Fluorescent TAPY-bodipy dyads as tools for imaging fungal mitochondria by confocal microscopy and flow cytometry

Area of Science:

• Cell Biology
• Mycology
• Biochemistry

Background:

• Mitochondria-targeted fluorescent probes are crucial for cell biology research.
• Current probes are primarily optimized for mammalian cells, limiting fungal studies.
• Pathogenic yeasts like Candida albicans have increasing clinical significance.

Purpose of the Study:

• To evaluate triarylpyridinium-bodipy (TAPY-BDP) conjugates as mitochondrial markers in fungi.
• To compare TAPY-BDP probes with existing triphenylphosphonium-bodipy (TPP-BDP) and non-targeted probes.
• To assess the efficacy of TAPY-BDP probes in Candida albicans and Pichia kudriavzevii.

Main Methods:

• Confocal microscopy and flow cytometry were used to quantify probe uptake and localization.
• Colocalization studies with MitoTracker™ Deep Red assessed targeting efficiency.
• The effect of carbonyl cyanide 3-chlorophenylhydrazone (CCCP) confirmed mitochondrial membrane potential-dependent accumulation.

Main Results:

• Most TAPY-BDP derivatives demonstrated efficient mitochondrial targeting with high colocalization coefficients (up to 0.9).
• TAPY-BDPs with specific substituents (R=H, Me, OMe, NMe2) showed superior mitochondrial fluorescence compared to lipophilic analogues and TPP-BDP.
• TAPY(OMe)-BDP maintained strong staining at lower concentrations (100 nM) where TPP-BDP signal diminished.

Conclusions:

• Triarylpyridinium (TAPY) serves as a viable alternative to triphenylphosphonium (TPP) for mitochondrial targeting in fungi.
• These findings expand the toolkit for bioanalytical studies in azole-resistant yeasts.
• The developed probes may facilitate mitochondria-directed therapeutic delivery strategies.