Printer Friendly

Possible involvement of granulocyte oxidative burst in Nrf2 signaling in cancer.

Respiratory burst of granulocytes

While it is well known that polymorphonuclear leukocytes play essential role in host defence against microorganisms (1), mechanisms by which granulocytes may be involved in the immune response against cancer are not well understood. Recently, it was reported that spontaneous regression/complete resistance to cancer cells is mediated by rapid infiltration of leukocytes, mostly as a consequence of innate immune response (2,3). In addition, the administration of granulocytes at the site of solid tumors can lead to tumor regression or can slower tumor growth and extend the overall survival of animals (4). The activation process of granulocytes is accompanied by the intense production of reactive oxygen species (ROS) (1,5) resulting in oxidative stress (6). ROS are normally formed in small quantities during metabolic processes (7,8). The level of cellular ROS that induces initial effects leading to protection of cells is only slightly lower than the levels that cause awful effects. Overproduction and accumulation of ROS is cytotoxic and damages macromolecules (DNA, proteins, sugars and lipids) (9,10), causing degeneration of tissues, premature aging, apoptosis, cellular transformation, mutagenicity, and cancer (11). Both prokaryotic and eukaryotic cells respond to this toxicity by the protection mechanisms coordinately inducing a series of genes encoding detoxifying and antioxidative stress enzymes/proteins that provide the necessary protection against oxidative and electrophilic stress (12,13). In eukaryotes, there are transcription factors known to be activated by ROS. Those transcription factors include NF-E2-related factors (Nrfs) (12). This review focuses on Nrf2 that regulates coordinated activation of a variety of genes in response to oxidative stress.

Nuclear factor-erythroid 2 (NF-E2)

NF-E2 belongs to the bZip (basic region leucine zipper) transcription factors. NF-E2 was purified from mouse erythroleukemia (MEL) cells and is composed of 45 kDa and 18 kDa subunit (14). NF-E2 is expressed only in erythroid cells, megakaryocytes, and mast cells. It binds to NF-E2 recognition site (GCTGAGTCA)

and regulates tissue specific expression of the globin genes (15,16). NF-E2 functions as a heterodimer with small Maf proteins that are ubiquitously expressed (17). Maf proteins are a family of nuclear transcription factors. Maf family constitutes from several genes (v-maf, c-maf and other v-maf related genes) and can be divided into large and small Maf subfamilies. They can act both as activators and repressors of a number of eukaryotic genes (18-20).

NF-E2 related factors (Nrfs)

Screening for a molecule that can interact with the NF-E2 binding motif led to the identification of NF-E2 related factors (Nrf1, Nrf2, and Nrf3) (21). All of these factors have highly conserved cap'n'colar (CNC) region, bZip region, basic and acidic region (21,22). It is known that the basic region is responsible for DNA binding, and that the acidic region is required for transcriptional activation. Many researchers have demonstrated Nrf2 to be the most prominent factor in activation and induction of antioxidant responsive element (ARE)- mediated genes in comparison with Nrf1 and Nrf3 (23,24).

Nrf2, a 68 kDa protein (21), consists of six highly conserved domains, Neh1-6. Few years ago Li et al characterized in Nrf2 a nuclear export signal (NES) in the leucine zipper domain (NESzip) (25) and in the Neh5 transactivation domain ([NES.sub.TA]) (26). A nuclear localization signal (NLS) was also identified as a bipartite NLS in the basic region (bNLS) of Nrf2 (25,27), or a monopartite at the N-terminus ([NLS.sub.N]) and C-terminus ([NLS.sub.C]) of Nrf2 (28). Based on the function of multiple Nrf2 NES/NLS motifs in the non-stimulated and oxidative stress conditions, Li et al proposed a new Keap1-independent Nrf2 signaling model (26,29). In this model, Nrf2 is a redox sensitive and consists of a constitutively active NESzip bNLS-[NLS.sub.N]-[NLS.sub.C] tandem and a conditional [NES.sub.TA] motif (28). Oxidative stress signals influence the reactive Cys in the [NES.sub.TA] and disable the [NES.sub.TA] resulting in Nrf2 nuclear translocation (29).

The Keap1-dependant Nrf2 signaling will be discussed later.

Detoxyfying enzimes and antioxidant responsive element

Against damage from ROS and xenobiotics, cells and tissues defend by enhancement of detoxifying enzymes. The generation of superoxides and electrophiles is a result of antioxidants and xenobiotics metabolism. There are two major groups of metabolizing enzymes, phase I (activation) and phase II (detoxifying) metabolizing enzymes. Phase I enzymes include oxidation, reduction, and hydrolysis. Throughout phase I detoxication structurally diversified chemicals are, with cytochrome P450 mono-oxygenase system, oxidated into their metabolically activated forms. Phase II include glucuronidation, sulfation, acetylation, methylation and conjugation with glutathione. Phase II detoxicating enzymes are induced by phase I metabolites as well as by antioxidants, which are very often electrophilic. Throughout phase II detoxification, the activated substrates are catalysed into nontoxic metabolites (30).

Phase II enzyme genes are induced through the ARE (31). This important regulatory element was also called the electrophile responsive element (EpRE) (32). The ARE was identified in the regulatory region of some Genes (33-36), and was also recognized as a DNA element that regulates basal expression and coordinated induction of genes encoding antioxidant enzymes in response to antioxidants and xenobiotics (12). The ARE sequence plays an essential role in the regulation of the cellular defense system. The minimum ARE core sequence was reported to be 5'-TGACNNNGC-3'. Other neighboring sequences and elements also affect ARE-mediated expression and induction of antioxidant genes (37-39).

