Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Inhibition of Cell Cycle Progression and Induction of Apoptosis in HeLa Cells by HY558-1, a Novel CDK Inhibitor Isolated from Penicillium minioluteum F558

  • Lim, Hae-Young (Department of Medical Genetics and Institute of Biomedical Science, Hanyang University) ;
  • Kim, Min-Kyoung (Department of Medical Genetics and Institute of Biomedical Science, Hanyang University) ;
  • Cho, Youl-Hee (Department of Medical Genetics and Institute of Biomedical Science, Hanyang University) ;
  • Kim, Jung-Mogg (Department of Microbiology, College of Medicine, Hanyang University) ;
  • Lim, Yoong-Ho (Bio/Molecular Informatics Center, Konkuk University) ;
  • Lee, Chul-Hoon (Department of Medical Genetics and Institute of Biomedical Science, Hanyang University)
  • Published : 2004.10.01
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Abstract

In the course of screening for a novel inhibitor of CDC2, HY558-1 was isolated from a culture broth of Penicillium minioluteum F558. Moreover, it was found that HY558-1 had an effect on both the cell cycle regulation and apoptosis of human cervical adenocarcinoma HeLa cells. A flow cytometric analysis of HeLa cells revealed appreciable cell cycle arrest at the G1 and G2/M phases following treatment with HY558-1. Furthermore, DNA fragmentation due to apoptosis was observed in HeLa cells treated with HY558-1. To obtain further information on the cell cycle arrest and apoptotic induction induced by HY558-1, the expression of certain cell cycle and apoptosis-associated proteins was examined using a Western blot analysis. The results revealed that HY558-1 inhibited the phosphorylation of pRb and decreased the expression levels of CDK2, CDC2, and cyclin A in the cell cycle progression. It was also shown that the level of $p21^{WAF1/CIP1}$ was increased in HeLa cells treated with 0.52 mM of HY558-1. Accordingly, HY558-1 was found to inhibit the proliferation of HeLa cells through the induction of G1 phase arrest by inhibiting pRb phosphorylation via an upregulation of $p21^{WAF1/CIP1}$, and G2/M phase arrest by directly inhibiting CDC2 and cyclin A. Moreover, HeLa cells treated with 0.52 mM of HY558-1 exhibited apoptotic induction associated with the cleavage of Bid and release of cytochrome c from mitochondria into the cytosol. Subsequent investigation of the activation of caspase-3 and cleavage of poly (ADP-ribose) polymerase (PARP) suggested that the mitochondrial pathway was primarily involved in the HY558-1-induced apoptosis in HeLa cells.

