Ctt1 catalase activity potentiates antifungal azoles in the emerging opportunistic pathogen Saccharomyces cerevisiae

Dorival Martins,Dao Nguyen,Ann M English

doi:10.1038/s41598-019-45070-w

Dorival Martins, Dao Nguyen + Show 1 more

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https://doi.org/10.1038/s41598-019-45070-w

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Abstract

Fungi respond to antifungal drugs by increasing their antioxidant stress response. How this impacts antifungal efficacy remains controversial and not well understood. Here we examine the role of catalase activity in the resistance of Saccharomyces cerevisiae to the common antifungals, fluconazole and miconazole, for which we report minimum inhibitory concentrations (MICs) of 104 and 19 μM, respectively. At sub-MIC concentrations, fluconazole and miconazole stimulate catalase activity 2-3-fold but, unexpectedly, deletion of cytosolic catalase (ctt1) makes cells more resistant to these azoles and to clotrimazole, itraconazole and posaconazole. On the other hand, upregulating Ctt1 activity by preconditioning with 0.2 mM H2O2 potentiates miconazole 32-fold and fluconazole 4-fold. Since H2O2 preconditioning does not alter the resistance of ctt1Δ cells, which possess negligible catalase activity, we link azole potentiation with Ctt1 upregulation. In contrast, sod2Δ cells deleted for mitochondrial superoxide dismutase are 4–8-fold more azole sensitive than wild-type cells, revealing that Sod2 activity protects cells against azole toxicity. In fact, the ctt1Δ mutant has double the Sod2 activity of wild-type cells so ctt1 deletion increases azole resistance in part by Sod2 upregulation. Notably, deletion of peroxisomal/mitochondrial cta1 or cytosolic sod1 does not alter fluconazole or miconazole potency.

Highlights

Antimicrobial challenge appears to induce the rewiring of microbial metabolic networks and stress-response pathways regardless of the primary drug-target interaction[1,2]
C. albicans possesses a single peroxisomal/mitochondrial catalase (Cta1)[29] together with six superoxide dismutases (Sods)[13] while S. cerevisiae produces cytosolic Ctt[1] in addition to Cta[1] but just two Sods, cytosolic CuZnSod[1], which localizes to the mitochondrial intermembrane space[30,31], and mitochondrial MnSod[232]
An azole is classified as fungicidal if 1xMIC or 2xMIC promotes a ≥103-fold reduction in the viable cfu/mL and Table S2 shows that the imidazoles are fungicidal under the present experimental conditions, whereas the triazoles are fungistatic with the exception of voriconazole

Summary

Introduction

Antimicrobial challenge appears to induce the rewiring of microbial metabolic networks and stress-response pathways regardless of the primary drug-target interaction[1,2]. Deletion of respiratory enzymes or inhibition of cellular respiration, a major source of reactive oxygen species (ROS)[1,2,18], decreases antimicrobial lethality. Combined, these observations are consistent with the belief that cidal antibiotics[1,19] and antifungals[2,20,21] increase ROS levels. The ΔrelA ΔspoT mutant of Pseudomonas aeruginosa, which is deficient in the (p)ppGpp alarmone, exhibits depressed catalase and superoxide dismutase (Sod) activities and is hypersensitive to antibiotics[3,4,22]. A comparison of how deletion of specific antioxidant enzymes alters antifungal potency in these well-characterized yeasts provides an excellent opportunity to gain new insights into pathogen survival strategies and the evolution of antifungal resilience

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