Nrf2 plays an important role in the resistance to xenobiotic toxicity by the regulation of ARE-mediated gene expression (40,41). In response to antioxidants, xenobiotics, metals, and UV irradiation, Nrf2 protein binds strongly to ARE sequence and regulates ARE-mediated antioxidant enzyme gene expression and induction (13,42,43).

In the interaction between Nrf2 and ARE sequence, Nrf2 requires other cofactors, by forming heterodimer with a member of the small Maf proteins (MafK and MafG) (41,44). Dimerization with one of the small Maf proteins (transactivation) allows Nrf2 binding to the ARE. Some studies have shown the role of Nrf2-MafK heterodimers in the activation of ARE-mediated gene expression (41).

Keap1 acts as a negative regulator of Nrf2 (Keap1Nrf2 complex)

Keap1 (Kelch-like ECH-associated protein 1) or INrf2 (inhibitor of Nrf2) is a cytosolic inhibitor of Nrf2 (45,46). Keap1 is a dimer and retains Nrf2 in the cytoplasm. Keap1 was identified as a direct binding partner of Nrf2, using the Neh2 domain as a bait (45). N-terminal Neh2 domain is necessary for the recruitment of Nrf2 negative regulator (Keap1). Keap1 consists of five domains. One of them is Kelch domain or the double glycine repeat (DGR) domain, through which Keap1 binds Neh2 and actin (45,47). Normally, Nrf2 associates with Keap1 in the cytoplasm and represses Nrf2 transactivation activity (45). After direct attack by electrophiles or ROS or indirect actions such as phosphorylation, Nrf2 is released from Keap1 repression, translocated into nucleus where it activates the transcription of a variety of detoxifying enzyme genes through the ARE45. After dissociation from Nrf2, Keap1 remains in the cytosol (46). Under basal conditions, Nrf2 is present at low concentrations due to CUL3-dependent E3-ubiquitin ligase mediated ubiquitination of Nrf2 (48). Mutational studies revealed three Keap1 cysteine residues (Cys 151, Cys 273 and Cys 288) (49) that are essential for degrading Nrf2 by Keap1-mediated ubiquitination. Nrf2 dissociation and Keap1 ubiquitination, mediated by electrophiles or ROS, are abolished by the mutation of Cys 15149. This finding was further verified in vivo (50). The mechanisms by which Cys 273 and Cys 288 affect Keap1 function are not well understood (51). Recently, Chen et al (52) described that interaction between Nrf2 and p21 interferes with Keap1-Nrf2 binding and thus upregulates Nrf2-mediated antioxidant response. The antioxidant function of p21, mediated through activation of Nrf2 by stabilizing the Nrf2 protein, was further verified by in vivo studies using p21-deficient mice (52).

Several reports have shown that Nrf2 binding to Keap1 leads to degradation of Nrf2 (53-56). After Nrf2 is released from Keap1, it has a strong transactivation potential such as before binding with Keap1, which suggests that any modification of Nrf2 is only required for the release of Nrf2 from Keap1 (56). Under oxidative stress conditions, besides Keap1-dependent degradation, Nrf2 can be also degraded by proteosomal degradation (Keap1-independent degradation) (55,57).

There is an autoregulatory loop, Nrf2:INrf2 (Keap1), between stress sensors Keap1 and Nrf2 in the Nrf2 pathway (58). Namely, ARE that binds to Nrf2 regulates Keap1 gene expression and induction and Keap1 controls Nrf2 by serving as an adaptor for Degradation (59).

The involvement of MAPKs, PKC, PI3K and GSK3[beta] in the activation of Nrf2

The mitogen-activated protein kinases (MAPK) are important cellular signaling components that transform various extracellular signals into intracellular responses through phosphorylation cascades. MAPK pathways that are activated by MEKK1, TAK1, and ASK1 may link chemical signals to Nrf2 which then leads to the activation of ARE-dependent genes (60). The correlation between MAPK activation and Nrf2-mediated detoxifying enzyme suggests that Nrf2 could be a downstream target of activated MAPKs. If MAPK are inhibited prior to exposure to detoxifying enzyme inducers, Nrf2 accumulation is decreased (54) and Nrf2 translocation to the nucleus is reduced (61). Recently, it was reported that chemosensitivity of lung cancer cells to anti-cancer agents may be modulated by HO-1 through the MAPK-Nrf2 pathway (62).

The importance of protein kinase C (PKC) is in phosphorylation of many target proteins that control cell growth and differentiation. A classic PKC activator can activate ARE-mediated gene expression and this activation can be inhibited by PKC inhibitors (63). Posttranslational modification of Nrf2 may also represent an important regulatory step in the activation of Nrf2, for instance, phosphorylation of Nrf2 (63). PKC phosphorylation site in Nrf2 protein is Ser-40 (64) and is located in the Keap1-interacting Neh2 domain. The phosphorylation of this site by PKC disrupts the interaction of Nrf2 with Keap1 leading to Nrf2 release from Keap1-Nrf2 complex. This phosphorilation is not necessary for Nrf2 stabilization or transcriptional activation of ARE-mediated gene expression (56,64).

