Role of a Mitogen-activated Protein Kinase Pathway in the Induction of Phase II Detoxifying Enzymes by Chemicals

Yu, Rong; Wei, Lei; Mandlekar, Sandhya; Weber, Michael J.; Der, Channing J.; Wu, Jie; Kong, Ah Ng Tony

doi:10.1074/jbc.274.39.27545

Cited by 267 publications

(191 citation statements)

References 45 publications

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“…Furthermore, using inhibitors of the MAPK/ERK pathway, it appears that this pathway may lead to phosphorylation of Nrf2 and its increased stability. These findings confirm previous reports implicating the MAPK/ ERK pathway in the regulation of ARE-responsive genes (21,22). Although PKC can phosphorylate Nrf2, these kinases did not seem to affect its stability.…”

Section: Discussionsupporting

confidence: 91%

“…It has been previously reported that the MAPK/ERK signaling pathway may be involved in the regulation of the ARE response (21,22). We sought to determine if phosphorylation events mediated by this pathway may have a role in promoting Nrf2 stability.…”

Section: Stabilization Of Nrf2 Is Dependent On Phosphorylation By Thementioning

confidence: 99%

“…These include the mitogen-activated protein kinase cascades (21)(22)(23), the phosphatidylinositol 3-kinase (PI3K)-dependent pathways (24 -26), and the protein kinase C (PKC)-dependent pathway (27). We have previously reported that activation of Nrf2 in HepG2 cells treated with tBHQ, ␤-NF, or 12-O-tetradecanoylphorbol-13-acetate involves in part a phosphorylation-dependent mechanism mediated by PKC, which appears to promote translocation of the transcription factor into the nucleus (27).…”

mentioning

confidence: 99%

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Increased Protein Stability as a Mechanism That Enhances Nrf2-mediated Transcriptional Activation of the Antioxidant Response Element

Nguyen¹,

Sherratt²,

Huang³

et al. 2003

Journal of Biological Chemistry

554

382

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Nrf2 (NF-E2Similarly, the protein phosphatase inhibitor okadaic acid also caused an accumulation of Nrf2, whereas the reverse effects were observed with PD 98059 and U 0126, two compounds that block the activation of the MAPK/ ERK signaling cascade. These data suggest that Nrf2 is degraded by the ubiquitin-dependent pathway and that phosphorylation of Nrf2 leads to an increase in its stability and subsequent transactivation activity.

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Section: Discussionsupporting

confidence: 91%

Section: Stabilization Of Nrf2 Is Dependent On Phosphorylation By Thementioning

confidence: 99%

mentioning

confidence: 99%

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Increased Protein Stability as a Mechanism That Enhances Nrf2-mediated Transcriptional Activation of the Antioxidant Response Element

Nguyen¹,

Sherratt²,

Huang³

et al. 2003

Journal of Biological Chemistry

554

382

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show abstract

“…Previous studies implicating the involvement of various kinase pathways in ARE-mediated gene expression have failed to identify any cellular kinase targets (26,27,31). To ascertain whether Nrf2 can be phosphorylated in vivo, we performed metabolic labeling of HepG2 cells with [ 32 P]orthophosphate, followed by immunoprecipitation with an anti-Nrf2 antibody of the cell lysates.…”

Section: Phosphorylation Of Nrf2 In Vivomentioning

confidence: 99%

Regulation of the antioxidant response element by protein kinase C-mediated phosphorylation of NF-E2-related factor 2

Huang¹,

Nguyen²,

Pickett³

2000

Proc. Natl. Acad. Sci. U.S.A.

