growing cells were treated with the indicated concentrations of H2O2 for 2 hours. Cells were 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. (C) Deletion of HIR1 reduces voriconazole (Voric.) susceptibility. The hat1hir1Δ/Δ double deletion strain mimics lack of Hat1. Loss of Cac2 has only a minor effect and deletion of RTT106 does not alter azole susceptibility. Experiment was performed as described in (A). (D) Increased azole tolerance of hat1Δ/Δ, hir1Δ/Δ and hat1hir1Δ/Δ was confirmed using a liquid growth inhibition assay. Logarithmically growing cells were diluted into medium containing the indicated concentrations of voriconazole (Voric.) and incubated at 30°C for 18 hours. OD600 was determined and growth inhibition relative to untreated samples was calculated. Data are shown as mean + SD from three independent experiments. (E) Lack of Spt6 reduces H2O2 susceptibility. Experiment was performed as described in (B). Cells were treated with 10 mM H2O2. Data are shown as mean + SD from two independent experiments. (F) Deletion of SPT6 increases H2O2 resistance and azole tolerance. Fivefold serial dilutions of the indicated strains were spotted on agar plates containing the indicated substances and pictures were taken after incubation at 30°C for 5 days. (G) Reduction of histone gene dosage decreases H2O2 and azole susceptibility. Experiment was performed as described in (A). (B, D, E) *P<0.05, **P<0.01 and ***P<0.001 relative to the corresponding wild-type (Student's t-test).
Mentions: Strikingly, lack of CAC2, a subunit of the CAF-1 histone chaperone complex, also strongly increased resistance to H2O2 and tBOOH (Fig 2A and S1D Fig). Furthermore, a quantification of H2O2 resistance by determination of the survival rate in liquid culture confirmed this result (Fig 2B). Interestingly however, spot dilution assays suggested a minor effect on azole susceptibility of cac2Δ/Δ cells (Fig 2C). By contrast, deletion of HIR1, a component of the HIR histone chaperone complex, dramatically increased tolerance to voriconazole, but did not alter H2O2 susceptibility (Fig 2A and 2C). Importantly, HAT1 and HIR1 are epistatic, since a double deletion strain failed to show increased azole resistance when compared to the corresponding single deletions based on spot dilution assays and growth inhibition in liquid culture (Fig 2C and 2D).

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.


Related in: MedlinePlus