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The time course of Akt and ERK activation on XIAP expression in HEK 293 cell line

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Abstract

The cell growth is controlled by the interaction of survival and cell growth arrest pathways as well as apoptosis mechanisms which determine the outcome of cell faith as proliferation or apoptosis. In this study, we have studied the activity of survival pathways, i.e., Akt and ERK1/2 with regard to XIAP (inhibitor of apoptosis) in serum starved and stimulated conditions. The HEK-293 cells were cultured in RPMI + 10% FBS. The cells were serum starved by switching to medium with 1% FBS for 24 h and serum stimulated by changing the medium to 10% FBS following serum starvation. The expression of p-Akt, p-ERK, Akt, ERK and XIAP was studied in various time points using western blot. The apoptosis was evaluated by DNA condensation using Hoechst 33258 and Caspase-3 assay. In serum starved condition expression of p-Akt and XIAP is very low. Serum stimulation increases p-Akt and p-ERK within 5 min and sustains a high level for 30 min. The expression of total Akt and ERK1/2 has not changed significantly for 24 h. XIAP expression starts at 6 h after serum stimulation, reaches to maximum level at 12 h and decreases to baseline within 24 h. Furthermore, serum starvation for 24 h does not induced apoptosis and DNA condensation. Taken together, the results indicate that serum activates Akt and ERK pathways earlier than XIAP expression. Furthermore, XIAP expression is low in serum starvation unlike p-ERK which suggests a survival role for ERK in serums starvation. The expression pattern of XIAP indicates induction by Akt and/or ERK activation which requires further studies.

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References

  1. Song G, Ouyang G, Bao S (2005) The activation of Akt/PKB signaling pathway and cell survival. J Cell Mol Med 9:59–71. doi:10.1111/j.1582-4934.2005.tb00337.x

    Article  CAS  PubMed  Google Scholar 

  2. Downward J (1998) Mechanisms and consequences of activation of protein kinase B/Akt. Curr Opin Cell Biol 10:262–267. doi:10.1016/S0955-0674(98)80149-x

    Article  CAS  PubMed  Google Scholar 

  3. Kerkhoff E, Rapp UR (1998) Cell cycle targets of Ras/Raf signalling. Oncogene 17:1457–1462. doi:10.1038/sj.onc.1202185

    Article  CAS  PubMed  Google Scholar 

  4. Shapiro P (2001) Ras-MAP kinase signaling pathways and control of cell proliferation: relevance to cancer therapy. Crit Rev Clin Lab Sci 39:285–330. doi:10.1080/10408360290795538

    Article  Google Scholar 

  5. Shaul YD, Seger R (2007) The MEK/ERK cascade: from signaling specificity to diverse functions. Biochim Biophys Acta 1773:1213–1226. doi:10.1016/j.bbamcr.2006.10.005

    Article  CAS  PubMed  Google Scholar 

  6. Wennstrom S, Downward J (1999) Role of phosphoinositide 3-kinase in activation of ras and mitogen-activated protein kinase by epidermal growth factor. Mol Cell Biol 19:4279–4288

    CAS  PubMed  Google Scholar 

  7. McCubrey JA, Lee JT, Steelman LS et al (2001) Interactions between the PI3K and Raf signaling pathways can result in the transformation of hematopoietic cells. Cancer Detect Prev 25:375–393

    CAS  PubMed  Google Scholar 

  8. Sheng H, Shao J, DuBois RN (2001) Akt/PKB activity is required for Ha-Ras-mediated transformation of intestinal epithelial cells. J Biol Chem 276:14498–14504

    CAS  PubMed  Google Scholar 

  9. Guan KL, Figueroa C, Brtva TR et al (2000) Negative regulation of the serine/threonine kinase B-Raf by Akt. J Biol Chem 275:27354–27359

    CAS  PubMed  Google Scholar 

  10. Reusch HP, Zimmermann S, Schaefer M et al (2001) Regulation of Raf by Akt controls growth and differentiation in vascular smooth muscle cells. J Biol Chem 276:33630–33637. doi:10.1074/jbc.M105322200

    Article  CAS  PubMed  Google Scholar 

  11. Zimmermann S, Moelling K (1999) Phosphorylation and regulation of Raf by Akt (protein kinase B). Science 286:1741–1744. doi:10.1126/science.286.5445.1741

    Article  CAS  PubMed  Google Scholar 

  12. Deveraux QL, Reed JC (1999) IAP family proteins–suppressors of apoptosis. Genes Dev 13:239–252. doi:10.1101/gad.13.3.239

    Article  CAS  PubMed  Google Scholar 

  13. Eiben LJ, Duckett CS (1998) The IAP family of apoptotic regulators. Results Probl Cell Differ 24:91–104

    CAS  PubMed  Google Scholar 

  14. Richter BW, Duckett CS (2000) The IAP proteins: caspase inhibitors and beyond. Sci STKE 2000:PE1. doi:10.1126/stke.2000.44.pe1

  15. Yang YL, Li XM (2000) The IAP family: endogenous caspase inhibitors with multiple biological activities. Cell Res 10:169–177. doi:10.1038/sj.cr.7290046

