Development of an in vitro model to dissect the mechanism of fungal persistence in the cystic fibrosis lung

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Cystic fibrosis (CF) is the most common fatal genetically inherited disease in Caucasian populations. This disease is caused by mutations in the cystic fibrosis transmembrane regulator gene (CFTR) which produces defective ion fluxes and calcium homeostasis in the epithelia. Patients with cystic fibrosis are susceptible to lung infections caused by the fungal pathogen Aspergillus fumigatus.Animal models play a central role in the study of cystic fibrosis. Researchers have used mice, ferrets, pigs and sheep to model the inflammatory response seen in human patients with this disease. Although such models generate useful knowledge on the pathophysiology of this disease, animal models are limited for a number of ethical reasons.Here we propose to establish a new in vitro model of isogenic CF and healthy controls bronchial epithelial cell lines carrying different mutations in the CFTR gene using CRISPR/Cas9 mutagenesis. Using an established methodological approach in our laboratory including in vitro infection, confocal microscopy, expression analysis, cytokine measurements and competitive fitness analysis the student will be able to define critical stages of the interaction between the cystic fibrosis epithelia and A. fumigatus compared to healthy cells and the fungal factors governing this process.The successful completion of this project will achieve a number of scientific objectives. Firstly, the development of an isogenic cell culture system of epithelial cell lines will be of great value for researchers focused on the study of epithelia-pathogen interactions in CF. Secondly, screening of A. fumigatus null libraries on CF epithelia will increase our knowledge on the pathogen factors leading persistence in the context of disease. Additionally, the student will be enrolled in a programme that will give the student the opportunity to enroll with industrial partners in Manchester and our fungal community to promote the implementation on this resource in other laboratories at an international level.