Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Manuscript
  • Published:

Constitutively active Akt1 protects HL60 leukemia cells from TRAIL-induced apoptosis through a mechanism involving NF-κB activation and cFLIPL up-regulation

Leukemia volume 17, pages 379–389 (2003)Cite this article

Abstract

TRAIL is a member of the tumor necrosis factor superfamily which induces apoptosis in cancer but not in normal cells. Akt1 promotes cell survival and blocks apoptosis. The scope of this paper was to investigate whether a HL60 human leukemia cell clone (named AR) with constitutively active Akt1 was resistant to TRAIL. We found that parental (PT) HL60 cells were very sensitive to a 6 h incubation in the presence of TRAIL and died by apoptosis. In contrast, AR cells were resistant to TRAIL concentrations as high as 2 μg/ml for 24 h. Two pharmacological inhibitors of PI3K, Ly294002 and wortmannin, restored TRAIL sensitivity of AR cells. AR cells stably overexpressing PTEN had lower Akt1 activity and were sensitive to TRAIL. Conversely, PT cells stably overexpressing a constitutive active form of Akt1 became TRAIL resistant. TRAIL activated caspase-8 but not caspase-9 or -10 in HL60 cells. We did not observe a protective effect of Bcl-XL or Bcl-2 against the cytotoxic activity of TRAIL, even though TRAIL induced cleavage of BID. There was a close correlation between TRAIL sensitivity and intranuclear presence of the p50 subunit of NF-κB. Higher levels of the FLICE inhibitory protein, cFLIPL, were observed in TRAIL-resistant cells. Both the cell permeable NF-κB inhibitor SN50 and cycloheximide lowered cFLIPLexpression and restored sentivity of AR cells to TRAIL. Our results suggest that Akt1 may be an important regulator of TRAIL sensitivity in HL60 cells through the activation of NF-κB and up-regulation of cFLIPL synthesis

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8
Figure 9

Similar content being viewed by others

References

  1. Wiley SR, Schooley K, Smolak PJ, Din WS, Huang CP, Nicholl JK, Sutherland GR, Rauch C, Smith CA . Identification and characterization of a new member of the TNF family that induces apoptosis. Immunity 1995; 3: 673–682.

    Article  CAS  PubMed  Google Scholar 

  2. Pitti RM, Marsters SA, Ruppert S, Donahue CJ, Moore A, Ashkenazi A . Induction of apoptosis by Apo-2 ligand, a new member of the tumor necrosis factor cytokine family. J Biol Chem 1996; 271: 12687–12690.

    Article  CAS  PubMed  Google Scholar 

  3. Walczak H, Miller RE, Ariail K, Gliniak B, Griffith TS, Kubin M, Chin W, Jones J, Woodward A, Le T, Smith C, Smolak P, Goodwin RG, Rauch CT, Schuh JC, Lynch DH . Tumoricidal activity of tumor necrosis factor-related apoptosis-inducing ligand in vivo. Nat Med 1999; 5: 157–163.

    Article  CAS  PubMed  Google Scholar 

  4. Ashkenazi A, Pai RC, Fong S, Lueng S, Lawrence DA, Marsters SA, Blackie C, Chang L, McMurtrey AE, Hebert A, DeForge L, Koumenis IL, Lewis D, Harris L, Bussiere J, Koeppen H, Shahrokh Z, Schwall RH . Safety and antitumor activity of recombinant soluble Apo2 ligand. J Clin Invest 1999; 104: 155–162.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Griffith TS, Lynch DH . TRAIL: a molecule with multiple receptors and control mechanisms. Curr Opin Immunol 1998; 254: 439–459.

    Google Scholar 

  6. De Maria R, Zeuner A, Eramo A, Domenichelli C, Bonci D, Grignani F, Srinivasula SM, Alnemri ES, Testa U, Peschle C . Negative regulation of erythropoiesis by caspase-mediated cleavage of GATA-1. Nature 1999; 401: 489–493.

    Article  CAS  PubMed  Google Scholar 

  7. Song K, Chen Y, Goke R, Wilmen A, Seidel C, Goke A, Hilliard B, Chen Y . Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is an inhibitor of autoimmune inflammation and cell cycle progression. J Exp Med 2000; 191: 1095–1103.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Jo M, Kim T-H, Seol D-W, Esplen JE, Dorko K, Billiar TR, Strom SC . Apoptosis induced in normal human hepatocytes by tumor necrosis factor-related apoptosis-inducing ligand. Nat Med 2000; 6: 564–567.

