MassIVEapplication/xmlftp://massive-ftp.ucsd.edu/v01/MSV000079314/primaryOK2000000ProteomicsDavid StokoeOrbitrap FusionHomo Sapiens (ncbitaxon:9606)https://massive.ucsd.edu/ProteoSAFe/dataset.jsp?task=8c37f6a7a76746efb090fa76c4460415MassIVE128MS:1002864 - No post-translational-modifications are included in the identified peptides of this datasetThe KEAP1/Nrf2 pathway senses and responds to changes in intracellular oxidative stress. Mutations that result in constitutive activation of Nrf2 are present in several human tumors, especially non-small cell lung cancer. Therefore, compounds that inhibit Nrf2 activity might be beneficial in treating patients whose tumors show activation of this pathway. Recent reports suggest that the natural product brusatol can potently and selectively inhibit Nrf2 activity, resulting in cell cytotoxicity, and can be effectively combined with chemotherapeutic agents. Here, we analyzed the effects of brusatol on the cellular proteome in the KEAP1 mutant non-small cell lung cancer cell line A549. Brusatol was found to rapidly and potently decrease the expression of the majority of detected proteins, including Nrf2. The most dramatically decreased proteins are those that display a short half-life, like Nrf2. This effect was confirmed by restricting the analysis to newly synthesized proteins using a labeled methionine analogue. Moreover, brusatol increased the expression of multiple components of the ribosome, suggesting that it regulates the function of this macromolecular complex. Finally, we show that brusatol induces its potent cellular cytotoxicity effects on multiple cancer cell lines in a manner independent of KEAP1/Nrf2 activity and with a profile similar to the protein translation inhibitor silvestrol. In conclusion, our data show that the activity of brusatol is not restricted to Nrf2 but, rather, functions as a global protein synthesis inhibitor.Application of Mass Spectrometry Profiling to Establish Brusatol as an Inhibitor of Global Protein Synthesis.Vartanian Steffan S, Ma Taylur P TP, Lee James J, Haverty Peter M PM, Kirkpatrick Donald S DS, Yu Kebing K, Stokoe David DMass Spectrum Analysis, Svc, Spectroscopy, Manuscripts., Mass Spectrum, Bru, MS, Col4a-1, Raw, Analyses, Mass, Spectrometry, Del(8)44H, Analysis, Spectrum Analyses, Mass Spectrum Analyses, Spectrum Analysis, Mass SpectroscopyRaw., Svc, Bru, Del(8)44H, Col4a-1DNA Oxidative, protein translation, Ribosomal Peptide, (11 beta, nucleocytoplasm, KEAP1, Activity, determination, Non-Small Cell Lung Cancer, Nitrative Stress, ribosomal RNA, Ribosomal Protein, Tumor, Damage, l(3)j5E7, 11, Mutations, dmTAF[[II]]230, Lung Carcinomas, L-Isomer Methionine, Methionine, Oxidative DNA, KIAA0132, Oxidative, Dmel_CG13826, Nitro-Oxidative Stress, Non-Small-Cell Lung Carcinoma, Line, NFT2, Inhibitors, Nitro-Oxidative Stresses, Non-Small Cell Lung Carcinoma, Oxidative Injury, present in fewer numbers in organism, DNA Damage, increased, Oxidative Injuries, Ribosomal, A-549, Man (Taxonomy), TFIID TAF250, mRNA Translation, cel, M, Oxidative Cleavage, CG3962, Non-Small Cell Lung, RCH04A07, Oxidative DNA Damages, keap1, Non Small Cell Lung Carcinoma, Anti-oxidative, Oxidative Stress Injuries, Oxidative Stresses, decreased, Halflifes, malignant neoplasm, Half Life, Dmel_CG17894, Malignancies, Racemethionine, intracellular, Tumors, dKEAP1, Carcinoma, protein anabolism, dTAF[[II]]230, protein biosynthetic process, 12- trihydroxy-15-((3-methyl-1-oxo-2-butenyl)oxy)- 2, Modern, 2-Amino-4-(methylthio)butyric acid, TAF200, macromolecule complex, TAFII-250, TAF250/230, l(3)03921, NRF2A, non-small cell lung cancer, shortened, TAFII250, Benign, Half-Lifes, Oxidative and Nitrosative Stress, dKeap1, protein formation, Methionin, DNA Oxidative Damages, Hmet, internal