Phosphatidylinositol 3-kinase (PI3K) is important in cell growth, differentiation and apoptosis. PI3K is involved in the regulation of ARE and detoxifying enzymes: when PI3K is inhibited, ARE reporter expression is also inhibited. Overexpression of constitutively active PI3K always leads to the activation of ARE activity in a dose dependent manner. This upregulatory effect was completely blocked by dominant-negative Nrf2 (65). A partial explanation of PI3K function in the cytoprotection machinery is that PI3K is involved in the regulation of nuclear translocation of Nrf2 protein (65,66). PI3K regulates Nrf2 through actin rearrangement in response to oxidative stress (67).

A delayed response of electrophilic/oxidative stress activates glycogen synthase kinase-3 beta (GSK-3[beta]) that acts upstream of Fyn kinase controlling the nuclear export of Nrf2 (68). Phosphorilation of Fyn, mediated by GSK-3p\ is followed by phosphorilated Fyn translocation into the nucleus where it exerts its activity by phosphorilating Nrf2 at Tyr 568 thus leading to nuclear export and degradation of Nrf2 (68). GSK-3[beta] was shown to be essential in the down-regulation of the antioxidant cell defense elicited by Nrf2 after oxidant injury (69).

Nrf2 in cancer prevention/promotion

Many studies have described the role of Nrf2 in cancer prevention. Numerous chemopreventive compounds have been identified as Nrf2 inducers (e.g., sulforaphane) (70) and their list is continuously growing. After their application, Nrf2-dependant adaptive response is induced, thus exerting its protective role from genotoxic damage caused by carcinogens (71). The role of Nrf2 in cancer prevention was confirmed by in vitro studies on Nrf2-null mice that had enhanced sensitivity to carcinogens in comparison to wild-type mice (72-75). The incidence and multiplicity of tumors as well as the tumor volume were enhanced in Nrf2-null mice (73-78). However, in the last few years there are papers describing the tumor promoting role of Nrf2 proposing its dual role in cancer (71). Nrf2 overexpression was found in the later stages of cancer (79). Furthermore, Nrf2 was found to be upregulated in hepatocellular Carcinoma (80). Somatic mutations of Keap1 (81-83) and low level expressions of Keap1 (82) were identified in lung cancer tissues and cell lines, leading to enhanced nuclear accumulation and constitutive activation of Nrf2. These results suggest that elevated Nrf2 could play a role in the evolution of cancer (84). Moreover, Shibata et al found Keap1 gene alterations in biliary tract and gallbladder cancer (85). These alterations were especially frequent in gallbladder cancer leading to a loss of Nrf2 repression activity and constitutive activation of Nrf2 (85). Finally, Keap1 mutation that impairs its ability for Nrf2 repression was also found in breast cancer (86). However, another recently published study revealed that most of breast cancer specimens examined had high Cul3/low Nrf2 signature (87) that can significantly contribute to increased cellular sensitivity to chemotherapeutic drugs. Nevertheless, to date, there is no evidence suggesting that the compounds used to induce Nrf2 signaling pathway have adverse impacts on tumor growth (78).

The involvement of oxidative stress in Nrf2 signaling pathway in an early stage of cancer is discussed below.

The involvement of granulocyte respiratory burst in Nrf2 pathway in cancer

ROS are reported to have an important role in tumor biology (88). Persistent oxidative stress may activate antioxidant systems, constitutively activate transcription factors and induce expression of proto-oncogenes. Furthermore, it may lead to genomic instability and facilitate tumor invasion and metastasis (88). Previously we have reported the involvement of oxidative stress in tumor progression (89,90) or regression (3). We have shown that respiratory burst of granulocytes is significantly increased in tumor-bearing animals in an early stage of tumor development. Furthermore, we found that tumor progression is associated with the constant increase in the oxidative burst of granulocytes, whereas in animals with tumor regression the respiratory burst of granulocytes decreased to normal values following the tumor disappearance (3). ROS, present in the high levels in an early stage of tumor development, may consequently influence the induction of nuclear factor-[kappa]B (NF-[kappa]B) intracellular signaling (91) repressing the Nrf2-ARE pathway at transcriptional level (92). NF-[kappa]B is ubiquitously expressed transcription factor mostly present in cytoplasm, bound to its inhibitor-kappa B (I[kappa]B) (93). Activated NF-[kappa]B and its activating signaling pathways regulate cell adhesion, differentiation, growth, apoptosis, malignant transformation and inflammatory response (94). Granulocytes were shown to be the first cells that rapidly infiltrate the site of tumor transplantation thus supporting the possibility that granulocyte response to early stage of cancer is important for spontaneous regression (3). In the presence of tumor cells activated granulocytes produce higher amounts of ROS (89) thus inhibiting the tumor cell proliferation. One of the mechanisms by which oxidative burst of granulocytes may lead to tumor destruction could be by influencing the Nrf2 signaling pathway (Fig.). Namely, tumor cells are more resistant to oxidative stress by the process of antioxidant systems activation. ROS produced by oxidative burst of granulocytes influence the NF-[kappa]B signaling pathway by repressing the Nrf2-ARE pathway and thus leading to malignant destruction. However, further increase in ROS levels may facilitate tumor promotion. Namely, polyunsaturated fatty acids that are esterified in membrane or storage lipids are subject to ROS-induced peroxidation resulting in the destruction of biomembranes. Final products of lipid peroxidation are reactive aldehydes among which is 4-hydroxy-2-nonenal (HNE), denoted the "second toxic messenger of free radicals" (95-97). Lipid-derived aldehydes are more stable than ROS and can therefore diffuse to targets far from the initial oxidative injury. Many deleterious effects have been attributed to HNE (98) such as modulation of cell growth (99,100) differentiation (101) apoptosis (101) and cell signaling (102,103) HNE is also very effective in binding to DNA or proteins leading to adduct formation, eliciting mutagenic or carcinogenic effects (9,104). Finally, HNE is also reported to have a role in Nrf2 signaling by inducing Nrf2 expression (105), which may further support proposed Nrf2 tumor promoting role (70) in the later stages of cancer (79). However, in an early stage of tumor development ROS may be of crucial importance in repressing the Nrf2 pathway thus leading to tumor regression.



