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Cyclin-dependent protein kinase 2 activity is required for mitochondrial translocation of Bax and disruption of mitochondrial transmembrane potential during etoposide-induced apoptosis

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Abstract

Previous studies have suggested that upregulation of Cyclin A-dependent protein kinase 2 (Cdk2) activity is an essential event in apoptotic progression and the mitochondrial permeability transition in human cancer cells. Here, we show that upregulated Cyclin A/Cdk2 activity precedes the proteolytic cleavage of PARP and is correlated with the mitochondrial translocation of Bax and the loss of mitochondrial transmembrane potential (Δψm) during etoposide-induced apoptosis in human cervical adenocarcinoma (HeLa) cells. Etoposide-induced apoptotic cell death is efficiently prevented in cells that overexpress a dominant negative mutant of Cdk2 (Cdk2-dn) or p21WAF1/CIP1, a specific Cdk inhibitor. Conversely, apoptotic cell death is promoted in Cyclin A-expressing cells. Disruption of the mitochondrial transmembrane potential in etoposide-induced cells is prevented in cells that overexpress Cdk2-dn or p21WAF1/CIP1, while this transition is prominently promoted in Cyclin A-expressing cells. We screened for mitochondrial Cdk2 targets in the etoposide-induced cells and found that the mitochondrial level of Bax is elevated by more than three fold in etoposide-treated cells and this elevation is effectively prevented in cells expressing Cdk2-dn under the same conditions. Thus, we suggest that Cdk2 activity is involved in the mitochondrial translocation of Bax, which plays an important role in the mitochondrial membrane permeability transition during apoptotic progression.

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References

  1. Bruce A, Edgar TL, Terry LO (2001) Endoreplication cell cycles: more for less. Cell 105:297–306

    Article  Google Scholar 

  2. Grana X, Reddy EP (1995) Cell cycle control in mammalian cells: role of Cyclins, Cyclin dependent kinases (CDKs), growth suppressor genes, and Cyclin-dependent kinase inhibitors (CKIs). Oncogene 11:211–219

    PubMed  CAS  Google Scholar 

  3. Van den Heuvel S, Harlow F (1993) Distinct roles for Cyclin-dependent kinases in cell cycle control. Science 262:2050–2054

    Article  PubMed  CAS  Google Scholar 

  4. Hakem A, Sasaki T, Kozieradzki I, Penninger JM (1999) The Cyclin-dependent kinase Cdk2 regulates thymocyte apoptosis. J Exp Med 189:957–968

    Article  PubMed  CAS  Google Scholar 

  5. Jin Z, El-Deiry WS (2005) Overview of cell death signaling pathways. Cancer Biol Ther 4:139–163

    Article  PubMed  CAS  Google Scholar 

  6. Fulda S, Debatin KM (2006) Extrinsic versus intrinsic apoptosis pathways in anticancer chemotherapy. Oncogene 25:4798–4811

    Article  PubMed  CAS  Google Scholar 

  7. Thorburn A (2004) Death receptor-induced cell killing. Cell Signal 16:139–144

    Article  PubMed  CAS  Google Scholar 

  8. Green DR, Reed JC (1998) Mitochondria and apoptosis. Science 281:1309–1312

    Article  PubMed  CAS  Google Scholar 

  9. Saelens X, Festjens N, Vande Walle L, van Gurp M, van Loo G, Vandenabeele P (2004) Toxic proteins released from mitochondria in cell death. Oncogene 23:2861–2874

    Article  PubMed  CAS  Google Scholar 

  10. Adachi S, Ito H, Tamamori-Adachi M et al (2001) Cyclin A/cdk2 activation is involved in hypoxia-induced apoptosis in cardiomyocytes. Circ Res 88:408–414

    PubMed  CAS  Google Scholar 

  11. Kim SG, Kim SN, Jong HS et al (2001) Caspase-mediated Cdk2 activation is a critical step to execute transforming growth factor-beta1-induced apoptosis in human gastric cancer cells. Oncogene 20:1254–1265

    Article  PubMed  CAS  Google Scholar 

  12. Jin YH, Yoo KJ, Lee YH, Lee SK (2000) Caspase 3-mediated cleavage of p21WAF1/CIP1-associated with the Cyclin A-Cyclin-dependent kinase 2 complex is a prerequisite for apoptosis in SK-HEP-1 cells. J Biol Chem 275:30256–30263

    Article  PubMed  CAS  Google Scholar 

  13. Shen SC, Huang TS, Jee SH, Kuo ML (1998) Taxol-induced p34cdc2 kinase activation and apoptosis inhibited by 12-O-tetradecanoylphorbol-13-acetate in human breast MCF-7 carcinoma cells. Cell Growth Differ 9:23–29