to cell, non-small cell lung carcinoma (disease), DmelCG43286, activation, Lines, Carcinomas, Ribosomal Protein Synthesis, Synthesis Antagonist, Antagonists, dkeap1, Benign Neoplasms, CG17603, TAF[[II]], mKIAA0132, human, Non Small Cell Lung, Malignant Neoplasms, Nitro-Oxidative, non-small cell carcinoma of lung, Protein Synthesis, DmelCG3962, Protein Translation, Patient, Taf250, protein synthesis, SR3-5, Oxidative Stress Injury, DNA, 20-epoxy-3, short, Liquimeth, Inhibitor, Proteomes, humans, TAF230, accessory, Oxidative Damage, CG13826, Protein Biosynthesis, Oxidative Stress, d230, human being, 2-amino-4-(methylsulfanyl)butanoic acid, anon-WO0153538.7, anon-WO0153538.6, Antioxidative, non-small cell lung carcinoma, Neoplasms, INRF2, INrf2, Benign Neoplasm, L Isomer, Antioxidative Stress, Synthesis, Gene, dTAFII250, Non-Small-Cell Lung, non-small cell carcinoma of the lung, Malignant, EfW1, RCB0098, supernumerary, Stresses, Human, Inrf2, non-small cell cancer of the lung, Translation, NSCLC cell line, Oxidative Nitrative, Homo sapiens, reduced, E4TF1-60, Stress Injury, dmTAF1, Taf230, subnumerary, CG4566, Gene Products, Oxidative Cleavages, L-Methionine, NSCLC - non-small cell lung cancer, CNC, Cnc, Man, Oxidative Damages, Pedameth, TAF250, protoplasm, KLHL19, Genetic Translations, Taf200, Injury, dTAF[[II]]250, Genetic, protoplast, non small cell lung cancer cell line, Malignancy, Ribosomal Protein Biosynthesis, cell, stubby, Peptide Biosynthesis, inhibiteur, NRF2, Nrf2, Taf1p, 16-dioxopicras-3-en-21-oic acid, decreased number, Antagonist, Lung Carcinoma, 15 beta)-13, Protein Synthesis Antagonists., Neoplasias, dTAF250, Halflife, NSCLC, 2-amino-4-(methylthio)butanoic acid, inhibidor, Anti oxidative Stress, Oxidative DNA Damage, Antioxidative Stresses, Clients, Nonsmall Cell Lung Cancer, A549 cell, Ribosomal Peptide Biosynthesis, TAF, metionina, Non-Small-Cell Lung Carcinomas, Genetic Translation, mRNA Translations, Cancer, inhibitors, TAF[[II]]250, A549, Malignant Neoplasm, DL-Methionine, protein complex, Proteins, Cell Lines, methyl ester, total expressed protein, l(3)84Ab, inhibitor, BG:DS00004.13, function, Anti-oxidative Stresses, L-Isomer, Client, Cell, dTAF230, Synthesis Inhibitor, BcDNA:RE05559, Oxidative Nitrative Stress, MT, 12 alpha, Oxidative Nitrative Stresses, p230, Protein, chemical analysis, Neoplasm, Protein Synthesis Antagonist, TAF[[II]]250/230, TFIID, protein biosynthesis, CG43286, Non-small cell lung cancer, membrane bound ribosome, CG17894, Met, free ribosome, Taf[[II]]250, Protein Synthesis Inhibitor, Ribosome, macromolecular complex, primary cancer, Anti-oxidative Stress, TAF[[II]]230, DNA Oxidative Damage, Biosynthesis, mRNA, increased number, protein containing complex, CG4578, CNC_DROME, TAF[II]250, Cancers, Cleavage, malignant tumor, Protein Gene Products, AI194320, Gene Proteins, present in greater numbers in organism, 5134, DmelCG17603, Modern Man, Stress, alpha-amino-gamma-methylmercaptobutyric acid, E4TF1A, assay, non-small cell cancer of lung, Nitro Oxidative Stress, General activity, Neoplasia, protein-protein complex, Non-Small-Cell, TAF1Mass Spectrum Analysis, inhibitors, protein translation, Mass Spectrum, protein anabolism, (11 beta, protein biosynthetic process, Analyses, 12- trihydroxy-15-((3-methyl-1-oxo-2-butenyl)oxy)- 2, Spectrometry, methyl ester, inhibiteur, inhibitor, Spectrum Analyses, 16-dioxopicras-3-en-21-oic acid, Spectrum Analysis, 15 beta)-13, 11, Spectroscopy, protein synthesis., MS, inhibidor, 12 alpha, Mass, protein formation, protein biosynthesis, Analysis, 20-epoxy-3, Mass Spectrum Analyses, Mass Spectroscopy0PXD003389trueVartanian et al. 2015 raw dataa list of raw mass spectrometry data files for manuscript authored by Vartanian et al.MSV00007931426711467