title: Proteasome inhibition and oxidative reactions disrupt cellular homeostasis during heme stress.
abstract: Dual control of cellular heme levels by extracellular scavenger proteins and degradation by heme oxygenases is essential in diseases associated with increased heme release. During severe hemolysis or rhabdomyolysis, uncontrolled heme exposure can cause acute kidney injury and endothelial cell damage. The toxicity of heme was primarily attributed to its pro-oxidant effects; however additional mechanisms of heme toxicity have not been studied systematically. In addition to redox reactivity, heme may adversely alter cellular functions by binding to essential proteins and impairing their function. We studied inducible heme oxygenase (Hmox1)-deficient mouse embryo fibroblast cell lines as a model to systematically explore adaptive and disruptive responses that were triggered by intracellular heme levels exceeding the homeostatic range. We extensively characterized the proteome phenotype of the cellular heme stress responses by quantitative mass spectrometry of stable isotope-labeled cells that covered more than 2000 individual proteins. The most significant signals specific to heme toxicity were consistent with oxidative stress and impaired protein degradation by the proteasome. This ultimately led to an activation of the response to unfolded proteins. These observations were explained mechanistically by demonstrating binding of heme to the proteasome that was linked to impaired proteasome function. Oxidative heme reactions and proteasome inhibition could be differentiated as synergistic activities of the porphyrin. Based on the present data a novel model of cellular heme toxicity is proposed, whereby proteasome inhibition by heme sustains a cycle of oxidative stress, protein modification, accumulation of damaged proteins and cell death.
authors.affiliation: Division of Internal Medicine, University of Zurich, CH-8091 Zurich, Switzerland.
authors.forename: F
authors.initials: F
authors.lastname: Vallelian
authors.affiliation: Division of Internal Medicine, University of Zurich, CH-8091 Zurich, Switzerland.
authors.forename: J W
authors.initials: JW
authors.lastname: Deuel
authors.affiliation: Functional Genomics Center Zurich, Swiss Federal Institute of Technology Zurich/University of Zurich, Zurich, Switzerland.
authors.forename: L
authors.initials: L
authors.lastname: Opitz
authors.affiliation: 1] Division of Internal Medicine, University of Zurich, CH-8091 Zurich, Switzerland [2] Institute of Anesthesiology, University of Zurich, Zurich, Switzerland.
authors.forename: C A
authors.initials: CA
authors.lastname: Schaer
authors.affiliation: 1] Division of Internal Medicine, University of Zurich, CH-8091 Zurich, Switzerland [2] Functional Genomics Center Zurich, Swiss Federal Institute of Technology Zurich/University of Zurich, Zurich, Switzerland.
authors.forename: M
authors.initials: M
authors.lastname: Puglia
authors.affiliation: School of Medical Sciences, Discipline of Pathology and Bosch Institute, University of Sydney, Sydney, New South Wales, Australia.
authors.forename: M
authors.initials: M
authors.lastname: Lönn
authors.affiliation: Division of Internal Medicine, University of Zurich, CH-8091 Zurich, Switzerland.
authors.forename: W
authors.initials: W
authors.lastname: Engelsberger
authors.affiliation: Functional Genomics Center Zurich, Swiss Federal Institute of Technology Zurich/University of Zurich, Zurich, Switzerland.
authors.forename: S
authors.initials: S
authors.lastname: Schauer
authors.affiliation: Laboratory of Biochemistry and Vascular Biology, Center of Biologics Evaluation and Research (CBER), FDA, Bethesda, MD, USA.
authors.forename: E
authors.initials: E
authors.lastname: Karnaukhova
authors.affiliation: Institute of Anesthesiology, University of Zurich, Zurich, Switzerland.
authors.forename: D R
authors.initials: DR
authors.lastname: Spahn
authors.affiliation: 1] School of Medical Sciences, Discipline of Pathology and Bosch Institute, University of Sydney, Sydney, New South Wales, Australia [2] Victor Chang Cardiac Research Institute and University of New South Wales, Sydney, New South Wales, Australia.
authors.forename: R
authors.initials: R
authors.lastname: Stocker
authors.affiliation: Laboratory of Biochemistry and Vascular Biology, Center of Biologics Evaluation and Research (CBER), FDA, Bethesda, MD, USA.
authors.forename: P W
authors.initials: PW
authors.lastname: Buehler
authors.affiliation: 1] Division of Internal Medicine, University of Zurich, CH-8091 Zurich, Switzerland [2] Center of Evolutionary Medicine, University of Zurich, Zurich, Switzerland.
authors.forename: D J
authors.initials: DJ
authors.lastname: Schaer
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D000069286:Bortezomib / Q000494:pharmacology; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002942:Circular Dichroism; D057809:HEK293 Cells; D006360:Heat-Shock Proteins / Q000378:metabolism; D006418:Heme / Q000494:pharmacology*; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D018345:Mice, Knockout; D018384:Oxidative Stress / Q000187:drug effects*; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry / Q000378:metabolism*; D061988:Proteasome Inhibitors / Q000494:pharmacology; D011485:Protein Binding; D000071456:Sequestosome-1 Protein; D013056:Spectrophotometry, Ultraviolet; D025801:Ubiquitin / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D061988:Proteasome Inhibitors; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse; D025801:Ubiquitin; D006418:Heme; D000069286:Bortezomib; D051547:Heme Oxygenase-1; C496542:Hmox1 protein, mouse; D046988:Proteasome Endopeptidase Complex