The study was supported by Croatian Ministry of Science, Education and Sports.

Received May 8, 2009


(1.) Babior BM. Phagocytes and oxidative stress. Am J Med 2000; 109: 33-44.

(2.) Hicks AM, Riedlinger G, Willingham MC, Alexander-Miller MA, Von Kap-Herr C, Pettenati MJ, et al. Transferable anticancer innate immunity in spontaneous regression/ complete resistance mice. Proc Natl Acad Sci USA 2006; 103: 7753-8.

(3.) Jaganjac M, Poljak-Blazi M, Zarkovic K, Schaur RJ, Zarkovic N. The involvement of granulocytes in spontaneous regression of Walker 256 carcinoma. Cancer Lett 2008; 260: 180-6.

(4.) Jaganjac M, Poljak-Blazi M, Kirac I, Borovic S, Schaur RJ, Zarkovic N. Granulocytes as effective anticancer agent in experimental solid tumor models. Immunobiology 2010; (in press).

(5.) Sbarra AJ, Karnovsky ML. The biochemical basis of phagocytosis. I. Metabolic changes during the ingestion of particles by polymorphonuclear leukocytes. J Biol Chem 1959; 234: 1355-62.

(6.) Sies H, editor. Oxidative stress oxidants and antioxidants. London: Academic Press; 1991. p. 15-22.

(7.) Braughler JM, Hall ED. Central nervous system trauma and stroke. I. Biochemical considerations for oxygen radical formation and lipid peroxidation. Free Radic Biol Med 1989; 6: 289-301.

(8.) Hall ED, Braughler JM. Central nervous system trauma and stroke. II. Physiological and pharmacological evidence for involvement of oxygen radicals and lipid peroxidation. Free Radic Biol Med 1989; 6: 303-13.

(9.) Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal, malonaldehyde and related aldehydes. Free Radic Biol Med 1991; 11: 81-128.

(10.) Venkatesh S, Deecaraman M, Kumar R, Shamsi MB, Dada R. Role of reactive oxygen species in the pathogenesis of mitochondrial DNA (mtDNA) mutations in male infertility. Indian J Med Res 2009; 129: 127-37.

(11.) Breen AP, Murphy JA. Reactions of oxyl radicals with DNA. Free Radic Biol Med 1995; 18: 1033-77.

(12.) Talalay P, Dinkova-Kostova AT, Holtzclaw WD. Importance of phase 2 gene regulation in protection against electrophile and reactive oxygen toxicity and carcinogenesis. Adv Enzyme Regul 2003; 43: 121-34.

(13.) Jaiswal AK. Regulation of genes encoding NAD(P)H:quinone oxidoreductases. Free Radic Biol Med 2000; 29: 254-62.

(14.) Andrews NC, Erdjument-Bromage H, Davidson MB, Tempst P, Orkin SH. Erythroid transcription factor NF-E2 is a haematopoietic-specific basic-leucine zipper protein. Nature 1993; 362: 722-8.

(15.) Liu D, Chang JC, Moi P, Liu W, Kan YW, Curtin PT. Dissection of the enhancer activity of beta-globin 5' D-Nase I-hypersensitive site 2 in transgenic mice. Proc Natl Acad Sci USA 1992; 89: 3899-903.

(16.) Mignotte V, Eleouet JF, Raich N, Romeo PH. Cis- and transacting elements involved in the regulation of the erythroid promoter of the human porphobilinogen deaminase gene. Proc Natl Acad Sci USA 1989; 86: 6548-52.

(17.) Igarashi K, Kataoka K, Itoh K, Hayashi N, Nishizawa M, Yamamoto M. Regulation of transcription by dimerization of erythroid factor NF-E2 p45 with small Maf proteins. Nature 1994; 367: 568-72.

(18.) Kataoka K, Nishizawa M, Kawai S. Structure-function analysis of the maf oncogene product, a member of the b-Zip protein family. J Virol 1993; 67: 2133-41.

(19.) Kataoka K, Noda M, Nishizawa M. Maf nuclear oncoprotein recognizes sequences related to an AP-1 site and forms heterodimers with both Fos and Jun. Mol Cell Biol 1994; 14: 700-12.

(20.) Blank V, Kim MJ, Andrews NC. Human MafG is a functional partner for p45 NF-E2 in activating globin gene expression. Blood 1997; 89: 3925-35.

(21.) Moi P, Chan K, Asunis I, Cao A, Kan YW. Isolation of NF E2-related factor 2 (Nrf2), a NF-E2-like basic leucine zipper transcriptional activator that binds to the tandem NF-E2/AP1 repeat of the beta-globin locus control region. Proc Natl Acad Sci USA 1994; 91: 9926-30.