    Article  CAS  PubMed  Google Scholar 

  9. Schulze-Osheroff K, Ferrari D, Los M, Wesselborg S, Peter ME . Apoptosis signaling by death receptors. Apoptosis signaling by death receptors. Eur J Biochem 1998; 254: 439–459.

    Article  Google Scholar 

  10. Zhang XD, Nguyen T, Thomas WD, Sanders JE, Hersey P . Mechanisms of resistance of normal cells to TRAIL induced apoptosis vary between different cell types. FEBS Lett 2000; 482: 193–199.

    Article  CAS  PubMed  Google Scholar 

  11. Mitsiades N, Mitsiades CS, Poulaki V, Anderson KC, Treon SP . Intracellular regulation of tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in human multiple myeloma cells. Blood 2002; 99: 2162–2171.

    Article  CAS  PubMed  Google Scholar 

  12. Chen X, Thakkar H, Tyan F, Gim S, Robinson H, Lee C, Pandey SK, Nwekorie C, Onwudiwe N, Srivastava RK . Constitutively active Akt is an important regulator of TRAIL sensitivity in prostate cancer. Oncogene 2001; 20: 6073–6083.

    Article  CAS  PubMed  Google Scholar 

  13. Sheridan JP, Marsters SA, Pitti RM, Gurney A, Skubatch M, Baldwin D, Ramakrishnan L, Gray CL, Baker K, Wood WI, Goddard AD, Godowsli P, Ashkenazi A . Control of TRAIL-induced apoptosis by a family of signalling and decoy receptors. Science 1997; 277: 818–821.

    Article  CAS  PubMed  Google Scholar 

  14. Marsters SA, Sheridan JP, Pitti RM, Huang A, Skubatch M, Baldwin D, Yuan J, Gurney A, Goddard AD, Godowski P, Ashkenazi A . A novel receptor for Apo2L/TRAIL contains a truncated death domain. Curr Biol 1997; 7: 1003–1006.

    Article  CAS  PubMed  Google Scholar 

  15. Chan TO, Rittenhouse SE, Tsichlis PN . AKT/PKB and other D3 phosphoinositide-regulated kinases: kinase activation by phosphoinositide-dependent phosphorylation. Annu Rev Biochem 1999; 68: 965–1014.

    Article  CAS  PubMed  Google Scholar 

  16. Brazil DP, Hemmings BA . Ten years of protein kinase B signalling: a hard Akt to follow. Trends Biochem Sci 2001; 26: 657–664.

    Article  CAS  PubMed  Google Scholar 

  17. Nicholson KM, Anderson NG . The protein kinase B/Akt signalling pathway in human malignancy. Cell Signal 2002; 14: 381–395.

    Article  CAS  PubMed  Google Scholar 

  18. Cantrell DA . Phosphoinositide 3-kinase signalling pathways. J Cell Sci 2001; 114: 1439–1445.

    CAS  PubMed  Google Scholar 

  19. Lawlor MA, Alessi DR . PKB/Akt: a key mediator of cell proliferation, survival and insulin responses?. J Cell Sci 2001; 114: 2903–2910.

    CAS  PubMed  Google Scholar 

  20. Thakkar H, Chen X, Tyan F, Gim S, Robinson H, Lee C, Pandy SK, Nwokorie C, Onwudiwe N, Svrivastava RK . Pro-survival function of Akt/protein kinase B in prostate cancer cells: relationship with TRAIL resistance. J Biol Chem 2001; 276: 38361–38369.

    Article  CAS  PubMed  Google Scholar 

  21. Beresford SA, Davies MA, Gallick GE, Donato NJ . Differential effects of phosphatidylinositol-3/Akt-kinase inhibition on apoptotic sensitization to cytokines in LNCaP and PCc-3 prostate cancer cells. J Interfer Cytokine Res 2001; 21: 313–322.

    Article  CAS  Google Scholar 

  22. Nicoletti I, Migliorati G, Pagliacci MC, Grignani F, Riccardi C . A rapid and simple method for measuring thymocyte apoptosis by propidium iodide staining and flow cytometry. J Immunol Methods 1991; 139: 271–279.

    Article  CAS  PubMed  Google Scholar 

  23. Curti A, Ratta M, Corinti S, Girolomoni G, Ricci F, Tazzari P, Siena M, Grande A, Fogli M, Tura S, Lemoli RM . Interleukin-11 induces Th2 polarization of human CD4(+) T cells. Blood 2001; 97: 2758–2763.

