Cellular and Molecular Life Sciences

, Volume 67, Issue 20, pp 3523–3533 | Cite as

Involvement of aryl hydrocarbon receptor nuclear translocator in EGF-induced c-Jun/Sp1-mediated gene expression

Research Article


Aryl hydrocarbon receptor nuclear translocator (ARNT) binds to other basic helix-loop-helix Per/ARNT/Sim (bHLH-PAS) proteins to form functional transcriptional complexes in order to regulate specific biological pathways. Here, we report a novel mechanism that upon EGF treatment, ARNT associated with non-bHLH-PAS transcription factors, c-Jun/Sp1, and regulated gene expression, through forming a c-Jun/ARNT/Sp1 complex and binding to the Sp1 site of the gene promoter. EGF-induced promoter activity and the mRNA level of 12(S)-lipoxygenase as well as the association between c-Jun and Sp1 were reduced by ARNT knockdown. Notably, dominant negative c-Jun mutant, TAM-67, blocked ARNT-mediated 12(S)-lipoxygenase expression, demonstrating that c-Jun was responsible for the transcriptional activation. Moreover, ARNT knockdown also inhibited other EGF-induced c-Jun/Sp1 mediated gene expression, such as p21WAF1/CIP1. Our results reveal a novel mechanism by which ARNT acts as a modulator to bridge the c-Jun/Sp1 interaction and plays a role in EGF-mediated gene expression under normoxic conditions.


Epidermal growth factor (EGF) Aryl hydrocarbon receptor nuclear translocator (ARNT) Gene expression c-Jun/Sp1 Protein–DNA interaction 

Supplementary material

18_2010_392_MOESM1_ESM.ppt (126 kb)
Supplementary Figures (PPT 126 kb)