(22.) Motohashi H, O'Connor T, Katsuoka F, Engel JD, Yamamoto M. Integration and diversity of the regulatory network composed of Maf and CNC families of transcription factors. Gene 2002; 294: 1-12.

(23.) Zhang DD. Mechanistic studies of the Nrf2-Keap1 signaling pathway. Drug Metab Rev 2006; 38: 769-89.

(24.) Copple IM, Goldring CE, Kitteringham NR, Park BK. The Nrf2-Keap1 defence pathway: role in protection against drug-induced toxicity. Toxicology 2008; 246: 24-33.

(25.) Li W, Jain MR, Chen C, Yue X, Hebbar V, Zhou R, et al. Nrf2 Possesses a redox-insensitive nuclear export signal overlapping with the leucine zipper motif. J Biol Chem 2005; 280: 28430-8.

(26.) Li W, Yu SW, Kong AN. Nrf2 possesses a redox-sensitive nuclear exporting signal in the Neh5 transactivation domain. J Biol Chem 2006; 281: 27251-63.

(27.) Jain AK, Bloom DA, Jaiswal AK. Nuclear import and export signals in control of Nrf2. J Biol Chem 2005; 280: 29158-68.

(28.) Theodore M, Kawai Y, Yang J, Kleshchenko Y, Reddy SP, Villalta F, et al. Multiple nuclear localization signals function in the nuclear import of the transcription factor Nrf2. J Biol Chem 2008; 283: 8984-94.

(29.) Li W, Kong AN. Molecular mechanisms of Nrf2-mediated antioxidant response. Mol Carcinog 2009; 48: 91-104.

(30.) Chen C, Kong AN. Dietary chemopreventive compounds and ARE/EpRE signaling. Free Radic Biol Med 2004; 36: 1505-16.

(31.) Hayes JD, McMahon M. Molecular basis for the contribution of the antioxidant responsive element to cancer chemoprevention. Cancer Lett 2001; 174: 103-13.

(32.) Friling RS, Bensimon A, Tichauer Y, Daniel V. Xenobiotic-inducible expression of murine glutathione S-transferase Ya subunit gene is controlled by an electrophile-responsive element. Proc Natl Acad Sci USA 1990; 87: 6258-62.

(33.) Sherratt PJ, McLellan LI, Hayes JD. Positive and negative regulation of prostaglandin E2 biosynthesis in human colorectal carcinoma cells by cancer chemopreventive agents. Biochem Pharmacol 2003; 66: 51-61.

(34.) Sasaki H, Sato H, Kuriyama-Matsumura K, Sato K, Maebara K, Wang H, et al. Electrophile response element-mediated induction of the cystine/glutamate exchange transporter gene expression. J Biol Chem 2002; 277: 44765-71.

(35.) Zhang X, Jiao JJ, Bhavnani BR, Tam SP. Regulation of human apolipoprotein A-I gene expression by equine estrogens. J Lipid Res 2001; 42: 1789-800.

(36.) Miller KP, Chen YH, Hastings VL, Bral CM, Ramos KS. Profiles of antioxidant/electrophile response element (ARE/ EpRE) nuclear protein binding and c-Ha-ras transactivation in vascular smooth muscle cells treated with oxidative metabolites of benzo[a]pyrene. Biochem Pharmacol 2000; 60: 1285-96.

(37.) Li Y, Jaiswal AK. Regulation of human NAD(P)H:quinone oxidoreductase gene. Role of AP1 binding site contained within human antioxidant response element. J Biol Chem 1992; 267: 15097-104.

(38.) Prestera T, Holtzclaw WD, Zhang Y, Talalay P. Chemical and molecular regulation of enzymes that detoxify carcinogens. Proc Natl Acad Sci USA 1993; 90: 2965-9.

(39.) Wasserman WW, Fahl WE. Functional antioxidant responsive elements. Proc Natl Acad Sci USA 1997; 94: 5361-6.

(40.) Ishii T, Itoh K, Takahashi S, Sato H, Yanagawa T, Katoh Y, et al. Transcription factor Nrf2 coordinately regulates a group of oxidative stress-inducible genes in macrophages. J Biol Chem 2000; 275: 16023-9.

(41.) Itoh K, Chiba T, Takahashi S, Ishii T, Igarashi K, Katoh Y, et al. An Nrf2/small Maf heterodimer mediates the induction of phase II detoxifying enzyme genes through antioxidant response elements. Biochem Biophys Res Commun 1997; 236: 313-22.

(42.) Alam J, Stewart D, Touchard C, Boinapally S, Choi AM, Cook JL. Nrf2, a Cap'n'Collar transcription factor, regulates induction of the heme oxygenase-1 gene. Biol Chem 1999; 274: 26071-8.

(43.) Wild AC, Moinova HR, Mulcahy RT. Regulation of gammaglutamylcysteine synthetase subunit gene expression by the transcription factor Nrf2. J Biol Chem 1999; 274: 33627-36.

(44.) Kobayashi A, Ito E, Toki T, Kogame K, Takahashi S, Igarashi K, et al. Molecular cloning and functional characterization of a new Cap'n' collar family transcription factor Nrf3. J Biol Chem 1999; 274: 6443-52.

(45.) Itoh K, Wakabayashi N, Katoh Y, Ishii T, Igarashi K, Engel JD, et al. Keap1 represses nuclear activation of antioxidant responsive elements by Nrf2 through binding to the aminoterminal Neh2 domain. Genes Dev 1999; 13: 76-86.

