Skip to main content
SpringerLink
Log in

5-Phenylselenyl- and 5-methylselenyl-methyl-2′-deoxyuridine induce oxidative stress, DNA damage, and caspase-2-dependent apoptosis in cancer cells

  • Original Paper
  • Published:
Apoptosis Aims and scope Submit manuscript

Abstract

In the present study, we investigated the signaling pathways implicated in the induction of apoptosis by two modified nucleosides, 5-phenylselenyl-methyl-2′-deoxyuridine (PhSe-T) and 5-methylselenyl-methyl-2′-deoxyuridine (MeSe-T), using human cancer cell lines. The induction of apoptosis was associated with proteolytic activation of caspase-3 and -9, PARP cleavage, and decreased levels of IAP family members, including c-IAP-1 and c-IAP-2, but had no effect on XIAP and survivin. PhSe-T and MeSe-T also enhanced the activities of caspase-2 and -8, Bid cleavage, and the conformational activation of Bax. Additionally, nucleoside derivative-induced apoptosis was inhibited by the selective inhibitors of caspase-2, -3, -8, and -9 and also by si-RNAs against caspase-2, -3, -8, and -9; however, inhibition of caspase-2 and -3 was more effective at preventing apoptosis than inhibition of caspase-8 and -9. Moreover, the inhibition of caspase-2 activation by the pharmacological inhibitor z-VDVAD-fmk or by the knockdown of protein expression using siRNA suppressed nucleoside derivative-induced caspase-3 activation, but not vice versa. PhSe-T and MeSe-T also induced a Δψm loss via a CsA-insensitive mechanism, ROS production, and DNA damage, including strand breaks. Moreover, ROS scavengers such as NAC, tiron, and quercetin inhibited nucleoside derivative-induced ROS generation and apoptosis by blocking the sequential activation of caspase-2 and -3, indicating the role of ROS in caspase-2-mediated apoptosis. Taken together, these results indicate that caspase-2 acts upstream of caspase-3 and that caspase-2 functions in response to DNA damage in both PhSe-T- and MeSe-T-induced apoptosis. Our results also suggest that ROS are critical regulators of the sequential activation of caspase-2 and -3 in nucleoside derivative-treated cancer cells.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

Abbreviations

ANOVA:

Analysis of variance

CsA:

Cyclosporin A

DAPI:

4′-6-Diamidino-2-phenylindole

DiOC6:

3,3′-Dihexyloxacarbocyanine

DSBs:

Double-strand breaks

FACS:

Fluorescence-activated cell sorter

IAP:

Inhibitors of apoptosis proteins

LDH:

Lactate dehydrogenase

MeSe-T:

5-Methylselenyl-methyl-2′-deoxyuridine

MMP:

Mitochondrial membrane potential

NAC:

N-acetylcysteine

PhSe-T:

5-Phenylselenyl-methyl-2′-deoxyuridine

PI:

Propidium iodide

pNA:

p-Nitroanilide

ROS:

Reactive oxygen species

SCGE:

Single-cell gel electrophoresis

SSBs:

Single-strand breaks

tiron:

4,5-Dihydroxy-1,3-benzenedisulfonic acid

z-DEVD-fmk:

N-benzyloxycarbonyl-Asp-Glu-Val-Asp-fmk

z-IETD-fmk:

N-benzyloxycarbonyl-Ile-Glu-Thr-Asp-fmk

z-LEHD-fmk:

N-benzyloxycarbonyl-Leu-Glu-His-Asp-fmk

z-VDVAD-fmk:

N-benzyloxycarbonyl-Val-Asp-Val-Ala-Asp-fmk

References

  1. Zhu C, Johansson M, Karlsson A (2000) Incorporation of nucleoside analogs into nuclear or mitochondrial DNA is determined by the intracellular phosphorylation site. J Biol Chem 275:26727–26731

