Both terminal misfolding and polymerisation contribute to disease-relevant phenotypic changes in cell models of α ₁ -antitrypsin deficiency-associated liver disease

Polymerisation of α ₁ -antitrypsin within hepatocytes is considered central to pathogenesis in α ₁ -antitrypsin deficiency-associated liver fibrosis, most commonly seen in homozygotes for the Z (p.Glu342Lys) allele. Polymerisation proceeds via self-association of monomeric intermediate states. In parallel, >50% of synthesised Z α ₁ -antitrypsin is instead recognized as terminally-misfolded and degraded. It is unclear whether this contributes to Z α ₁ -antitrypsin deficiency-associated liver disease. We characterised the relationships between polymer formation, terminal misfolding and their cellular consequences, using ion-mobility and label-free proteomics mass spectrometry, light and electron microscopy, and cellular assays. IM-MS findings supported the relationship between polymerisation propensity of different variants and expansion of the protein consistent with opening of its β-sheet A. Proteomic analyses of well-established CHO cell models of hepatocyte handling of α ₁ -antitrypsin variants indicated that cellular responses to the Z mutation were surprisingly similar to those seen with the Null _HongKong variant (NHK), which can only misfold terminally and cannot polymerise. A minor set of proteins showed increases associated with Z and not NHK α ₁ -antitrypsin expression, consistent with a polymer-specific response, characterized by association with increased organellar organization and vesicle-mediated transport. Conversely, proteostatic and pro-fibrotic integrin-associated pathways increased with terminal misfolding tendency of the expressed α ₁ -antitrypsin variant. Bioenergetic pathway changes indicated concomitant switching from oxidative to glycolytic metabolism. Cell studies further correlated fibrosis-associated behaviours with terminal misfolding rather than polymerisation. Terminal misfolding, as well as polymerisation behaviour, may therefore be important for pro-fibrotic responses including metabolic reprogramming and senescence in Z α ₁ -antitrypsin deficiency. Molecular therapies may prove most efficacious for associated liver disease if they address terminal misfolding as well as polymerisation consequences.