(46.) Dhakshinamoorthy S, Jaiswal AK. Functional characterization and role of INrf2 in antioxidant response element-mediated expression and antioxidant induction of NAD(P)H:quinone oxidoreductase1 gene. Oncogene 2001; 20: 3906-17.

(47.) Kang MI, Kobayashi A, Wakabayashi N, Kim SG, Yamamoto M. Scaffolding of Keap1 to the actin cytoskeleton controls the function of Nrf2 as key regulator of cytoprotective phase 2 genes. Proc Natl Acad Sci USA 2004; 101: 2046-51.

(48.) Zhang DD, Lo SC, Cross JV, Templeton DJ, Hannink M. Keap1 is a redox-regulated substrate adaptor protein for a Cul3-dependent ubiquitin ligase complex. Mol Cell Biol 2004; 24: 10941-53.

(49.) Zhang DD, Hannink M. Distinct cysteine residues in Keap1 are required for Keap1-dependent ubiquitination of Nrf2 and for stabilization of Nrf2 by chemopreventive agents and oxidative stress. Mol Cell Biol 2003; 23: 8137-51.

(50.) Yamamoto T, Suzuki T, Kobayashi A, Wakabayashi J, Maher J, Motohashi H, et al. Physiological significance of reactive cysteine residues of Keap1 in determining Nrf2 activity. Mol Cell Biol 2008; 28: 2758-70.

(51.) Sekhar KR, Rachakonda G, Freeman ML. Cysteine-based regulation of the CUL3 adaptor protein Keap1. Toxicol Appl Pharmacol 2010; 244: 21-6.

(52.) Chen W, Sun Z, Wang XJ, Jiang T, Huang Z, Fang D, et al. Direct interaction between Nrf2 and p21(Cip1/WAF1) upregulates the Nrf2-mediated antioxidant response. Mol Cell 2009; 34: 663-73.

(53.) Stewart D, Killeen E, Naquin R, Alam S, Alam J. Degradation of transcription factor Nrf2 via the ubiquitin-proteasome pathway and stabilization by cadmium. J Biol Chem 2003; 278: 2396-402.

(54.) Nguyen T, Sherratt PJ, Huang HC, Yang CS, Pickett CB. Increased protein stability as a mechanism that enhances Nrf2mediated transcriptional activation of the antioxidant response element. Degradation of Nrf2 by the 26 S proteasome. J Biol Chem 2003; 278: 4536-41.

(55.) Itoh K, Wakabayashi N, Katoh Y, Ishii T, O'Connor T, Yamamoto M. Keap1 regulates both cytoplasmic-nuclear shuttling and degradation of Nrf2 in response to electrophiles. Genes Cells 2003; 8: 379-91.

(56.) Bloom DA, Jaiswal AK. Phosphorylation of Nrf2 at Ser40 by protein kinase C in response to antioxidants leads to the release of Nrf2 from INrf2, but is not required for Nrf2 stabilization/ accumulation in the nucleus and transcriptional activation of antioxidant response element-mediated NAD(P)H:quinone oxidoreductase-1 gene expression. J Biol Chem 2003; 278: 44675-82.

(57.) McMahon M, Itoh K, Yamamoto M, Hayes JD. Keap1-dependent proteasomal degradation of transcription factor Nrf2 contributes to the negative regulation of antioxidant response element-driven gene expression. J Biol Chem 2003; 278: 21592-600.

(58.) Lee OH, Jain AK, Papusha V, Jaiswal AK. An auto-regulatory loop between stress sensors INrf2 and Nrf2 controls their cellular abundance. J Biol Chem 2007; 282: 36412-20.

(59.) Kaspar JW, Niture SK, Jaiswal AK. Nrf2:INrf2 (Keap1) signaling in oxidative stress. Free Radic Biol Med 2009; 47: 1304-9.

(60.) Yu R, Chen C, Mo YY, Hebbar V, Owuor ED, Tan TH, et al. Activation of mitogen-activated protein kinase pathways induces antioxidant response element-mediated gene expression via a Nrf2-dependent mechanism. J Biol Chem 2000; 275: 39907-13.

(61.) Zipper LM, Mulcahy RT. Erk activation is required for Nrf2 nuclear localization during pyrrolidine dithiocarbamate induction of glutamate cysteine ligase modulatory gene expression in HepG2 cells. Toxicol Sci 2003; 73: 124-34.

(62.) Kim HR, Kim S, Kim EJ, Park JH, Yang SH, Jeong ET, et al. Suppression of Nrf2-driven heme oxygenase-1 enhances the chemosensitivity of lung cancer A549 cells toward cisplatin. Lung Cancer 2008; 60: 47-56.

(63.) Huang HC, Nguyen T, Pickett CB. Regulation of the antioxidant response element by protein kinase C-mediated phosphorylation of NF-E2-related factor 2. Proc Natl Acad Sci USA 2000; 97: 12475-80.

(64.) Huang HC, Nguyen T, Pickett CB. Phosphorylation of Nrf2 at Ser-40 by protein kinase C regulates antioxidant response element-mediated transcription. J Biol Chem 2002; 277: 42769-74.

(65.) Lee JM, Moehlenkamp JD, Hanson JM, Johnson JA. Nrf2-dependent activation of the antioxidant responsive element by tert-butylhydroquinone is independent of oxidative stress in IMR-32 human neuroblastoma cells. Biochem Biophys Res Commun 2001; 280: 286-92.