    PubMed  CAS  Google Scholar 

  2. Zhivotovsky B, Orrenius S (2005) Caspase-2 function in response to DNA damage. Biochem Biophys Res Commun 331:859–867

    Article  PubMed  CAS  Google Scholar 

  3. Troy CM, Shelanski ML (2003) Caspase-2 redux. Cell Death Differ 10:101–107

    Article  PubMed  CAS  Google Scholar 

  4. Shi M, Vivian CJ, Lee KJ, Ge C, Morotomi-Yano K, Manzl C et al (2009) DNA-PKcs-PIDDosome: a nuclear caspase-2-activating complex with role in G2/M checkpoint maintenance. Cell 136:508–520

    Article  PubMed  CAS  Google Scholar 

  5. Lin CF, Chen CL, Chang WT, Jan MS, Hsu LJ, Wu RH et al (2004) Sequential caspase-2 and caspase-8 activation upstream of mitochondria during ceramide and etoposide-induced apoptosis. J Biol Chem 279:40755–40761

    Article  PubMed  CAS  Google Scholar 

  6. Kim BM, Hong SH (2011) Sequential caspase-2 and caspase-8 activation is essential for saikosaponin a-induced apoptosis of human colon carcinoma cell lines. Apoptosis 16:184–197

    Article  PubMed  Google Scholar 

  7. Burney S, Niles JC, Dedon PC, Tannenbaum SR (1999) DNA damage in deoxynucleosides and oligonucleotides treated with peroxynitrite. Chem Res Toxicol 12:513–520

    Article  PubMed  CAS  Google Scholar 

  8. Prasad V, Chandele A, Jagtap JC, Sudheer Kumar P, Shastry P (2006) ROS-triggered caspase-2 activation and feedback amplification loop in beta-carotene-induced apoptosis. Free Radic Biol Med 41:431–442

    Article  PubMed  CAS  Google Scholar 

  9. Madesh M, Zong WX, Hawkins BJ, Ramasamy S, Venkatachalam T, Mukhopadhyay P et al (2009) Execution of superoxide-induced cell death by the proapoptotic Bcl-2-related proteins Bid and Bak. Mol Cell Biol 29:3099–3112

    Article  PubMed  CAS  Google Scholar 

  10. Hong IS, Greenberg MM (2004) Mild generation of 5-(2′-deoxyuridinyl)methyl radical from a phenyl selenide precursor. Org Lett 6:5011–5013

    Article  PubMed  CAS  Google Scholar 

  11. Hong IS, Greenberg MM (2005) Efficient DNA interstrand cross-link formation from a nucleotide radical. J Am Chem Soc 127:3692–3693

    Article  PubMed  CAS  Google Scholar 

  12. Hong IS, Greenberg MM (2005) DNA interstrand cross-link formation initiated by reaction between singlet oxygen and a modified nucleotide. J Am Chem Soc 127:10510–10511

    Article  PubMed  CAS  Google Scholar 

  13. Hong IS, Ding H, Greenberg MM (2006) Oxygen independent DNA interstrand cross-link formation by a nucleotide radical. J Am Chem Soc 128:485–491

    Article  PubMed  CAS  Google Scholar 

  14. Peng X, Hong IS, Li H, Seidman MM, Greenberg MM (2008) Interstrand cross-link formation in duplex and triplex DNA by modified pyrimidines. J Am Chem Soc 130:10299–10306

    Article  PubMed  CAS  Google Scholar 

  15. Rode AB, Kim BM, Park SH, Hong IS, Hong SH (2011) Potent radiosensitizing agents: 5-methylselenyl- and 5-phenylselenyl-methyl-2′-deoxyuridine. Bioorg Med Chem Lett 21:1151–1154

    Article  PubMed  CAS  Google Scholar 

  16. Yamaguchi H, Wang HG (2001) The protein kinase PKB/Akt regulates cell survival and apoptosis by inhibiting Bax conformational change. Oncogene 20:7779–7786

    Article  PubMed  CAS  Google Scholar 

  17. Gao Z, Tian Y, Wang J, Yin Q, Wu H, Li YM et al (2007) A dimeric Smac/diablo peptide directly relieves caspase-3 inhibition by XIAP. Dynamic and cooperative regulation of XIAP by Smac/Diablo. J Biol Chem 282:30718–30727

