title: Expression of xCT and activity of system xc(-) are regulated by NRF2 in human breast cancer cells in response to oxidative stress.
abstract: Cancer cells adapt to high levels of oxidative stress in order to survive and proliferate by activating key transcription factors. One such master regulator, the redox sensitive transcription factor NF E2 Related Factor 2 (NRF2), controls the expression of cellular defense genes including those encoding intracellular redox-balancing proteins involved in glutathione (GSH) synthesis. Under basal conditions, Kelch-like ECH-associated protein 1 (KEAP1) targets NRF2 for ubiquitination. In response to oxidative stress, NRF2 dissociates from KEAP1, entering the nucleus and binding to the antioxidant response element (ARE) in the promoter of its target genes. Elevated reactive oxygen species (ROS) production may deplete GSH levels within cancer cells. System xc(-), an antiporter that exports glutamate while importing cystine to be converted into cysteine for GSH synthesis, is upregulated in cancer cells in response to oxidative stress. Here, we provided evidence that the expression of xCT, the light chain subunit of system xc(-), is regulated by NRF2 in representative human breast cancer cells. Hydrogen peroxide (H2O2) treatment increased nuclear translocation of NRF2, also increasing levels of xCT mRNA and protein and extracellular glutamate release. Overexpression of NRF2 up-regulated the activity of the xCT promoter, which contains a proximal ARE. In contrast, overexpression of KEAP1 repressed promoter activity and decreased xCT protein levels, while siRNA knockdown of KEAP1 up-regulated xCT protein levels and transporter activity. These results demonstrate the importance of the KEAP1/NRF2 pathway in balancing oxidative stress in breast cancer cells through system xc(-). We have previously shown that xCT is upregulated in various cancer cell lines under oxidative stress. In the current investigation, we focused on MCF-7 cells as a model for mechanistic studies.
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada L8S 4L8.
authors.forename: Eric
authors.initials: E
authors.lastname: Habib
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada L8S 4L8.
authors.forename: Katja
authors.initials: K
authors.lastname: Linher-Melville
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada L8S 4L8.
authors.forename: Han-Xin
authors.initials: HX
authors.lastname: Lin
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada L8S 4L8. Electronic address: singhg@mcmaster.ca.
authors.forename: Gurmit
authors.initials: G
authors.lastname: Singh
mesh_terms: D027182:Amino Acid Transport System y+ / Q000235:genetics / Q000378:metabolism*; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D002467:Cell Nucleus / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D018698:Glutamic Acid / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D000072019:Kelch-Like ECH-Associated Protein 1; D061986:MCF-7 Cells; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects; D011401:Promoter Regions, Genetic; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D027182:Amino Acid Transport System y+; D047908:Intracellular Signaling Peptides and Proteins; C431364:KEAP1 protein, human; D000072019:Kelch-Like ECH-Associated Protein 1; D051267:NF-E2-Related Factor 2; D021122:Protein Subunits; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; C118895:SLC7A11 protein, human; D018698:Glutamic Acid; D006861:Hydrogen Peroxide; D005978:Glutathione
keywords: Hydrogen peroxide; KEAP1; NRF2; Oxidative stress; System x(c)(−); xCT
references.citation: Mol Cancer. 2013;12(1):138
references.pmid: 24238102
references.citation: Proc Natl Acad Sci U S A. 1996 Nov 26;93(24):13943-8
references.pmid: 8943040
references.citation: Toxicol Sci. 2010 Aug;116(2):590-603
references.pmid: 20421342
references.citation: Biochim Biophys Acta. 2013 Jun;1832(6):729-41
references.pmid: 23429074
references.citation: Environ Toxicol. 2014 Nov;29(11):1292-300
references.pmid: 23649983
references.citation: FEBS Lett. 1974 May 15;42(1):68-72
references.pmid: 4859511
references.citation: Cancer Res. 1991 Feb 1;51(3):794-8
references.pmid: 1846317
references.citation: J Neurochem. 2011 Nov;119(3):569-78
references.pmid: 21883218
references.citation: Neurochem Res. 2013 Jul;38(7):1375-93
references.pmid: 23585122
references.citation: Neuropharmacology. 2004 Feb;46(2):273-84
references.pmid: 14680765
references.citation: Biochem Pharmacol. 2000 Jun 1;59(11):1433-9
references.pmid: 10751553
references.citation: J Gastroenterol. 2014 May;49(5):853-63
references.pmid: 23771433
references.citation: Mol Cell Biol. 2004 Dec;24(24):10941-53
references.pmid: 15572695
references.citation: Leuk Lymphoma. 1995 Mar;17(1-2):101-9
references.pmid: 7773147
