Maintenance of cellular protein homeostasis (proteostasis) depends on a complex network of molecular chaperones, proteases and other regulatory factors. Proteostasis deficiency develops during normal aging and predisposes individuals for many diseases, including neurodegenerative disorders. Here we describe sensor proteins for the comparative measurement of proteostasis capacity in different cell types and model organisms. These sensors are increasingly structurally destabilized versions of firefly luciferase. Imbalances in proteostasis manifest as changes in sensor solubility and luminescence activity. We used EGFP-tagged constructs to monitor the aggregation state of the sensors and the ability of cells to solubilize or degrade the aggregated proteins. A set of three sensor proteins serves as a convenient toolkit to assess the proteostasis status in a wide range of experimental systems, including cell and organism models of stress, neurodegenerative disease and aging.
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We thank M. Hipp, Y.E. Kim and M. Hayer-Hartl for discussion, F. Buchholz (Max Planck Institute for Molecular Cell Biology and Genetics) for providing Hsc70 endoribonuclease-prepared small interfering (esi)RNA, and H. Wagner (Institut für Med. Mikrobiologie, Immunologie und Hygiene, Technische Universität München) for the HSP70-Luc plasmid. We acknowledge the expert technical assistance of V. Marcus. This work has been supported by the European Commission within the 7th framework program Proteomics Specification in Time and Space.
F.U.H is a paid consultant of Proteostasis Therapeutics Inc. A.V and D.G are employees of Proteostasis Therapeutics Inc.
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Gupta, R., Kasturi, P., Bracher, A. et al. Firefly luciferase mutants as sensors of proteome stress. Nat Methods 8, 879–884 (2011). https://doi.org/10.1038/nmeth.1697
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