(66.) Nakaso K, Yano H, Fukuhara Y, Takeshima T, Wada-Isoe K, Nakashima K. PI3K is a key molecule in the Nrf2-mediated regulation of antioxidative proteins by hemin in human neuroblastoma cells. FEBSLett 2003; 546: 181-4.

(67.) Kang KW, Lee SJ, Park JW, Kim SG. Phosphatidylinositol 3-kinase regulates nuclear translocation of NF-E2-related factor 2 through actin rearrangement in response to oxidative stress. Mol Pharmacol 2002; 62: 1001-10.

(68.) Jain AK, Jaiswal AK. GSK-3beta acts upstream of Fyn kinase in regulation of nuclear export and degradation of NF-E2 related factor 2. J Biol Chem 2007; 282: 16502-10.

(69.) Rojo AI, Sagarra MR, Cuadrado A. GSK-3beta down-regulates the transcription factor Nrf2 after oxidant damage: relevance to exposure of neuronal cells to oxidative stress. J Neurochem 2008; 105: 192-202.

(70.) Dinkova-Kostova AT, Holtzclaw WD, Cole RN, Itoh K, Wakabayashi N, Katoh Y, et al. Direct evidence that sulfhydryl groups of Keap1 are the sensors regulating induction of phase 2 enzymes that protect against carcinogens and oxidants. Proc Natl Acad Sci USA 2002; 99: 11908-13.

(71.) Lau A, Villeneuve NF, Sun Z, Wong PK, Zhang DD. Dual roles of Nrf2 in cancer. Pharmacol Res 2008; 58: 262-70.

(72.) Ramos-Gomez M, Kwak MK, Dolan PM, Itoh K, Yamamoto M, Talalay P, et al. Sensitivity to carcinogenesis is increased and chemoprotective efficacy of enzyme inducers is lost in nrf2 transcription factor-deficient mice. Proc Natl Acad Sci USA 2001; 98: 3410-5.

(73.) Iida K, Itoh K, Kumagai Y, Oyasu R, Hattori K, Kawai K, et al. Nrf2 is essential for the chemopreventive efficacy of oltipraz against urinary bladder carcinogenesis. Cancer Res 2004; 64: 6424-31.

(74.) Iida K, Itoh K, Maher JM, Kumagai Y, Oyasu R, Mori Y, et al. Nrf2 and p53 cooperatively protect against BBN-induced urinary bladder carcinogenesis. Carcinogenesis 2007; 28: 2398-403.

(75.) Khor TO, Huang MT, Prawan A, Liu Y, Hao X, Yu S, et al. Increased susceptibility of Nrf2 knockout mice to colitis-associated colorectal cancer. Cancer Prev Res (Phila Pa) 2008; 1: 187-91.

(76.) Osburn WO, Karim B, Dolan PM, Liu G, Yamamoto M, Huso DL, et al. Increased colonic inflammatory injury and formation of aberrant crypt foci in Nrf2-deficient mice upon dextran sulfate treatment. Int J Cancer 2007; 121: 1883-91.

(77.) Xu C, Huang MT, Shen G, Yuan X, Lin W, Khor TO, et al. Inhibition of 7,12-dimethylbenz(a)anthracene-induced skin tumorigenesis in C57BL/6 mice by sulforaphane is mediated by nuclear factor E2-related factor 2. Cancer Res 2006; 66: 8293-6.

(78.) Kwak MK, Kensler TW. Targeting NRF2 signaling for cancer chemoprevention. Toxicol Appl Pharmacol 2010; 244: 66-76.

(79.) Wang XJ, Sun Z, Villeneuve NF, Zhang S, Zhao F, Li Y, et al. Nrf2 enhances resistance of cancer cells to chemotherapeutic drugs, the dark side of Nrf2. Carcinogenesis 2008; 29: 1235-43.

(80.) Ikeda H, Nishi S, Sakai M. Transcription factor Nrf2/MafK regulates rat placental glutathione S-transferase gene during hepatocarcinogenesis. Biochem J 2004; 380: 515-21.

(81.) Padmanabhan B, Tong KI, Ohta T, Nakamura Y, Scharlock M, Ohtsuji M, et al. Structural basis for defects of Keap1 activity provoked by its point mutations in lung cancer. Mol Cell 2006; 21: 689-700.

(82.) Ohta T, Iijima K, Miyamoto M, Nakahara I, Tanaka H, Ohtsuji M, et al. Loss of Keap1 function activates Nrf2 and provides advantages for lung cancer cell growth. Cancer Res 2008; 68: 1303-9.

(83.) Singh A, Misra V, Thimmulappa RK, Lee H, Ames S, Hoque MO, et al. Dysfunctional KEAP1-NRF2 interaction in non small-cell lung cancer. PLoS Med 2006; 3: e420.

(84.) Hayes JD, McMahon M. The double-edged sword of Nrf2: subversion of redox homeostasis during the evolution of cancer. Mol Cell 2006; 21: 732-4.

(85.) Shibata T, Kokubu A, Gotoh M, Ojima H, Ohta T, Yamamoto M, et al. Genetic alteration of Keap1 confers constitutive Nrf2 activation and resistance to chemotherapy in gallbladder cancer. Gastroenterology 2008; 135: 1358-68.

(86.) Nioi P, Nguyen T. A mutation of Keap1 found in breast cancer impairs its ability to repress Nrf2 activity. Biochem Biophys Res Commun 2007; 362: 816-21.

