The glutathionylation of p65 modulates NF-κB activity in 15-deoxy-Δ12,14-prostaglandin J2-treated endothelial cells

Lin, Yuan-Chun; Huang, Guan-Da; Hsieh, Chia-Wen; Wung, Being‐Sun

doi:10.1016/j.freeradbiomed.2012.02.028

Cited by 29 publications

(21 citation statements)

References 43 publications

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“…5). Lin and coworkers also showed that multiple cysteine residues within p65 are targets for S -glutathionylation and that Cys 38, 160, 216 are critical to transcriptional activity in endothelial cells [43,44]. Our mass spectrometry and transcriptional activation data support their findings.…”

Section: Discussionsupporting

confidence: 83%

N-acetyl-l-cysteine sensitizes pancreatic cancers to gemcitabine by targeting the NFκB pathway

Qanungo

Uys

Manevich

et al. 2014

Biomedicine & Pharmacotherapy

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First-line therapy for pancreatic cancer is gemcitabine. Although tumors may initially respond to the gemcitabine treatment, soon tumor resistance develops leading to treatment failure. Previously, we demonstrated in human MIA PaCa-2 pancreatic cancer cells that N-acetyl-l-cysteine (NAC), a glutathione (GSH) precursor, prevents NFκB activation via S-glutathionylation of p65-NFκB, thereby blunting expression of survival genes. In this study, we documented the molecular sites of S-glutathionylation of p65, and we investigated whether NAC can suppress NFκB signaling and augment a therapeutic response to gemcitabine in vivo. Mass spectrometric analysis of S-glutathionylated p65-NFκB protein in vitro showed post-translational modifications of cysteines 38, 105, 120, 160 and 216 following oxidative and nitrosative stress. Circular dichroism revealed that S-glutathionylation of p65-NFκB did not change secondary structure of the protein, but increased tryptophan fluorescence revealed altered tertiary structure. Gemcitabine and NAC individually were not effective in decreasing MIA PaCa-2 tumor growth in vivo. However, combination treatment with NAC and gemcitabine decreased tumor growth by approximately 50%. NAC treatment also markedly enhanced tumor apoptosis in gemcitabine-treated mice. Compared to untreated tumors, gemcitabine treatment alone increased p65-NFκB nuclear translocation (3.7-fold) and DNA binding (2.5-fold), and these effects were blunted by NAC. In addition, NAC plus gemcitabine treatment decreased anti-apoptotic XIAP protein expression compared to gemcitabine alone. None of the treatments, however, affected extent of tumor hypoxia, as assessed by EF5 staining. Together, these results indicate that adjunct therapy with NAC prevents NFκB activation and improves gemcitabine chemotherapeutic efficacy.

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Section: Discussionsupporting

confidence: 83%

N-acetyl-l-cysteine sensitizes pancreatic cancers to gemcitabine by targeting the NFκB pathway

Qanungo

Uys

Manevich

et al. 2014

Biomedicine & Pharmacotherapy

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“…Specifically, GSH upregulation via 15d-PGJ 2 -induced Nrf2 activation, as discussed above, promotes the modification of p65 at Cys38, Cys160, or Cys216, each of which is a target of electrophilic glutathionylation (79). Beyond inhibiting p65 DNA binding, 15d-PGJ 2 also inhibits degradation of IκB-α, leading to the sequestration of NF-κB in the cytoplasm (80, 81).…”

Section: Pleiotropic Signaling Actions Of Electrophilic Fatty Acidsmentioning

confidence: 99%

Redox-Dependent Anti-Inflammatory Signaling Actions of Unsaturated Fatty Acids

Delmastro-Greenwood

Freeman

Wendell

2014

Annu. Rev. Physiol.

107

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Unsaturated fatty acids are metabolized to reactive products that can act as pro- or anti-inflammatory signaling mediators. Electrophilic fatty acid species, including nitro- and oxo-containing fatty acids, display salutary anti-inflammatory and metabolic actions. Electrophilicity can be conferred by both enzymatic and oxidative reactions, via the homolytic addition of nitrogen dioxide to a double bond or via the formation of α,β-unsaturated carbonyl and epoxide substituents. The endogenous formation of electrophilic fatty acids is significant and influenced by diet, metabolic, and inflammatory reactions. Transcriptional regulatory proteins and enzymes can sense the redox status of the surrounding environment upon electrophilic fatty acid adduction of functionally significant, nucleophilic cysteines. Through this covalent and often reversible posttranslational modification, gene expression and metabolic responses are induced. At low concentrations, the pleiotropic signaling actions that are regulated by these protein targets suggest that some classes of electrophilic lipids may be useful for treating metabolic and inflammatory diseases.