474

308

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A coordinated cellular response to oxidative stress occurs in part through transcriptional regulation via a cis-acting sequence known as the antioxidant response element (ARE). NF-E2-related factor 2 (Nrf2), a member of the Cap'n'Collar family of basic region-leucine zipper (bZIP) transcription factors, has been implicated as an essential component of an ARE-binding transcriptional complex, but the signaling pathway leading to its activation has remained unclear. Using a reporter gene assay, we found that ARE-directed transcription was activated by phorbol 12-myristate 13-acetate (PMA), but completely suppressed by staurosporine and Ro-32–0432, selective inhibitors of protein kinase C (PKC). Immunocytochemistry and subcellular fractionation revealed that PMA, like tert -butylhydroquinone (tBHQ), promoted the nuclear localization of Nrf2, a process that was blocked by staurosporine or Ro-32–0432. We showed that Nrf2, a previously unidentified kinase target, was phosphorylated in HepG2 cells. PMA transiently activated Nrf2 phosphorylation, whereas the addition of tBHQ or β-naphthoflavone (βNF) led to a persistent stimulation, which was abolished by staurosporine, but not by U0126 and SB203580, respective inhibitors of MEK and p38 kinases. Purified Nrf2 was phosphorylated in vitro by the catalytic subunit of PKC, or by PKC immunoprecipitated from cell lysates. Significantly, PKC precipitated from tBHQ- or βNF-treated cells showed enhanced activity against Nrf2. These findings indicate an important role of the PKC pathway in the ARE-mediated gene expression, and suggest that PKC-directed phosphorylation of Nrf2 may be a critical event for the nuclear translocation of this transcription factor in response to oxidative stress.

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“…Another potential mediator of this survival pathway is the mitogen-activated protein kinase (MAPK) as it was reported that dicumarol blocks MAPK activation in HeLa cells (Cross et al, 1999) and inhibition of the MAPK cascade sensitizes HeLa cells to Fas-induced apoptosis (Holmstrom et al, 1999). Interestingly, extracellular-regulated kinase (ERK), p38 and c-Jun N-terminal kinase (JNK) have been implicated in ARE activation in a complex network that suggest ARE specificity, inducer specificity and cell type/tissue specificity (Yu et al, 1999(Yu et al, , 2000a.…”

Section: Discussionmentioning

confidence: 99%

Nrf2 is an inhibitor of the Fas pathway as identified by Achilles' Heel Method, a new function-based approach to gene identification in human cells

et al. 2003

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Here we describe the Achilles' Heel Method (AHM), a new function-based approach for identification of inhibitors of signaling pathways, optimized for human cells. The principle of AHM is the identification of 'sensitizing' cDNAs based on their decreased abundance following selection. As a proof of principle, we have employed AHM for the identification of Fas/CD95/APO-1 pathway inhibitors. HeLa cells were transfected with an antisense cDNA expression library in an episomal vector followed by selection with a suboptimal dose of the apoptotic inducer. Antisense inactivation of Fas inhibitors rendered the cells more sensitive to apoptosis resulting in their preferential death and consequent loss of their sensitizing episomes that were identified by subtraction. We show that the resulting products were enriched for sensitizing cDNAs as seven out of eight candidates tested were confirmed as inhibitors of Fas-induced killing either by transfection or by pharmacological inhibition. Furthermore, we demonstrate by multiple approaches that one candidate, NF-E2 related factor 2 (Nrf2), is an inhibitor of Fas-induced apoptosis. Inactivation of Nrf2 by antisense or by a membrane permeable dominantnegative polypeptide sensitized cells while overexpression of Nrf2 protected cells from Fas-induced apoptosis. In addition, dicumarol, an inhibitor of the phase II detoxifying enzyme NQO1, a downstream target of Nrf2, sensitized cells. Nrf2 induces the production of Glutathione (GSH) and we demonstrated that N-acetyl L-cysteine (NAC), a precursor to GSH, protected cells from Fas-mediated killing. Taken together, AHM is a powerful approach for the identification of inhibitors of a signaling pathway with a low rate of false positives that opens new avenues for function profiling of human genes and discovery of new drug targets.

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Role of a Mitogen-activated Protein Kinase Pathway in the Induction of Phase II Detoxifying Enzymes by Chemicals

Cited by 267 publications

References 45 publications

Increased Protein Stability as a Mechanism That Enhances Nrf2-mediated Transcriptional Activation of the Antioxidant Response Element

Increased Protein Stability as a Mechanism That Enhances Nrf2-mediated Transcriptional Activation of the Antioxidant Response Element

Regulation of the antioxidant response element by protein kinase C-mediated phosphorylation of NF-E2-related factor 2

Nrf2 is an inhibitor of the Fas pathway as identified by Achilles' Heel Method, a new function-based approach to gene identification in human cells

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