(87.) Loignon M, Miao W, Hu L, Bier A, Bismar TA, Scrivens PJ, et al. Cul3 overexpression depletes Nrf2 in breast cancer and is associated with sensitivity to carcinogens, to oxidative stress, and to chemotherapy. Mol Cancer Ther 2009; 8: 2432-40.

(88.) Toyokuni S, Okamoto K, Yodoi J, Hiai H. Persistent oxidative stress in cancer. FEBSLett 1995; 358: 1-3.

(89.) Zivkovic M, Poljak-Blazi M, Egger G, Sunjic SB, Schaur RJ, Zarkovic N. Oxidative burst and anticancer activities of rat neutrophils. Biofactors 2005; 24: 305-12.

(90.) Zivkovic M, Poljak-Blazi M, Zarkovic K, Mihaljevic D, Schaur RJ, Zarkovic N. Oxidative burst of neutrophils against melanoma B16-F10. Cancer Lett 2007; 246: 100-8.

(91.) Poljak-Blazi M, Jaganjac M, Mustapic M, Pivac N, Muck-Seler D. Acute immunomodulatory effects of iron polyisomaltosate in rats. Immunobiology 2009; 214: 121-8.

(92.) Liu GH, Qu J, Shen X. NF-KB/p65 antagonizes Nrf2ARE pathway by depriving CBP from Nrf2 and facilitating recruitment of HDAC3 to MafK. Biochim BiophysActa 2008; 1783: 713-27.

(93.) Ghosh S, May MJ, Kopp EB. NF-kappa B and Rel proteins: evolutionarily conserved mediators of immune responses. Annu Rev Immunol 1998; 16: 225-60.

(94.) Chen F, Beezhold K, Castranova V. Tumor promoting or tumor suppressing of NF-kappa B, a matter of cell context dependency. Int Rev Immunol 2008; 27: 183-204.

(95.) Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal, malonaldehyde and related aldehydes. Free Radic Biol Med 1991; 11: 81-128.

(96.) Zarkovic N, Zarkovic K, Schaur RJ, Stolc S, Schlag G, Redl H, et al. 4-Hydroxynonenal as a second messenger of free radicals and growth modifying factor. Life Sci 1999; 65: 1901-4.

(97.) Uchida K. 4-Hydroxy-2-nonenal: a product and mediator of oxidative stress. Prog Lipid Res 2003; 42: 318-43.

(98.) Zarkovic N. 4-hydroxynonenal as a bioactive marker of pathophysiological processes. Mol Aspects Med 2003; 24: 281-91.

(99.) Zarkovic N, Tillian MH, Schaur J, Waeg G, Jurin M, Esterbauer H. Inhibition of melanoma B16-F10 growth by lipid peroxidation product 4-hydroxynonenal. Cancer Biother 1995; 10: 153-6.

(100.) Zarkovic N, Ilic Z, Jurin M, Schaur RJ, Puhl H, Esterbauer H. Stimulation of HeLa cell growth by physiological concentrations of 4-hydroxynonenal. Cell Biochem Funct 1993; 11: 279-86.

(101.) Borovic Sunjic S, Cipak A, Rabuzin F, Wildburger R, Zarkovic N. The influence of 4-hydroxy-2-nonenal on proliferation, differentiation and apoptosis of human osteosarcoma cells. Biofactors 2005; 24: 141-8.

(102.) Awasthi YC, Yang Y, Tiwari NK, Patrick B, Sharma A, Li J, et al. Regulation of 4-hydroxynonenal-mediated signaling by glutathione S-transferases. Free Radic Biol Med 2004; 37: 607-19.

(103.) Dwivedi S, Sharma A, Patrick B, Sharma R, Awasthi YC. Role of 4-hydroxynonenal and its metabolites in signaling. Redox Rep 2007; 12: 4-10.

(104.) Sovic A, Borovic S, Loncaric I, Kreuzer T, Zarkovic K, Vukovic T, et al. The carcinostatic and proapoptotic potential of 4-hydroxynonenal in HeLa cells is associated with its conjugation to cellular proteins. Anticancer Res 2001; 21: 1997-2004.

(105.) Numazawa S, Ishikawa M, Yoshida A, Tanaka S, Yoshida T. Atypical protein kinase C mediates activation of NF-E2-related factor 2 in response to oxidative stress. Am J Physiol Cell Physiol 2003; 285: C334-42.

Reprint requests: Dr Morana Jaganjac, Research Associate, Laboratory for Oxidative Stress, Division of Molecular Medicine Rudjer Boskovic Institute, Bijenicka 54, HR-10002 Zagreb, Croatia e-mail:

Morana Jaganjac

Division of Molecular Medicine, Rudjer Boskovic Institute, Zagreb, Croatia
COPYRIGHT 2010 Indian Council of Medical Research
No portion of this article can be reproduced without the express written permission from the copyright holder.
Copyright 2010 Gale, Cengage Learning. All rights reserved.

Article Details
Printer friendly Cite/link Email Feedback
Author:Jaganjac, Morana
Publication:Indian Journal of Medical Research
Article Type:Report
Geographic Code:4EXCR
Date:May 1, 2010
Previous Article:New vistas in treating diabetes--insight into a holistic approach.
Next Article:Evidence-based national vaccine policy.

Terms of use | Privacy policy | Copyright © 2021 Farlex, Inc. | Feedback | For webmasters