    PubMed  CAS  Google Scholar 

  14. Chadebech P, Truchet I, Brichese L, Valette A (2000) Up-regulation of cdc2 protein during paclitaxel-induced apoptosis. Int J Cancer 87:779–786

    Article  PubMed  CAS  Google Scholar 

  15. Shu CH, Yang WK, Shih YL, Kuo ML, Huang TS (1997) Cell cycle G2/M arrest and activation of Cyclin-dependent kinases associated with low-dose paclitaxel-induced sub-G1 apoptosis. Apoptosis 2:463–470

    Article  PubMed  CAS  Google Scholar 

  16. Shimizu T, O'Connor PM, Kohn KW, Pommier Y (1995) Unscheduled activation of Cyclin B1/Cdc2 kinase in human promyelocytic leukemia cell line HL60 cells undergoing apoptosis induced by DNA damage. Cancer Res 55:228–231

    PubMed  CAS  Google Scholar 

  17. Levkau B, Koyama H, Raines EW et al (1998) Cleavage of p21Cip1/Waf1 and p27Kip1 mediates apoptosis in endothelial cells through activation of Cdk2: role of a caspase cascade. Mol Cell 1:553–563

    Article  PubMed  CAS  Google Scholar 

  18. Hiromura K, Pippin JW, Blonski MJ, Roberts JM, Shankland SJ (2002) The subcellular localization of Cyclin dependent kinase 2 determines the fate of mesangial cells: role in apoptosis and proliferation. Oncogene 21:1750–1758

    Article  PubMed  CAS  Google Scholar 

  19. Finkielstein CV, Chen LG, Maller JL (2002) A role for G1/S Cyclin-dependent protein kinases in the apoptotic response to ionizing radiation. J Biol Chem 277:38476–38485

    Article  PubMed  CAS  Google Scholar 

  20. Park JA, Kim KW, Kim SI, Lee SK (1998) Caspase-3 specifically cleaves p21WAF1/CIP1 in the earlier stage of apoptosis in SK-HEP-1 human hepatoma cells. Eur J Biochem 257:242–248

    Article  PubMed  CAS  Google Scholar 

  21. Jin YH, Yim H, Park JH, Lee SK (2003) Cdk2 activity is associated with depolarization of mitochondrial membrane potential during apoptosis. Biochem Bioph Res Co 305:974–980

    Article  CAS  Google Scholar 

  22. Reed JC (1997) Cytochrome c: can't live with it-can't live without it. Cell 91:559–562

    Article  PubMed  CAS  Google Scholar 

  23. Kroemer G, Dallaporta B, Resche-Rigon M (1998) The mitochondrial death/life regulator in apoptosis and necrosis. Annu Rev Physiol 60:619–642

    Article  PubMed  CAS  Google Scholar 

  24. Susin SA, Lorenzo HK, Zamzami N et al (1999) Molecular characterization of mitochondrial apoptosis-inducing factor. Nature 397:441–446

    Article  PubMed  CAS  Google Scholar 

  25. Hirsch T, Susin SA, Marzo I, Marchetti P, Zamzami N, Kroemer G (1998) Mitochondrial permeability transition in apoptosis and necrosis. Cell Biol Toxicol 14:141–145

    Article  PubMed  CAS  Google Scholar 

  26. Ly JD, Grubb DR, Lawen A (2003) The mitochondrial membrane potential (Dm) in apoptosis; an update. Apoptosis 8:115–128

    Article  PubMed  CAS  Google Scholar 

  27. Zamzami N, Kroemer G (2001) The mitochondrion in apoptosis: how Pandora's box opens. Nat Rev Mol Cell Bio 2:67–71

    Article  CAS  Google Scholar 

  28. Nomura M, Shimizu S, Ito T, Narita M, Matsuda H, Tsujimoto Y (1999) Apoptotic cytosol facilitates Bax translocation to mitochondria that involves cytosolic factor regulated by Bcl-2. Cancer Res 59:5542–5548

    PubMed  CAS  Google Scholar 

  29. Gottlieb RA, Granville DJ (2002) Analyzing mitochondrial changes during apoptosis. Methods 26:341–347

    Article  PubMed  CAS  Google Scholar 

  30. Naour F, Hohenkirk L, Grolleau A et al (2001) Profiling changes in gene expression during differentiation and maturation of monocyte-derived dendritic cells using both oligonucleotide microarrays and proteomics. J Biol Chem 276:17920–17931

    Article  PubMed  Google Scholar 

  31. Lewis TS, Hunt JB, Aveline LD et al (2000) Identification of novel MAP kinase pathway signaling targets by functional proteomics and mass spectrometry. Mol Cell 6:1343–1354

