Dataset Information

Mechanisms of Nrf2 protection in astrocytes as identified by quantitative proteomics and siRNA screening.

ABSTRACT: The Nrf2 (NF-E2 related factor 2)-ARE (antioxidant response element) pathway controls a powerful array of endogenous cellular antioxidant systems and is an important pathway in the detoxification of reactive oxygen species (ROS) in the brain. Using a combination of quantitative proteomics and siRNA screening, we have identified novel protective mechanisms of the Nrf2-ARE pathway against oxidative stress in astrocytes. Studies from our lab and others have shown Nrf2 overexpression protects astrocytes from oxidative stress. However, the exact mechanisms by which Nrf2 elicits these effects are unknown. In this study, we show that induction of Nrf2 reduces levels of reactive oxygen species (ROS) produced by various oxidative stressors and results in robust cytoprotection. To identify the enzymes responsible for these effects, we used stable isotope labeling by amino acids in cell culture (SILAC) and quantitative shotgun proteomics to identify 72 Nrf2-regulated proteins in astrocytes. We hypothesized a subset of these proteins might play a critical role in Nrf2 protection. In order to identify these critical proteins, we used bioinformatics to narrow our target list of proteins and then systematically screened each candidate with siRNA to assess the role of each in Nrf2 protection. We screened each target against H2O2, tert-butyl hydroperoxide, and 4-hydroxynonenal and subsequently identified three enzymes-catalase, prostaglandin reductase-1, and peroxiredoxin-6-that are critical for Nrf2-mediated protection in astrocytes.

SUBMITTER: Dowell JA

PROVIDER: S-EPMC3726381 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Mechanisms of Nrf2 protection in astrocytes as identified by quantitative proteomics and siRNA screening.

Dowell James A JA Johnson Jeffrey A JA

PloS one 20130729 7

The Nrf2 (NF-E2 related factor 2)-ARE (antioxidant response element) pathway controls a powerful array of endogenous cellular antioxidant systems and is an important pathway in the detoxification of reactive oxygen species (ROS) in the brain. Using a combination of quantitative proteomics and siRNA screening, we have identified novel protective mechanisms of the Nrf2-ARE pathway against oxidative stress in astrocytes. Studies from our lab and others have shown Nrf2 overexpression protects astroc ...[more]

PMID: 23922950

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Project description:RelevanceMolecular network changes are the hallmark of the pathogenesis of ovarian cancers (OCs). Network-based biomarkers benefit for the effective treatment of OC.PurposeThis study sought to identify key pathway-network alterations and network-based biomarkers for clarification of molecular mechanisms and treatment of OCs.MethodsIngenuity Pathway Analysis (IPA) platform was used to mine signaling pathway networks with 1198 human tissue mitochondrial differentially expressed proteins (mtDEPs) and compared those pathway network changes between OCs and controls. The mtDEPs in important cancer-related pathway systems were further validated with qRT-PCR and Western blot in OC cell models. Moreover, integrative analysis of mtDEPs and Cancer Genome Atlas (TCGA) data from 419 patients was used to identify hub molecules with molecular complex detection method. Hub molecule-based survival analysis and multiple multivariate regression analysis were used to identify survival-related hub molecules and hub molecule signature model.ResultsPathway network analysis revealed 25 statistically significant networks, 192 canonical pathways, and 5 significant molecular/cellular function models. A total of 52 canonical pathways were activated or inhibited in cancer pathogenesis, including antigen presentation, mitochondrial dysfunction, GP6 signaling, EIF2 signaling, and glutathione-mediated detoxification. Of them, mtDEPs (TPM1, CALR, GSTP1, LYN, AKAP12, and CPT2) in those canonical pathway and molecular/cellular models were validated in OC cell models at the mRNA and protein levels. Moreover, 102 hub molecules were identified, and they were regulated by post-translational modifications and functioned in multiple biological processes. Of them, 62 hub molecules were individually significantly related to OC survival risk. Furthermore, multivariate regression analysis of 102 hub molecules identified significant seven hub molecule signature models (HIST1H2BK, ALB, RRAS2, HIBCH, EIF3E, RPS20, and RPL23A) to assess OC survival risks.ConclusionThese findings provided the overall signaling pathway network profiling of human OCs; offered scientific data to discover pathway network-based cancer biomarkers for diagnosis, prognosis, and treatment of OCs; and clarify accurate molecular mechanisms and therapeutic targets. These findings benefit for the discovery of effective and reliable biomarkers based on pathway networks for OC predictive and personalized medicine.

Dataset Information

Mechanisms of Nrf2 protection in astrocytes as identified by quantitative proteomics and siRNA screening.

Publications

Mechanisms of Nrf2 protection in astrocytes as identified by quantitative proteomics and siRNA screening.

Similar Datasets

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