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“…3B). Extensive data exists to support a functional role for PSSG of NF-κB components in controlling the activity of this pathway [22–25], including work in our laboratory demonstrating the importance of IKKβ S-glutathionylation (IKKβ-SSG) [23, 43]. Therefore, we next examined the glutathionylation status of specific proteins in the NF-κB pathway after IL-17A stimulation.…”

Section: Resultsmentioning

confidence: 99%

“…This modification has considerable implications for biological processes, as it can alter the function of various proteins, and both PSSG-induced activation and inactivation have been reported, depending on the protein target [21]. Work by our laboratory and others has demonstrated that the NF-κB pathway, as well as upstream adaptor molecules such as TRAF6, are negatively regulated via PSSG [22–26]. Under physiological conditions, the thiol oxidoreductase glutaredoxin-1 (Grx1) can deglutathionylate proteins, thus restoring the native thiol form of the target cysteine residue [27].…”

Section: Introductionmentioning

confidence: 99%

“…Under physiological conditions, the thiol oxidoreductase glutaredoxin-1 (Grx1) can deglutathionylate proteins, thus restoring the native thiol form of the target cysteine residue [27]. Numerous studies initially focused on oxidant-induced PSSG, but more recent studies have demonstrated the role of this modification in the context of biologically relevant stimuli [22, 28–30]. …”

Section: Introductionmentioning

confidence: 99%

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The glutaredoxin/S-glutathionylation axis regulates interleukin-17A-induced proinflammatory responses in lung epithelial cells in association with S-glutathionylation of nuclear factor κB family proteins

Nolin

Tully

Hoffman

et al. 2014

Free Radical Biology and Medicine

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Interleukin-17A (IL-17A) is a newly emerging player in the pathogenesis of chronic lung diseases that amplifies inflammatory responses and promotes tissue remodeling. Stimulation of lung epithelial cells with IL-17A leads to activation of the transcription factor, nuclear factor kappa B (NF-κB), a key player in the orchestration of lung inflammation. We have previously demonstrated the importance of the redox-dependent post-translational modification, S-glutathionylation, in limiting activation of NF-κB and downstream gene induction. Under physiological conditions, the enzyme glutaredoxin 1 (Grx1) acts to deglutathionylate NF-κB proteins, which restores functional activity. In this study, we sought to determine the impacts of S-glutathionylation on IL-17A-induced NF-κB activation and expression of pro-inflammatory mediators. C10 mouse lung alveolar epithelial cells, or primary mouse tracheal epithelial cells exposed to IL-17A show rapid activation of NF-κB, and the induction of pro-inflammatory genes. Upon IL-17A exposure, sulfenic acid formation and S-glutathionylated proteins increased. Assessment of S-glutathionylation of NF-κB pathway components revealed S-glutathionylation of RelA (RelA-SSG) and inhibitory kappa B kinase alpha (IKKα-SSG) after stimulation with IL-17A. SiRNA-mediated ablation of Grx1 increased both RelA-SSG and IKKα-SSG and acutely increased nuclear content of RelA, and tended to decrease nuclear RelB. SiRNA mediated ablation or genetic ablation of Glrx1 decreased the expression of NF-κB regulated genes, KC and CCL20, in response to IL-17A, but conversely increased the expression of IL-6. Lastly, siRNAmediated ablation of IKKα attenuated nuclear RelA and RelB content and decreased expression of KC and CCL20 in response to IL-17A. Together, these data demonstrate a critical role for the S-glutathionylation/Grx1 redox axis in regulating IKKα and RelA S-glutathionylation and the responsiveness of epithelial cells to IL-17A.

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The glutathionylation of p65 modulates NF-κB activity in 15-deoxy-Δ12,14-prostaglandin J2-treated endothelial cells

Cited by 29 publications

References 43 publications

N-acetyl-l-cysteine sensitizes pancreatic cancers to gemcitabine by targeting the NFκB pathway

N-acetyl-l-cysteine sensitizes pancreatic cancers to gemcitabine by targeting the NFκB pathway

Redox-Dependent Anti-Inflammatory Signaling Actions of Unsaturated Fatty Acids

The glutaredoxin/S-glutathionylation axis regulates interleukin-17A-induced proinflammatory responses in lung epithelial cells in association with S-glutathionylation of nuclear factor κB family proteins

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