    Article  CAS  PubMed  Google Scholar 

  24. Borgatti P, Martelli AM, Bellacosa A, Casto R, Massari L, Capitani S, Neri LM . Translocation of Akt/PKB to the nucleus of osteoblast-like MC3T3-E1 cells exposed to proliferative growth factors. FEBS Lett 2000; 477: 27–32.

    Article  CAS  PubMed  Google Scholar 

  25. Martelli AM, Bortul R, Bareggi R, Grill V, Narducci P, Zweyer M . Biochemical and morphological changes in the nuclear matrix prepared from apoptotic HL-60 cells. Effect of different stabilizing procedures. J Cell Biochem 1999; 74: 99–110.

    Article  CAS  PubMed  Google Scholar 

  26. Martelli AM, Bortul R, Bareggi R, Tabellini G, Grill V, Baldini G, Narducci P . The pro-apoptotic drug camptothecin stimulates phospholipase D activity and diacylglycerol production in the nucleus of HL-60 human promyelocytic leukemia cells. Cancer Res 1999; 59: 3961–3967.

    CAS  PubMed  Google Scholar 

  27. Mitsiades CS, Treon SP, Mitsiades N, Shima Y, Richardson P, Schlossman R, Hideshima T, Anderson KC . TRAIL/Apo2L ligand selectively induces apoptosis and overcomes drug resistance in multiple myeloma : therapeutic applications. Blood 2001; 98: 795–804.

    Article  CAS  PubMed  Google Scholar 

  28. Lamothe B, Aggarwal BB . Ectopic expression of Bcl-2 and Bcl-XL inhibits apoptosis induced by TNF-related apoptosis-inducing ligand (TRAIL) through suppression of caspases-8, 7, and 3 and BID cleavage in human acute myelogenous leukaemia cell line HL-60. J Interfer Cytokine Res 2002; 22: 269–279.

    Article  CAS  Google Scholar 

  29. Cuvillier O, Levade T . Sphingosine 1-phosphate antagonizes apoptosis of human leukaemia cells by inhibiting release of cytochrome c and Smac/DIABLO from mitochondria. Blood 2001; 98: 2828–2836.

    Article  CAS  PubMed  Google Scholar 

  30. Leslie NR, Downes CP . PTEN: the down side of PI 3-kinase signalling. Cell Signal 2002; 14: 285–295.

    Article  CAS  PubMed  Google Scholar 

  31. Wen J, Ramadevi N, Nguyen D, Perkins CM, Worthington E, Bhalla K . Antileukemic drugs increase death receptor 5 levels and enhance Apo-2L-induced apoptosis of human acute leukaemia cells. Blood 2000; 96: 3900–3906.

    CAS  PubMed  Google Scholar 

  32. Panka Dj, Mano T, Suhara T, Walsh K, Mier JW . Phosphatidylinositol 3-kinase/Akt activity up-regulates cFLIP expression in tumor cells. J Biol Chem 2001; 276: 6893–6896.

    Article  CAS  PubMed  Google Scholar 

  33. Kreuz S, Siegmund D, Scheurich P, Wajant H . NF-κB upregulate cFLIP, a cycloheximide-sensitive inhibitor of death receptor signaling. Mol Cell Biol 2001; 21: 3964–3973.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Bernard D, Quatannens B, Vandenbunder B, Abbadie C . Rel/NF-κB transcription factors protect against tumor necrosis facor (TNF)-related apoptosis-inducing ligand (TRAIL)-induced apoptosis by up-regulating the TRAIL decoy receptor DcR1. J Biol Chem 2001; 276: 27322–27328.

    Article  CAS  PubMed  Google Scholar 

  35. Kischkel FC, Lawrence DA, Chuntharapai A, Schow P, Kim Kj, Ashkenazi A . Apo2L/TRAIL-dependent recruitment of endogenous FADD and caspase-8 to death receptors 4 and 5. Immunity 2000; 12: 611–620.

    Article  CAS  PubMed  Google Scholar 

  36. Sprick MR, Weigand MA, Rieser E, Rauch CT, Juo P, Blenis J, Krammer PH, Walczak H . FADD/MORT 1 and caspase-8 are recruited to TRAIL receptors 1 and 2 and are essential for apoptosis mediated by TRAIL receptor 2. Immunity 2000; 12: 599–609.

