Different Stress-Induced Calcium Signatures Are Reported by Aequorin-Mediated Calcium Measurements in Living Cells of Aspergillus fumigatus

• Published in: PloS one Vol. 10; no. 9; p. e0138008
• Main Authors: Muñoz, Alberto, Bertuzzi, Margherita, Bettgenhaeuser, Jan, Iakobachvili, Nino, Bignell, Elaine M, Read, Nick D
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 24.09.2015; Public Library of Science (PLoS)
• Subjects: Aequorin - genetics; Aequorin - metabolism; Age; Aspergillus; Aspergillus fumigatus; Aspergillus fumigatus - physiology; Bacteriology; Biology; Biomass; Calcium; Calcium (extracellular); Calcium (intracellular); Calcium - metabolism; Calcium ions; Calcium signalling; Calcium-binding protein; Calmodulin; Candida albicans; Causes of; Chelating Agents - pharmacology; Chemical treatment; Development and progression; Disease; Drug resistance; Egtazic Acid - analogs & derivatives; Egtazic Acid - pharmacology; Fungal infections; Fungi; Gene Expression; Genetic aspects; Health aspects; Homeostasis; Homologous recombination; Homology; Inflammation; Intracellular Space - metabolism; Kinases; Lungs; Mycoses; Neurospora crassa; Osmotic Pressure; Osmotic shock; Oxidative Stress; Pathogenesis; Phenotype; Physiological aspects; Proteins; Renilla-luciferin 2-monooxygenase; Research parks; Signal transduction; Signatures; Stimuli; Stress response; Stress, Mechanical; Stress, Physiological; Trifluoperazine; United Kingdom; United Kingdom > UK; Edinburgh Scotland
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0138008

Aspergillus fumigatus is an inhaled fungal pathogen of human lungs, the developmental growth of which is reliant upon Ca2+-mediated signalling. Ca2+ signalling has regulatory significance in all eukaryotic cells but how A. fumigatus uses intracellular Ca2+ signals to respond to stresses imposed by the mammalian lung is poorly understood. In this work, A. fumigatus strains derived from the clinical isolate CEA10, and a non-homologous recombination mutant ΔakuBKU80, were engineered to express the bioluminescent Ca2+-reporter aequorin. An aequorin-mediated method for routine Ca2+ measurements during the early stages of colony initiation was successfully developed and dynamic changes in cytosolic free calcium ([Ca2+]c) in response to extracellular stimuli were measured. The response to extracellular challenges (hypo- and hyper-osmotic shock, mechanical perturbation, high extracellular Ca2+, oxidative stress or exposure to human serum) that the fungus might be exposed to during infection, were analysed in living conidial germlings. The 'signatures' of the transient [Ca2+]c responses to extracellular stimuli were found to be dose- and age-dependent. Moreover, Ca2+-signatures associated with each physico-chemical treatment were found to be unique, suggesting the involvement of heterogeneous combinations of Ca2+-signalling components in each stress response. Concordant with the involvement of Ca2+-calmodulin complexes in these Ca2+-mediated responses, the calmodulin inhibitor trifluoperazine (TFP) induced changes in the Ca2+-signatures to all the challenges. The Ca2+-chelator BAPTA potently inhibited the initial responses to most stressors in accordance with a critical role for extracellular Ca2+ in initiating the stress responses.