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References

  1. Ashkenazi, A. and V. M. Dixit. 1998. Death receptors: signaling and modulation. Science 281: 1305- 1308.
  2. Dantzer, E., V. Schreiber, C. Niedergang, C. Trucco, E. Flatter, G. De La Rubia, J. Oliver, V. Rolli, J. Menissier-de Murcia, and G. de Murcia. 1999. Involvement of poly(ADPribose) polymerase in base excision repair. Biochimie 81: 69-75.
  3. El-Deiry, W. S., T. Tokino, V. E. Velculescu, D. B. Levy, R. Parsons, J. M. Trent,D. Lin, W. E. Mercer, K. W. Kinzler, and B. Voglstein. 1993. WAF1, a potential mediator of p53 tumor suppression. Cell 75: 817-825.
  4. Green, D. R. 1998. Apoptotic pathway: The roads to ruin. Cell 94: 695- 698.
  5. Green, D. R. and J. C. Reed. 1998. Mitochondria and apoptosis. Science 281: 1309- 1312.
  6. Gross, A., X. M. Yin, K. Wang, M. C. Wei, J. Jockel, C. Milliman, H. Erdjument-Bromage, P. Tempst, and S. J. Korsmeyer. 1999. Caspase cleaved Bid target mitochondria and is required for cytochrome c release, while Bcl-xl prevents this release but not tumor necrosis factor-Rl/Fas death. J. Biol. Chem. 274: 1156- 1163.
  7. Guadagno, T. M. and J. W. Newport. 1996. Cdk2 kinase is required for entry into mitosis as a positive regulator of cdc2-cyclin B kinaseactivity. Cell 84: 73- 82.
  8. Harper, J. W. and S. J. Elledge. 1996. Cdk inhibitor in development and cancer. Curro Opin. Genet. Dev. 6: 56- 64.
  9. Hunter, T. and J. Pines. 1994. Cyclins and cancer. CyclinD andCDKinhibitors come of age. Cell 79: 573- 582.
  10. Jeong, Y, K. Kim, J. Oh, J. Park, J. Bang, S. Choi, and J. Lee. 2003. Growth inhibition and apoptosis induction of gastric cancer by copper (II) glycinate complex. J. Microbiol. Biotechnol. 13: 394- 399.
  11. Kim, M., Y Cho, J. Kim, M. Chun, S. Lee, Y Lim, and C.-H. Lee. 2003. Inhibition of cell-cycle progression in human promyeloc:ytic leukemia HL-60 cells by MCS-C2, novel cyclin-dependent kinase inhibitor.J. Microbiol. Biotechnol. 13: 607- 612.
  12. King, R. W., P. K. Jackson, and M. W. Kirschner. 1994. Mitosis in transition. Cell 79: 563- 571.
  13. Knudsen, E. S. and J. Y Wang. 1997. Dual mechanisms for the inhibition of E2F binding to RB by cyclin-dependent kinase-mediated RB phosphorylation. Mol. Cell. BioI. 17: 5771-5783.
  14. Lee, C.-H., H. Lim, M. K. Kim, Y-H. Cho, D.-K. Oh, C.-J. Kim, and Y. Lim. 2002. Isolation and biological properties of novel cell cycle inhibitor, HY558, isolated from Penicillium minioluteum F558. J. Microbiol. Biotechnol. 12: 470- 475.
  15. Lee, C.-H., M. Lee, Y. Cho, H. Lim, J. Jung, K. H. Kim, and Y. Lim. 200I. In vitro detection of apoptosis in human promyelocytic leukemia HL-60 cells by H-NMR. J. Microbiol. Biotechnol. 11: 539- 542.
  16. Lee, H., M. Jang, J. Choi, Y Lee, H. Park, Y. Lee, S. Kim, and M. Ahn. 2003. Proliferation, apoptosis, and telomerase activity in human cord blood CD34(+) cells cultured with combinations of various cytokines. J. Microbial. Biotechnol. 13(3): 422- 428.
  17. Li, H., H. Zhu, C. J. Xu, and J. Yuan. 1998. Cleavage of Bid by caspase 8 mediates the motochondrial damage in the Fas pathway of apoptosis, Cell 94: 491- 501.
  18. Lundberg, A. S. and R. A. Weinberg. 1998. Functional inactivation of the retinoblastoma protein requires sequential modification by at least two distinct cyclin-cdk complexes. Mol. Cell. BioI. 18: 753- 761.
  19. Luo, X., I. Budihardjo, H. Zou, C. Slaughter, and X. Wang. 1998. Bid, a Bcl2 interacting protein, mediates cytochrome c release from mitochondria in response to activation of cell death receptors. Cell 94: 481- 490.
  20. Matsushime, H., M. F. Roussel, R. A. Ashmun, and C. J. Sherr. 1991. Colony-stimulating factor 1 regulates novel cyclins during the G1 phase of the cell cycle. Cell 65: 701-713.
  21. Morgan, D. O. 1995. Principles of CDK regulation. Nature 374: 131- 134. https://doi.org/10.1038/374131a0
  22. Park, H., D. Jun, and Y. Kim. 2002. Apoptotic activity of insect pathogenic fungus Paecilomyces japonica toward human acute leukemia jurkat T cells is associated with mitochondria-dependent caspase-3 activation regulated by Bci-2. J. Microbiol. Biotechnol. 12: 950- 956.
  23. Polyak, K., M. H. Lee, H. Erdjument-Bromage, A. Koff, J. M. Roberts, P. Tempst, and J. Massague. 1994. Cloning of p27Kipl, a cyclin-dependent kinase inhibitor and a potential mediator of extracellular antimitogenic signals. Cell 78: 5966.
  24. Rhee, W. J. and T. H. Park. 2001. Flow cytometric analysis of silkworm hemolymph on the baculovirus-induced insect cell apoptosis. J. Microbiol. Biotechnol. 11: 853- 857.
  25. Sherr, C. J. 1993. Mammalian G1 cyclins. Cell 73: 1059-1065.
  26. Sherr, C. J. and J. M. Roberts. 1999. CDK inhibitors: Positive and negative regulators of Gl phase progression. Genes Dev. 13: 1501-1512.
  27. Song, J., S. Kim, Y Lee, K. Lee, H. Jung, M. Kim, K. Kim, R. Brown, and Y. Kim. 2002. Adenovirus-mediated antisense expression of telomerase template RNA induces apoptosis in lung cancer cells. J. Microbiol. Biotechnol. 12(1): 89- 95.
  28. Stennicke, H. R., J. M. Jurgensmeier, H. Shin, Hwain Shin, Q. Deveraux, B. B. Wolf, X. Yang, Q. Zhou, H. M. Ellerby, L. M. Ellerby, D. Bredesen, D. R. Green, J. C. Reed, C. J. Froelich, and G. S. Salvesen. 1998. Pro-caspase-3 is a major physiologic target of caspase-8. J. BioI. Chem. 273: 27084-27090.
  29. Stennicke, H. R. and G. S. Salvesen. 1998. Properties of the caspases. Biochim. Biophys. Acta 1387: 17-31.
  30. Tang, D., J. M. Lahti, and V. J. Kidd. 2000. Caspase-8 activation and Bid cleavage contribute to MCF7 cellular execution in a caspase-3-dependent manner during Staurosporine-mediated apoptosis. J. BioI. Chem. 275: 9303- 9307.
  31. Viswanath, V., Y Wu, R. Boonplueang, S. Chen, F. F. Stevenson, F. Yantiri, L. Yang, F. Beal, and J. K. Anderson. 2001. Caspase-9 activation results in downstream caspase-8 activation and Bid cleavage in I-methyl-4-phenyl-l,2,3,6tetrahydropyridine- induced Parkinsons disease. J. Neurosci. 21: 9519- 9528.
  32. Weinberg, R. A. 1995. The retinoblastoma protein and cell cycle control. Cell 81: 323- 330. https://doi.org/10.1016/0092-8674(95)90385-2
  33. Zha, J., S. Weiler, K. J. Oh, M. C. Wei, and S. J. Korsmeyer. 2000. Posttranslational N-myristoylation of Bid as a molecular switch for targeting mitochondria and apoptosis. Science 290: 1761-1765.