    Article  CAS  PubMed  Google Scholar 

  16. Dan HC, Sun M, Kaneko S et al (2004) Akt phosphorylation and stabilization of X-linked inhibitor of apoptosis protein (XIAP). J Biol Chem 279:5405–5412. doi:10.1074/jbc.M312044200

    Article  CAS  PubMed  Google Scholar 

  17. Tian S, Mewani RR, Kumar D et al (2006) Interaction and stabilization of X-linked inhibitor of apoptosis by Raf-1 protein kinase. Int J Oncol 29:861–867

    CAS  PubMed  Google Scholar 

  18. Carter BZ, Milella M, Tsao T et al (2003) Regulation and targeting of antiapoptotic XIAP in acute myeloid leukemia. Leukemia 17:2081–2089. doi:10.1038/sj.leu.2403113

    Article  CAS  PubMed  Google Scholar 

  19. Downward J (2004) PI 3-kinase, Akt and cell survival. Semin Cell Dev Biol 15:177–182. doi:10.1016/j.semcdb.2004.01.002

    Article  CAS  PubMed  Google Scholar 

  20. Gagnon V, St-Germain ME, Parent S et al (2003) Akt activity in endometrial cancer cells: regulation of cell survival through cIAP-1. Int J Oncol 23:803–810

    CAS  PubMed  Google Scholar 

  21. Kim D, Kim S, Koh H et al (2001) Akt/PKB promotes cancer cell invasion via increased motility and metalloproteinase production. FASEB J 15:1953–1962. doi:10.1096/fj.01-0198com

    Article  CAS  PubMed  Google Scholar 

  22. Liu W, Bagaitkar J, Watabe K (2007) Roles of AKT signal in breast cancer. Front Biosci 12:4011–4019. doi:10.2741/2367

    Article  CAS  PubMed  Google Scholar 

  23. Toker A, Yoeli-Lerner M (2006) Akt signaling and cancer: surviving but not moving on. Cancer Res 66:3963–3966. doi:10.1158/0008-5472.CAN-06-0743

    Article  CAS  PubMed  Google Scholar 

  24. Asselin E, Mills GB, Tsang BK (2001) XIAP regulates Akt activity and caspase-3-dependent cleavage during cisplatin-induced apoptosis in human ovarian epithelial cancer cells. Cancer Res 61:1862–1868

    CAS  PubMed  Google Scholar 

  25. Hu P, Han Z, Couvillon AD et al (2004) Critical role of endogenous Akt/IAPs and MEK1/ERK pathways in counteracting endoplasmic reticulum stress-induced cell death. J Biol Chem 279:49420–49429. doi:10.1074/jbc.M407700200

    Article  CAS  PubMed  Google Scholar 

  26. Keramati AR, Goliaei A, Abkhezr M, et al. (2008) The role of Akt/PI3Kinase and ERK1/2 activation on expression of XIAP and proliferation of HT-29 cells. Proceeding of 10th Iranian Pharmaceutical Science Conference, Aug 21–24, 2006, p 308

  27. Rommel C, Clarke BA, Zimmermann S et al (1999) Differentiation stage-specific inhibition of the Raf-MEK-ERK pathway by Akt. Science 286:1738–1741. doi:10.1126/science.286.5445.1738

    Article  CAS  PubMed  Google Scholar 

  28. Mitsui H, Takuwa N, Maruyama T et al (2001) The MEK1-ERK map kinase pathway and the PI 3-kinase-Akt pathway independently mediate anti-apoptotic signals in HepG2 liver cancer cells. Int J Cancer 92:55–62. doi:10.1002/1097-0215(200102)9999:9999<::AID-IJC1143>3.0.CO;2-O

    Article  CAS  PubMed  Google Scholar 

  29. Amantana A, London CA, Iversen PL et al (2004) X-linked inhibitor of apoptosis protein inhibition induces apoptosis and enhances chemotherapy sensitivity in human prostate cancer cells. Mol Cancer Ther 3:699–707

    CAS  PubMed  Google Scholar 

  30. Liston P, Roy N, Tamai K et al (1996) Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes. Nature 379:349–353. doi:10.1038/379349a0

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

We thank Mr. Ali Reza Kazemi and Mr. Hassan Akbari for technical assistance. This work was supported by a Deputy of Research, Tehran University of Medical Sciences grant no 132/10450 to M.H.G.

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Authors and Affiliations

  1. Department of Pharmacology-Toxicology, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran University, New Building, Room J-309, 14155-6451, Tehran, Iran

    Mousa Abkhezr, Ali Reza Keramati, Seyed Nasser Ostad & Mohammad H. Ghahremani

  2. Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Iran

    Ali Reza Keramati

  3. Department of Biochemistry, Institute of Biochemistry and Biophysics, Tehran University, Tehran, Iran

    Jamshid Davoodi

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  1. Mousa Abkhezr
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  2. Ali Reza Keramati
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  3. Seyed Nasser Ostad
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  4. Jamshid Davoodi
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  5. Mohammad H. Ghahremani
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Correspondence to Mohammad H. Ghahremani.

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Abkhezr, M., Keramati, A.R., Ostad, S.N. et al. The time course of Akt and ERK activation on XIAP expression in HEK 293 cell line. Mol Biol Rep 37, 2037–2042 (2010). https://doi.org/10.1007/s11033-009-9658-4

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  • DOI: https://doi.org/10.1007/s11033-009-9658-4

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