Both terminal misfolding and polymerisation contribute to disease-relevant phenotypic changes in cell models of α1-antitrypsin deficiency-associated liver disease

Stylianou, P.; Adoni, K.; Nyon, M.-P.; Cryar, A.; Lee, N. O.; Haq, I.; Dickens, J. A.; Segeritz, C.-P.; Smith, G.; Ordonez, A.; Irving, J. A.; Lomas, D. A.; Leighton, K.-M.; Beasley, J.; Jones, A. W. E.; Rashid, S. T.; Vallier, L.; Thalassinos, K.; Gooptu, B.

doi:10.1101/2025.09.30.679226

bioRxiv · 10.1101/2025.09.30.679226

Both terminal misfolding and polymerisation contribute to disease-relevant phenotypic changes in cell models of α1-antitrypsin deficiency-associated liver disease

Stylianou, P.·Adoni, K.·Nyon, M.-P.·Cryar, A.·Lee, N. O.·Haq, I.·Dickens, J. A.·Segeritz, C.-P.·Smith, G.·Ordonez, A.·Irving, J. A.·Lomas, D. A.·Leighton, K.-M.·Beasley, J.·Jones, A. W. E.·Rashid, S. T.·Vallier, L.·Thalassinos, K.·Gooptu, B.

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Abstract

Polymerisation of 1-antitrypsin within hepatocytes is considered central to the pathogenesis of 1-antitrypsin deficiency-associated liver fibrosis, most commonly in homozygotes for the Z (p.Glu342Lys) allele. Polymerisation proceeds via self-association of monomeric intermediate states. In parallel, >50% of synthesised Z 1-antitrypsin is instead recognized as terminally-misfolded and degraded. It is unclear whether this contributes to Z 1-antitrypsin deficiency-associated liver disease. We characterised the relationships between polymer formation, terminal misfolding and their cellular consequences, using label-free proteomics mass spectrometry (MS), light and electron microscopy, and cellular assays. Proteomic analyses of well-established CHO cell models of hepatocyte handling of 1-antitrypsin variants indicated that cellular responses to the Z mutation were surprisingly similar to those seen with the NullHongKong variant (NHK), which can only misfold terminally and cannot polymerise. A minor set of proteins showed increases associated with Z and not NHK 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 the degree of terminal misfolding of the expressed 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 1-antitrypsin deficiency. Molecular therapies may prove most efficacious for associated liver disease if they address terminal misfolding as well as polymerisation.

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Stylianou, P., Adoni, K., Nyon, M.-P., Cryar, A., Lee, N. O., Haq, I., Dickens, J. A., Segeritz, C.-P., Smith, G., Ordonez, A., Irving, J. A., Lomas, D. A., Leighton, K.-M., Beasley, J., Jones, A. W. E., Rashid, S. T., Vallier, L., Thalassinos, K., Gooptu, B.. 2025-10-01. Both terminal misfolding and polymerisation contribute to disease-relevant phenotypic changes in cell models of α1-antitrypsin deficiency-associated liver disease. https://doi.org/10.1101/2025.09.30.679226

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