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Published on: June 13, 2021
Endosomal Trafficking Defects Can Induce Calcium-Dependent Azole Tolerance in Candida albicans
Arturo Luna-Tapia1, Hélène Tournu1, Tracy L Peters1
1Department of Clinical Pharmacy, Division of Clinical and Experimental Therapeutics, College of Pharmacy, University of Tennessee Health Sciences Center, Memphis, Tennessee, USA.
Abstract:
The azole antifungals arrest fungal growth through inhibition of ergosterol biosynthesis. We recently reported that a Candida albicans vps21Δ/Δ mutant, deficient in membrane trafficking through the late endosome/prevacuolar compartment (PVC), continues to grow in the presence of the azoles despite the depletion of cellular ergosterol. Here, we report that the vps21Δ/Δ mutant exhibits less plasma membrane damage upon azole treatment than the wild type, as measured by the release of a cytoplasmic luciferase reporter into the culture supernatant. Our results also reveal that the vps21Δ/Δ mutant has abnormal levels of intracellular Ca2+ and, in the presence of fluconazole, enhanced expression of a calcineurin-responsive RTA2-GFP reporter. Furthermore, the azole tolerance phenotype of the vps21Δ/Δ mutant is dependent upon both extracellular calcium levels and calcineurin activity. These findings underscore the importance of endosomal trafficking in determining the cellular consequences of azole treatment and indicate that this may occur through modulation of calcium- and calcineurin-dependent responses.
Insights
Candida albicans mutants lacking endosomal trafficking survive azole antifungal drugs. This tolerance is linked to calcium levels and calcineurin activity, revealing new drug targets.
Area of Science:
- Mycology
- Cell Biology
- Biochemistry
Background:
- Azole antifungals inhibit ergosterol biosynthesis, arresting fungal growth.
- A Candida albicans mutant (vps21Δ/Δ) deficient in endosomal trafficking tolerates azoles despite ergosterol depletion.
Purpose of the Study:
- Investigate the mechanisms behind azole tolerance in the vps21Δ/Δ mutant.
- Determine the role of endosomal trafficking in cellular responses to azole antifungals.
Main Methods:
- Assessing plasma membrane integrity using a luciferase reporter.
- Measuring intracellular calcium levels.
- Analyzing calcineurin-dependent gene expression (RTA2-GFP reporter).
- Evaluating the impact of extracellular calcium and calcineurin inhibition on azole tolerance.
Main Results:
- The vps21Δ/Δ mutant shows reduced plasma membrane damage upon azole treatment.
- This mutant exhibits altered intracellular calcium homeostasis.
- Fluconazole treatment enhances calcineurin-responsive reporter expression in the mutant.
- Azole tolerance in the vps21Δ/Δ mutant is dependent on extracellular calcium and calcineurin activity.
Conclusions:
- Endosomal trafficking is crucial for mediating cellular responses to azole antifungals.
- The vps21Δ/Δ mutant's azole tolerance involves calcium and calcineurin signaling pathways.
- Targeting these pathways may offer novel strategies for antifungal therapy.
