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Mitochondrial impairment triggers cytosolic oxidative stress and cell death following proteasome inhibition.

ABSTRACT: Dysfunctions of the mitochondria and the ubiquitin-proteasome system, as well as generation of reactive oxygen species (ROS), are linked to many aging-related neurodegenerative disorders. However, the order of these events remains unclear. Here, we show that the initial impairment occurs in mitochondria under proteasome inhibition. Fluorescent redox probe measurements revealed that proteasome inhibition led to mitochondrial oxidation followed by cytosolic oxidation, which could be prevented by a mitochondrial-targeted antioxidant or antioxidative enzyme. These observations demonstrated that proteasome dysfunction causes damage to mitochondria, leading them to increase their ROS production and resulting in cytosolic oxidation. Moreover, several antioxidants found in foods prevented intracellular oxidation and improved cell survival by maintaining mitochondrial membrane potential and reducing mitochondrial ROS generation. However, these antioxidant treatments did not decrease the accumulation of protein aggregates caused by inhibition of the proteasome. These results suggested that antioxidative protection of mitochondria maintains cellular integrity, providing novel insights into the mechanisms of cell death caused by proteasome dysfunction.

SUBMITTER: Maharjan S

PROVIDER: S-EPMC4116626 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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Mitochondrial impairment triggers cytosolic oxidative stress and cell death following proteasome inhibition.

Maharjan Sunita S Oku Masahide M Tsuda Masashi M Hoseki Jun J Sakai Yasuyoshi Y

Scientific reports 20140731

Dysfunctions of the mitochondria and the ubiquitin-proteasome system, as well as generation of reactive oxygen species (ROS), are linked to many aging-related neurodegenerative disorders. However, the order of these events remains unclear. Here, we show that the initial impairment occurs in mitochondria under proteasome inhibition. Fluorescent redox probe measurements revealed that proteasome inhibition led to mitochondrial oxidation followed by cytosolic oxidation, which could be prevented by a ...[more]

PMID: 25077633

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Project description:Control of intracellular heme levels by extracellular scavenger proteins and intracellular heme oxygenases are essential functions during disease states with enhanced extracellular heme release. During severe hemolysis or rhabdomyolysis uncontrolled heme exposure can cause acute kidney injury and endothelial damage. The cytotoxic activity of heme has been primarily attributed to its pro-oxidative potential. However, the mechanisms of heme toxicity have never been systematically explored. Besides its redox reactivity, heme could also adversely alter cellular functions through its broad binding affinity to multiple non-hemoproteins. Such interactions may impair protein functions and support heme toxicity. In this study we mapped the gene expression profile of Hb triggered acute kidney injury in old blood transfused guinea pigs by serial analysis of gene expression (SAGE). Additionally, the toxic heme response of mouse embryo fibroblasts was systematically characterized on the gene and protein expression levels by gene array experiments and quantitative mass-spectrometry of stable isotope labeled cells. In all these studies, in addition to oxidative stress signals, the most significant signals were reproducibly found for biologic networks related to altered protein degradation, which ultimately triggers the response to unfolded proteins and apoptosis. These screening data could be mechanistically explained by heme-proteasome interactions and a proteasome inhibitor activity of heme. Proteasome inhibition drastically reduced the threshold of cellular toxicity during heme exposure. We therefore propose a novel model of heme toxicity whereby proteasome inhibition by the porphyrin fuels a vicious cycle of oxidative protein modification, accumulation of damaged proteins, cell damage and apoptosis. A two color common reference design was chosen with 2-4 independent biological replicates of each condition. Each experimental sample (Cy5 labeled) was hybridized against a non-treated reference sample (Cy3 labeled). To compensate for dye bias control arrays with competitively hybridized Cy3- and Cy5-labeled non-treated reference samples were used. The latter allowed for a very robust statistical analysis with pair-wise comparison of treatment array replicates versus the corresponding control array replicates.

Project description:Control of intracellular heme levels by extracellular scavenger proteins and intracellular heme oxygenases are essential functions during disease states with enhanced extracellular heme release. During severe hemolysis or rhabdomyolysis uncontrolled heme exposure can cause acute kidney injury and endothelial damage. The cytotoxic activity of heme has been primarily attributed to its pro-oxidative potential. However, the mechanisms of heme toxicity have never been systematically explored. Besides its redox reactivity, heme could also adversely alter cellular functions through its broad binding affinity to multiple non-hemoproteins. Such interactions may impair protein functions and support heme toxicity. In this study we mapped the gene expression profile of Hb triggered acute kidney injury in old blood transfused guinea pigs by serial analysis of gene expression (SAGE). Additionally, the toxic heme response of mouse embryo fibroblasts was systematically characterized on the gene and protein expression levels by gene array experiments and quantitative mass-spectrometry of stable isotope labeled cells. In all these studies, in addition to oxidative stress signals, the most significant signals were reproducibly found for biologic networks related to altered protein degradation, which ultimately triggers the response to unfolded proteins and apoptosis. These screening data could be mechanistically explained by heme-proteasome interactions and a proteasome inhibitor activity of heme. Proteasome inhibition drastically reduced the threshold of cellular toxicity during heme exposure. We therefore propose a novel model of heme toxicity whereby proteasome inhibition by the porphyrin fuels a vicious cycle of oxidative protein modification, accumulation of damaged proteins, cell damage and apoptosis. A two color common reference design was chosen with 2-8 independent biological replicates of each condition. Each experimental sample (Cy5 labeled) was hybridized against a non-treated reference sample (Cy3 labeled). To compensate for dye bias control arrays with competitively hybridized Cy3- and Cy5-labeled non-treated reference samples were used. The latter allowed for a very robust statistical analysis with pair-wise comparison of treatment array replicates versus the corresponding control array replicates.

Dataset Information

Mitochondrial impairment triggers cytosolic oxidative stress and cell death following proteasome inhibition.

Publications

Mitochondrial impairment triggers cytosolic oxidative stress and cell death following proteasome inhibition.

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