    Article  CAS  PubMed  Google Scholar 

  37. Mitsiades N, Poulaki V, Tseleni-Balafouta S, Koutras DA, Stamenkovic I . Thyroid carcinoma cells are resistant to FAS-mediated apoptosis but sensitive to tumor necrosis factor-related apoptosis-inducing ligand. Cancer Res 2000; 60: 4122–4129.

    CAS  PubMed  Google Scholar 

  38. Mitsiades N, Poulaki V, Mitsiades C, Tsokos M . Ewing’s sarcoma family tumors are sensitive to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and express DR4 and DR5 receptors. Cancer Res 2001; 61: 2704–2712.

    CAS  PubMed  Google Scholar 

  39. Secchiero P, Gonelli A, Celeghini C, Mirandola P, Guidotti L, Visani G, Capitani S, Zauli G . Activation of nitric oxide synthase pathway represents a key component of tumor necrosis factor-related apoptosis inducing ligand-mediated cytoxicity on hematological malignancies. Blood 2001; 98: 2220–2228.

    Article  CAS  PubMed  Google Scholar 

  40. Kaufmann SH, Earnshaw WC . Induction of apoptosis by cancer chemotherapy. Exp Cell Res 2000; 256: 42–49.

    Article  CAS  PubMed  Google Scholar 

  41. Hengartner MO . The biochemistry of apoptosis. Nature 2000; 407: 770–776.

    Article  CAS  PubMed  Google Scholar 

  42. Lin YZ, Yao SY, Veach RA, Torgerson TR, Hawiger J . Inhibition of nuclear translocation of transcription factor NF-κB by a synthetic peptide containing a cell membrane-permeable motif and nuclear localization sequence. J Biol Chem 1995; 270: 14255–14258.

    Article  CAS  PubMed  Google Scholar 

  43. Olsson A, Diaz T, Aguilar-Santelises M, Osterborg A, Celsing F, Jondal M, Osorio LM . Sensitization to TRAIL-induced apoptosis and modulation of FLICE-inhibitory protein in B chronic lymphocytic leukemia by actinomycin D. Leukemia 2001; 15: 1868–1877.

    Article  CAS  PubMed  Google Scholar 

  44. Bin L, Li X, Xu L-G, Shu H-B . The short splice form of Casper/cFLIP is a major cellular inhibitor of TRAIL-induced apoptosis. FEBS Lett 2002; 510: 37–40.

    Article  CAS  PubMed  Google Scholar 

  45. Yuan XJ, Whang YE . PTEN sensitizes prostate cancer cells to death receptor-mediated and drug-induced apoptosis through a FADD-dependent pathway. Oncogene 2002; 21: 319–327.

    Article  CAS  PubMed  Google Scholar 

  46. Jeremias I, Kupatt C, Baumann B, Herr I, Wirth T, Debatin KM . Inhibition of nuclear factor κB attenuates apoptosis resistance in lymphoid cells. Blood 1998; 91: 4624–4631.

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from: AIRC, Italian MIUR Cofin 2001, Selected Topics Research Fund from Bologna University, CARISBO Foundation.

Author information

Authors and Affiliations

  1. Dipartimento di Morfologia Umana Normale, Università di Trieste, Trieste, Italy

    R Bortul & R Bareggi

  2. Servizio di Immunoematologia e Trasfusionale, Policlinico S Orsola-Malpighi, Bologna, Italy

    P L Tazzari

  3. Dipartimento di Scienze Anatomiche Umane e Fisiopatologia dell’Apparato Locomotore, Sezione di Anatomia, Cell Signalling Laboratory, Università di Bologna, Bologna, Italy

    A Cappellini, G Tabellini, A M Billi, L Manzoli, L Cocco & A M Martelli

  4. Istituto di Citomorfologia Normale e Patologica del CNR, c/o IOR, Bologna, Italy

    A M Martelli

Authors
  1. R Bortul
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. P L Tazzari
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A Cappellini
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. G Tabellini
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. A M Billi
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. R Bareggi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. L Manzoli
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. L Cocco
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. A M Martelli
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Bortul, R., Tazzari, P., Cappellini, A. et al. Constitutively active Akt1 protects HL60 leukemia cells from TRAIL-induced apoptosis through a mechanism involving NF-κB activation and cFLIPL up-regulation. Leukemia 17, 379–389 (2003). https://doi.org/10.1038/sj.leu.2402793

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.leu.2402793

Keywords

This article is cited by

Search

Advanced search

Quick links