  1. 1.
    Kewley RJ, Whitelaw ML, Chapman-Smith A (2004) The mammalian basic helix-loop-helix/PAS family of transcriptional regulators. Int J Biochem Cell Biol 36:189–204CrossRefPubMedGoogle Scholar
  2. 2.
    Maltepe E, Schmidt JV, Baunoch D, Bradfield CA, Simon MC (1997) Abnormal angiogenesis and responses to glucose and oxygen deprivation in mice lacking the protein ARNT. Nature 386:403–407CrossRefPubMedGoogle Scholar
  3. 3.
    Carmeliet P, Dor Y, Herbert J-M, Fukumura D, Brusselmans K, Dewerchin M, Neeman M, Bono F, Abramovitch R, Maxwell P, Koch CJ, Ratcliffe P, Moons L, Jain RK, Collen D, Keshet E (1998) Role of HIF-1[alpha] in hypoxia-mediated apoptosis, cell proliferation and tumour angiogenesis. Nature 394:485–490CrossRefPubMedGoogle Scholar
  4. 4.
    Denison MS, Nagy SR (2003) Activation of the aryl hydrocarbon receptor by structurally diverse exogenous and endogenous chemicals. Annu Rev Pharmacol Toxicol 43:309–334CrossRefPubMedGoogle Scholar
  5. 5.
    Woods SL, Whitelaw ML (2002) Differential activities of murine single minded 1 (SIM1) and SIM2 on a hypoxic response element. Cross-talk between basic helix-loop-helix/per-Arnt-Sim homology transcription factors. J Biol Chem 277:10236–10243CrossRefPubMedGoogle Scholar
  6. 6.
    Kozak KR, Abbott B, Hankinson O (1997) ARNT-deficient mice and placental differentiation. Dev Biol 191:297–305CrossRefPubMedGoogle Scholar
  7. 7.
    Moffett P, Pelletier J (2000) Different transcriptional properties of mSim-1 and mSim-2. FEBS Lett 466:80–86CrossRefPubMedGoogle Scholar
  8. 8.
    Wright CW, Duckett CS (2009) The aryl hydrocarbon nuclear translocator alters CD30-mediated NF-kappaB-dependent transcription. Science 323:251–255CrossRefPubMedGoogle Scholar
  9. 9.
    Chang KY, Shen MR, Lee MY, Wang WL, Su WC, Chang WC, Chen BK (2009) Epidermal growth factor-activated aryl hydrocarbon receptor nuclear translocator/HIF-1{beta} signal pathway up-regulates cyclooxygenase-2 gene expression associated with squamous cell carcinoma. J Biol Chem 284:9908–9916CrossRefPubMedGoogle Scholar
  10. 10.
    van Cruijsen H, Giaccone G, Hoekman K (2005) Epidermal growth factor receptor and angiogenesis: opportunities for combined anticancer strategies. Int J Cancer 117:883–888CrossRefPubMedGoogle Scholar
  11. 11.
    Salomon DS, Brandt R, Ciardiello F, Normanno N (1995) Epidermal growth factor-related peptides and their receptors in human malignancies. Crit Rev Oncol Hematol 19:183–232CrossRefPubMedGoogle Scholar
  12. 12.
    Yarden Y, Sliwkowski MX (2001) Untangling the ErbB signaling network. Nat Rev Mol Cell Biol 2:127–137CrossRefPubMedGoogle Scholar
  13. 13.
    Chen BK, Chang WC (2000) Functional interaction between c-Jun and promoter factor Sp1 in epidermal growth factor-induced gene expression of human 12(S)-lipoxygenase. Proc Natl Acad Sci USA 97:10406–10411CrossRefPubMedGoogle Scholar
  14. 14.
    Chen BK, Kung HC, Tsai TY, Chang WC (2000) Essential role of mitogen-activated protein kinase pathway and c-Jun induction in epidermal growth factor-induced gene expression of human 12-lipoxygenase. Mol Pharmacol l57:153–161Google Scholar
  15. 15.
    Nie D, Krishnamoorthy S, Jin R, Tang K, Chen Y, Qiao Y, Zacharek A, Guo Y, Milanini J, Pages G, Honn KV (2006) Mechanisms regulating tumor angiogenesis by 12-lipoxygenase in prostate cancer cells. J Biol Chem 281:18601–18609CrossRefPubMedGoogle Scholar
  16. 16.
    Kardassis D, Papakosta P, Pardali K, Moustakas A (1999) c-Jun transactivates the promoter of the human p21(WAF1/Cip1) gene by acting as a superactivator of the ubiquitous transcription factor Sp1. J Biol Chem 274:29572–29581CrossRefPubMedGoogle Scholar
  17. 17.
    Numayama-Tsuruta K, Kobayashi A, Sogawa K, Fujii-Kuriyama Y (1997) A point mutation responsible for defective function of the aryl-hydrocarbon-receptor nuclear translocator in mutant Hepa-1c1c7 cells. Eur J Biochem 246:486–495CrossRefPubMedGoogle Scholar
  18. 18.
    Liu YW, Arakawa T, Yamamoto S, Chang WC (1997) Transcriptional activation of human 12-lipoxygenase gene promoter is mediated through Sp1 consensus sites in A431 cells. Biochem J 324:133–140PubMedGoogle Scholar
  19. 19.
    Tsou JH, Chang KY, Wang WC, Tseng JT, Su WC, Hung LY, Chang WC, Chen BK (2008) Nucleolin regulates c-Jun/Sp1-dependent transcriptional activation of cPLA2alpha in phorbol ester-treated non-small cell lung cancer A549 cells. Nucleic Acids Res 36:217–227CrossRefPubMedGoogle Scholar
  20. 20.
    Andrews NC, Faller DV (1991) A rapid micropreparation technique for extraction of DNA-binding proteins from limiting numbers of mammalian cells. Nucleic Acids Res 19:2499CrossRefPubMedGoogle Scholar
  21. 21.
    Saccani S, Pantano S, Natoli G (2001) Two waves of nuclear factor {kappa}B recruitment to target promoters. J Exp Med 193:1351–1360CrossRefPubMedGoogle Scholar
  22. 22.