references.citation: Chem Biol Interact. 2014 Feb 25;209:56-67
references.pmid: 24361488
references.citation: Proc Natl Acad Sci U S A. 1996 Dec 10;93(25):14960-5
references.pmid: 8962164
references.citation: Glia. 2011 Oct;59(10):1387-401
references.pmid: 21590811
references.citation: Cancer Res. 2013 Apr 1;73(7):2199-210
references.pmid: 23382044
references.citation: Nat Rev Drug Discov. 2013 Dec;12(12):931-47
references.pmid: 24287781
references.citation: FASEB J. 2005 Dec;19(14):2085-7
references.pmid: 16234432
references.citation: J Biol Chem. 2009 Jan 9;284(2):1106-15
references.pmid: 19017641
references.citation: Antioxid Redox Signal. 2000 Winter;2(4):665-71
references.pmid: 11213471
references.citation: J Pathol. 2010 Mar;220(4):446-51
references.pmid: 19967722
references.citation: J Exp Med. 1987 Mar 1;165(3):628-40
references.pmid: 2880923
references.citation: Cell Death Dis. 2015;6:e1601
references.pmid: 25590811
references.citation: J Virol. 2015 Feb;89(4):2268-86
references.pmid: 25505069
references.citation: Mol Cancer Ther. 2009 Aug;8(8):2432-40
references.pmid: 19638449
references.citation: J Leukoc Biol. 2007 Apr;81(4):974-82
references.pmid: 17200146
references.citation: Mol Cell Biol. 2006 Jan;26(1):221-9
references.pmid: 16354693
references.citation: J Neurosci. 2003 Apr 15;23(8):3394-406
references.pmid: 12716947
references.citation: Biochem Biophys Res Commun. 2010 Jan 1;391(1):91-5
references.pmid: 19896463
references.citation: PLoS Med. 2006 Oct;3(10):e420
references.pmid: 17020408
references.citation: Curr Med Chem. 2014;21(16):1861-70
references.pmid: 24533814
references.citation: Biochem Biophys Res Commun. 1996 Apr 25;221(3):570-6
references.pmid: 8630002
references.citation: Cancer Res. 1991 Aug 15;51(16):4287-94
references.pmid: 1868449
references.citation: J Biol Chem. 2002 Nov 22;277(47):44765-71
references.pmid: 12235164
references.citation: Cancer. 1993 Aug 1;72(3):783-7
references.pmid: 8392905
references.citation: Cancer Res. 1994 Aug 1;54(15):4077-83
references.pmid: 8033140
references.citation: Biochem Biophys Res Commun. 2007 Nov 3;362(4):816-21
references.pmid: 17822677
references.citation: J Mol Cell Cardiol. 2010 Oct;49(4):576-86
references.pmid: 20685357
references.citation: Cancer Res. 2014 Apr 15;74(8):2295-305
references.pmid: 24686172
references.citation: Pain. 2014 Jan;155(1):28-36
references.pmid: 23999057
references.citation: Mol Cell Biol. 2014 Oct;34(20):3800-16
references.pmid: 25092871
references.citation: J Biol Chem. 1999 Apr 23;274(17):11455-8
references.pmid: 10206947
references.citation: Cancer Cell. 2012 Jul 10;22(1):66-79
references.pmid: 22789539
references.citation: J Virol. 2004 Nov;78(21):11926-38
references.pmid: 15479833
references.citation: Free Radic Biol Med. 2007 Nov 1;43(9):1335-47
references.pmid: 17893047
references.citation: Drug Metab Rev. 2006;38(4):769-89
references.pmid: 17145701
references.citation: J Neurosci. 2006 Oct 11;26(41):10514-23
references.pmid: 17035536
references.citation: Mol Cell Biol. 2014 Sep 15;34(18):3421-34
references.pmid: 25002527
references.citation: Mol Pharmacol. 2001 Apr;59(4):837-43
references.pmid: 11259628
references.citation: Amino Acids. 2012 Jan;42(1):171-9
references.pmid: 21369940
references.citation: Cell Biosci. 2014 Feb 10;4(1):8
references.pmid: 24507416
references.citation: BMC Cancer. 2014;14:415
references.pmid: 24913037
references.citation: Mol Cancer Ther. 2004 Jul;3(7):823-32
references.pmid: 15252143
references.citation: Histopathology. 2012 May;60(6):943-52
references.pmid: 22348534
references.citation: J Biol Chem. 1984 Feb 25;259(4):2435-40
references.pmid: 6142042
references.citation: J Neurochem. 2006 Aug;98(3):916-25
references.pmid: 16771835
references.citation: Cell Stress Chaperones. 2012 Sep;17(5):529-38
references.pmid: 22351438
references.citation: Biochim Biophys Acta. 2001 Jun 6;1512(2):335-44
references.pmid: 11406111
references.citation: J Biol Chem. 1984 Feb 25;259(4):2441-5
references.pmid: 6142043
references.citation: J Cell Sci. 2009 Dec 15;122(Pt 24):4452-64
references.pmid: 19920073
references.citation: Free Radic Biol Med. 2015 Jun;83:77-88
references.pmid: 25687825
references.citation: Cell Biochem Funct. 2004 Nov-Dec;22(6):343-52
references.pmid: 15386533
references.citation: Cancer Treat Rev. 2008 Oct;34(6):505-20
references.pmid: 18413281
references.citation: Int J Radiat Oncol Biol Phys. 1992;24(3):527-31
references.pmid: 1399740
references.citation: Proc Natl Acad Sci U S A. 2005 Jul 19;102(29):10070-5
references.pmid: 16006525
references.citation: Mol Cell. 2006 Mar 3;21(5):689-700
references.pmid: 16507366
references.citation: PLoS One. 2009;4(8):e6588
references.pmid: 19668370
references.citation: Cell Signal. 2014 Mar;26(3):512-20
references.pmid: 24308969