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The cellular target of histatin 5 on Candida albicans is the energized mitochondrion
E J Helmerhorst1, P Breeuwer, W van't Hof
1Academic Centre for Dentistry, Department of Oral Biochemistry, Vrije Universiteit, 1081 BT Amsterdam, The Netherlands. EJ.Helmerhorst.obc.acta@med.vu.nl
Abstract:
Histatin 5 is a human basic salivary peptide with strong fungicidal properties in vitro. To elucidate the mechanism of action, the effect of histatin 5 on the viability of Candida albicans cells was studied in relation to its membrane perturbing properties. It was found that both the killing activity and the membrane perturbing activity, studied by the influx of a DNA-specific marker propidium iodide, were inhibited by high salt conditions and by metabolic inhibitors, like sodium azide. In addition, exposure to histatin 5 resulted in a loss of the mitochondrial transmembrane potential in situ, measured by the release of the potential-dependent distributional probe rhodamine 123. Localization studies using tetramethylrhodamine isothiocyanate-labeled histatin 5 or fluorescein isothiocyanate-labeled histatin 5 showed a granular intracellular distribution of the peptide, which co-localized with mitotracker orange, a permeant mitochondria-specific probe. Like the biological effects, uptake of labeled histatin 5 was inhibited by mitochondrial inhibitors and high salt conditions. Our data indicate that histatin 5 is internalized, and targets to the energized mitochondrion.
Insights
Histatin 5, a salivary peptide, kills Candida albicans by entering cells and targeting mitochondria. Its fungicidal action depends on cell metabolism and is inhibited by high salt.
Area of Science:
- Biochemistry
- Microbiology
- Cell Biology
Background:
- Histatin 5 is a human salivary peptide with known fungicidal properties against Candida albicans in vitro.
- The precise mechanism of action for histatin 5's fungicidal activity remains incompletely understood.
Purpose of the Study:
- To investigate the mechanism of action of histatin 5 on Candida albicans viability.
- To determine the relationship between histatin 5's fungicidal and membrane-perturbing activities.
- To elucidate the cellular localization and target of histatin 5 within Candida albicans.
Main Methods:
- Studied the effect of histatin 5 on Candida albicans viability and membrane integrity using propidium iodide influx.
- Assessed mitochondrial transmembrane potential using rhodamine 123.
- Utilized labeled histatin 5 (TRITC or FITC) for intracellular localization studies, co-localizing with mitochondria-specific probes (Mitotracker Orange).
- Investigated the influence of high salt conditions and metabolic inhibitors (sodium azide) on histatin 5 activity and uptake.
Main Results:
- Histatin 5's fungicidal and membrane-perturbing activities were inhibited by high salt concentrations and metabolic inhibitors.
- Exposure to histatin 5 led to a loss of mitochondrial transmembrane potential in Candida albicans cells.
- Labeled histatin 5 showed granular intracellular distribution, co-localizing with mitochondria.
- Uptake of labeled histatin 5 was also inhibited by mitochondrial inhibitors and high salt conditions.
Conclusions:
- Histatin 5 is internalized by Candida albicans cells.
- The energized mitochondrion is a primary target for histatin 5.
- The fungicidal mechanism of histatin 5 involves mitochondrial disruption, dependent on cellular metabolic activity and salt concentration.