    Article  PubMed  CAS  Google Scholar 

  32. Gharahdaghi F, Weinberg CR, Meagher DA, Imai BS, Mische SM (1999) Mass spectrometric identification of proteins from silver-stained polyacrylamide gel: a method for the removal of silver ions to enhance sensitivity. Electrophoresis 2:601–605

    Article  Google Scholar 

  33. Shevchenko A, Wilm M, Vorm O, Mann M (1996) Mass spectrometric sequencing of proteins from silver-stained polyacrylamide gels. Anal Chem 68:850–858

    Article  PubMed  CAS  Google Scholar 

  34. Troyan MB, Gilman VR, Gay CV (1997) Mitochondrial membrane potential changes in osteoblasts treated with parathyroid hormone and estradiol. Exp Cell Res 233:274–280

    Article  PubMed  CAS  Google Scholar 

  35. Meikrantz W, Schlegel R (1996) Suppression of apoptosis by dominant negative mutants of Cyclin-dependent protein kinases. J Biol Chem 271:10205–10209

    Article  PubMed  CAS  Google Scholar 

  36. Granes F, Roig MB, Brady HJ, Gil-Gomez G (2004) Cdk2 activation acts upstream of the mitochondrion during glucocorticoid induced thymocyte apoptosis. Eur J Immunol 34:2781–2790

    Article  PubMed  CAS  Google Scholar 

  37. Zhou BB, Li H, Yuan J, Kirschner MW (1998) Caspase-dependent activation of Cyclin-dependent kinases during Fas-induced apoptosis in Jurkat cells. Proc Natl Acad Sci USA 95:6785–6790

    Article  PubMed  CAS  Google Scholar 

  38. Zinkel S, Gross A, Yang E (2006) BCL2 family in DNA damage and cell cycle control. Cell Death Differ 13:1351–1359

    Article  PubMed  CAS  Google Scholar 

  39. Upreti M, Lyle CS, Skaug B, Du L, Chambers TC (2006) Vinblastine-induced apoptosis is mediated by discrete alterations in subcellular location, oligomeric structure, and activation status of specific Bcl-2 family members. J Biol Chem 281:15941–15950

    Article  PubMed  CAS  Google Scholar 

  40. Hu H, Jiang C, Schuster T, Li GX, Daniel PT, Lu J (2006) Inorganic selenium sensitizes prostate cancer cells to TRAIL-induced apoptosis through superoxide/p53/Bax-mediated activation of mitochondrial pathway. Mol Cancer Ther 5:1873–1882

    Article  PubMed  CAS  Google Scholar 

  41. Tikhomirov O, Carpenter G (2005) Bax activation and translocation to mitochondria mediate EGF-induced programmed cell death. J Cell Sci 118:5681–5690

    Article  PubMed  CAS  Google Scholar 

  42. Van Laethem A, Van Kelst S, Lippens S et al (2004) Activation of p38 MAPK is required for Bax translocation to mitochondria, cytochrome c release and apoptosis induced by UVB irradiation in human keratinocytes. FASEB J 18:1946–1978

    PubMed  CAS  Google Scholar 

  43. Hetz C, Vitte PA, Bombrun A et al (2005) Bax channel inhibitors prevent mitochondrion-mediated apoptosis and protect neurons in a model of global brain ischemia. J Biol Chem 280:42960–42970

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

This work was supported by the National Research Laboratory Fund (M10104000129-02J000005910) of the Ministry of Science and Technology, Republic of Korea and by the NSFC (grant # 30640064). We thank Dr. Kwang-Yeol Lee (Chonnam National University, Republic Korea) for providing human full-length p21WAF1/CIP1 cDNA and Cdk2-dn cDNA.

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Authors and Affiliations

  1. Division of Pharmaceutical Biosciences, College of Pharmacy and The Research Institute for Pharmaceutical Sciences, Seoul National University, Seoul, 151-742, Korea

    Joon-Seok Choi & Seung-Ki Lee

  2. Division of Pharmaceutical Biosciences, College of Pharmacy, Seoul National University, Seoul, 151-742, Korea

    Soona Shin, Ying Hua Jin, Hyungshin Yim, Kyo-Tan Koo, Kwang-Hoon Chun, You-Take Oh & Won Hee Lee

  3. College of Life Science, Jilin University, Changchun, 130012, China

    Ying Hua Jin

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  1. Joon-Seok Choi
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Correspondence to Seung-Ki Lee.

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Choi, JS., Shin, S., Jin, Y.H. et al. Cyclin-dependent protein kinase 2 activity is required for mitochondrial translocation of Bax and disruption of mitochondrial transmembrane potential during etoposide-induced apoptosis. Apoptosis 12, 1229–1241 (2007). https://doi.org/10.1007/s10495-006-0047-3

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