Dynamic calcium-mediated stress response and recovery signatures in the fungal pathogen, Candida albicans

Calcium (Ca²⁺) is an important second messenger for activating stress response signaling and cell adaptation in eukaryotic cells yet intracellular Ca²⁺-dynamics in fungi are poorly understood due to lack of effective real-time Ca²⁺ reporters. We engineered the GCaMP6f construct for use in the fungal pathogen, Candida albicans, and used live-cell imaging to observe both dynamic Ca²⁺ spiking and slower changes in non-spiking Ca²⁺-GCaMP signals elicited by stress or gene deletion. Short-term exposure to membrane, osmotic or oxidative stress generated immediate stress-specific responses and repeated exposure revealed differential recovery signatures. Osmotic stress caused yeast cell shrinkage and no adaptation response, where Ca²⁺-GCaMP spiking was inhibited by 1 M NaCl but not by 0.666 M CaCl_2. Treatment with sodium dodecylsulfate (SDS) caused a spike-burst, raised the non-spiking Ca²⁺-GCaMP signals, and caused significant cell death, but surviving cells adapted over subsequent exposures. Treatment with 5 mM H₂O₂ abolished spiking and caused transient non-GCaMP-related autofluorescence, but cells adapted such that spiking returned and autofluorescence diminished on repeated exposure. Adaptation to H₂O₂ was dependent on Cap1, extracellular Ca²⁺, and calcineurin but not on its downstream target, Crz1. Ca²⁺-dynamics were not affected by H₂O₂ in the hog1Δ or yvc1Δ mutants, suggesting a pre-adapted, resistant state, possibly due to changes in membrane permeability. Live-cell imaging of Ca²⁺-GCaMP responses in individual cells has, therefore, revealed the dynamics of Ca²⁺-influx, signaling and homeostasis, and their role in the temporal stress response signatures of C. albicans.