    Zhu Y, Saunders MA, Yeh H, Deng W-g, Wu KK (2002) Dynamic regulation of cyclooxygenase-2 promoter activity by isoforms of CCAAT/enhancer-binding proteins. J Biol Chem 277:6923–6928CrossRefPubMedGoogle Scholar
  23. 23.
    Dong Z, Crawford HC, Lavrovsky V, Taub D, Watts R, Matrisian LM, Colburn NH (1997) A dominant negative mutant of jun blocking 12-O-tetradecanoylphorbol-13-acetate-induced invasion in mouse keratinocytes. Mol Carcinog 19:204–212CrossRefPubMedGoogle Scholar
  24. 24.
    Blaine SA, Wick M, Dessev C, Nemenoff RA (2001) Induction of cPLA2 in lung epithelial cells and non-small cell lung cancer is mediated by Sp1 and c-Jun. J Biol Chem 276:42737–42743CrossRefPubMedGoogle Scholar
  25. 25.
    Kardassis D, Papakosta P, Pardali K, Moustakas A (1999) c-Jun transactivates the promoter of the human p21WAF1/Cip1 gene by acting as a superactivator of the ubiquitous transcription factor Sp1. J Biol Chem 274:29572–29581CrossRefPubMedGoogle Scholar
  26. 26.
    Timar J, Raso E, Dome B, Li L, Grignon D, Nie D, Honn KV, Hagmann W (2000) Expression, subcellular localization and putative function of platelet-type 12-lipoxygenase in human prostate cancer cell lines of different metastatic potential. Int J Cancer 87:37–43CrossRefPubMedGoogle Scholar
  27. 27.
    Pidgeon GP, Lysaght J, Krishnamoorthy S, Reynolds JV, O’Byrne K, Nie D, Honn KV (2007) Lipoxygenase metabolism: roles in tumor progression and survival. Cancer Metastasis Rev 26:503–524CrossRefPubMedGoogle Scholar
  28. 28.
    Cordero-Erausquin M, Marubio LM, Klink R, Changeux JP (2000) Nicotinic receptor function: new perspectives from knockout mice. Trends Pharmacol Sci 21:211–217CrossRefPubMedGoogle Scholar
  29. 29.
    Leslie CC (1997) Properties and regulation of cytosolic phospholipase A2. J Biol Chem 272:16709–16712CrossRefPubMedGoogle Scholar
  30. 30.
    Leigh IM, Navsaria H, Purkis PE, McKay IA, Bowden PE, Riddle PN (1995) Keratins (K16 and K17) as markers of keratinocyte hyperproliferation in psoriasis in vivo and in vitro. Br J Dermatol 133:501–511CrossRefPubMedGoogle Scholar
  31. 31.
    Cheng M, Olivier P, Diehl JA, Fero M, Roussel MF, Roberts JM, Sherr CJ (1999) The p21(Cip1) and p27(Kip1) CDK ‘inhibitors’ are essential activators of cyclin D-dependent kinases in murine fibroblasts. EMBO J 18:1571–1583CrossRefPubMedGoogle Scholar
  32. 32.
    Giaccia A, Siim BG, Johnson RS (2003) HIF-1 as a target for drug development. Nat Rev Drug Discov 2:803–811CrossRefPubMedGoogle Scholar
  33. 33.
    Gunton JE, Kulkarni RN, Yim S, Okada T, Hawthorne WJ, Tseng Y-H, Roberson RS, Ricordi C, O’Connell PJ, Gonzalez FJ, Kahn CR (2005) Loss of ARNT/HIF1[beta] mediates altered gene expression and pancreatic-islet dysfunction in human type 2 diabetes. Cell 122:337–349CrossRefPubMedGoogle Scholar
  34. 34.
    Semenza GL (2001) Hypoxia-inducible factor 1: oxygen homeostasis and disease pathophysiology. Trends Mol Med 7:345–350CrossRefPubMedGoogle Scholar
  35. 35.
    Sutter CH, Yin H, Li Y, Mammen JS, Bodreddigari S, Stevens G, Cole JA, Sutter TR (2009) EGF receptor signaling blocks aryl hydrocarbon receptor-mediated transcription and cell differentiation in human epidermal keratinocytes. Proc Natl Acad Sci USA 106:4266–4271CrossRefPubMedGoogle Scholar
  36. 36.
    Chen BK, Huang CC, Chang WC, Chen YJ, Kikkawa U, Nakahama K-i, Morita I, Chang WC (2007) PP2B-mediated dephosphorylation of c-Jun C terminus regulates phorbol ester-induced c-Jun/Sp1 interaction in A431 cells. Mol Biol Cell 18:1118–1127CrossRefPubMedGoogle Scholar
  37. 37.
    Tojo M, Matsuzaki K, Minami T, Honda Y, Yasuda H, Chiba T, Saya H, Fujii-Kuriyama Y, Nakao M (2002) The aryl hydrocarbon receptor nuclear transporter is modulated by the SUMO-1 conjugation system. J Biol Chem 277:46576–46585CrossRefPubMedGoogle Scholar
  38. 38.
    Zhong H, De Marzo AM, Laughner E, Lim M, Hilton DA, Zagzag D, Buechler P, Isaacs WB, Semenza GL, Simons JW (1999) Overexpression of hypoxia-inducible factor 1alpha in common human cancers and their metastases. Cancer Res 59:5830–5835PubMedGoogle Scholar

Copyright information

© Springer Basel AG 2010

Authors and Affiliations

  • Wan-Chen Huang
    • 1
  • Shu-Ting Chen
    • 1
  • Wei-Chiao Chang
    • 2
    • 6
  • Kwang-Yu Chang
    • 3
  • Wen-Chang Chang
    • 1
    • 4
    • 5
  • Ben-Kuen Chen
    • 1
    • 4
    • 5
  1. 1.Department of Pharmacology, College of MedicineNational Cheng Kung UniversityTainanTaiwan, ROC
  2. 2.Graduate Institute of Medical GeneticsKaohsiung Medical UniversityKaohsiungTaiwan, ROC
  3. 3.National Institute of Cancer ResearchNational Health Research InstitutesTainanTaiwan, ROC
  4. 4.Center for Gene Regulation and Signal Transduction ResearchNational Cheng Kung UniversityTainanTaiwan, ROC
  5. 5.Institute of Biosignal Transduction, College of Bioscience and BiotechnologyNational Cheng Kung UniversityTainanTaiwan, ROC
  6. 6.Cancer CenterKaohsiung Medical University HospitalKaohsiungTaiwan, ROC

Personalised recommendations