Cellular homeostasis and healthy ageing: protein folding, redox and metabolism

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At a cellular level, healthy ageing depends upon the resilience of homoeostatic mechanisms. Pathways regulating clearance of misfolded proteins (proteostasis), redox responses, metabolic pathways and general injury responses appear important in this regard and may be linked. In tissue, collapse of these processes manifests as fibrosis or scarring. I plan to probe their relationships with each other. I propose to do this by using CRISPR editing of a promoter region to generate pairs of epithelial cell lines differing only in the degree of proteostatic challenge relating to expression of a single mucin protein, MUC5B. To fold properly this protein requires correct formation of three pairs of intramolecular disulphide bonds between six cysteine residues. This requires considerable engagement of the regulatory machinery of redox homoeostasis and protein folding within the endoplasmic reticulum (ER). Its overexpression in epithelial cells is associated with ER stress, and with dysfunctional injury and scarring responses in tissue. I will therefore use my cell models to correlate levels of homoeostatic challenge in protein folding, redox, and metabolic reprogramming. I further propose to assess the degree to which these responses relate to regulation of pro-fibrotic cross-talk between epithelial and mesenchymal cells by modulating ER stress, redox, and mitochondrial function.