    Article  PubMed  CAS  Google Scholar 

  18. Mehmet H (2000) Caspases find a new place to hide. Nature 403:29–30

    Article  PubMed  CAS  Google Scholar 

  19. Li H, Zhu H, Xu CJ, Yuan J (1998) Cleavage of BID by caspase 8 mediates the mitochondrial damage in the Fas pathway of apoptosis. Cell 94:491–501

    Article  PubMed  CAS  Google Scholar 

  20. Rogakou EP, Pilch DR, Orr AH, Ivanova VS, Bonner WM (1998) DNA double-stranded breaks induce histone H2AX phosphorylation on serine 139. J Biol Chem 273:5858–5868

    Article  PubMed  CAS  Google Scholar 

  21. Burma S, Chen BP, Murphy M, Kurimasa A, Chen DJ (2001) ATM phosphorylates histone H2AX in response to DNA double-strand breaks. J Biol Chem 276:42462–42467

    Article  PubMed  CAS  Google Scholar 

  22. Sedelnikova OA, Pilch DR, Redon C, Bonner WM (2003) Histone H2AX in DNA damage and repair. Cancer Biol Ther 2:233–235

    PubMed  CAS  Google Scholar 

  23. Rogakou EP, Boon C, Redon C, Bonner WM (1999) Megabase chromatin domains involved in DNA double-strand breaks in vivo. J Cell Biol 146:905–916

    Article  PubMed  CAS  Google Scholar 

  24. Zamzami N, Marchetti P, Castedo M, Zanin C, Vayssière JL, Petit PX et al (1995) Reduction in mitochondrial potential constitutes an early irreversible step of programmed lymphocyte death in vivo. J Exp Med 181:1661–1672

    Article  PubMed  CAS  Google Scholar 

  25. Zhang D, Berry MD, Paterson IA, Boulton AA (1999) Loss of mitochondrial membrane potential is dependent on the apoptotic program activated: prevention by R-2HMP. J Neurosci Res 58:284–292

    Article  PubMed  CAS  Google Scholar 

  26. Düssmann H, Rehm M, Kögel D, Prehn JH (2003) Outer mitochondrial membrane permeabilization during apoptosis triggers caspase-independent mitochondrial and caspase-dependent plasma membrane potential depolarization: a single-cell analysis. J Cell Sci 116:525–536

    Article  PubMed  Google Scholar 

  27. Guo Y, Srinivasula SM, Druilhe A, Fernandes-Alnemri T, Alnemri ES (2002) Caspase-2 induces apoptosis by releasing proapoptotic proteins from mitochondria. J Biol Chem 277:13430–13437

    Article  PubMed  CAS  Google Scholar 

  28. Paroni G, Henderson C, Schneider C, Brancolini C (2002) Caspase-2 can trigger cytochrome C release and apoptosis from the nucleus. J Biol Chem 277:15147–15161

    Article  PubMed  CAS  Google Scholar 

  29. Lassus P, Opitz-Araya X, Lazebnik Y (2002) Requirement for caspase-2 in stress-induced apoptosis before mitochondrial permeabilization. Science 297:1352–1354

    Article  PubMed  CAS  Google Scholar 

  30. Demple B, Harrison L (1994) Repair of oxidative damage to DNA: enzymology and biology. Annu Rev Biochem 63:915–948

    Article  PubMed  CAS  Google Scholar 

  31. Zha S, Sekiguchi J, Brush JW, Bassing CH, Alt FW (2008) Complementary functions of ATM and H2AX in development and suppression of genomic instability. Proc Natl Acad Sci USA 105:9302–9306

    Article  PubMed  CAS  Google Scholar 

  32. Dickey JS, Redon CE, Nakamura AJ, Baird BJ, Sedelnikova OA, Bonner WM (2009) H2AX: functional roles and potential applications. Chromosoma 118:683–692

    Article  PubMed  CAS  Google Scholar 

  33. Kruidering M, Evan GI (2000) Caspase-8 in apoptosis: the beginning of “the end”? IUBMB Life 50:85–90

    PubMed  CAS  Google Scholar 

  34. Luo X, Budihardjo I, Zou H, Slaughter C, Wang X (1998) Bid, a Bcl2 interacting protein, mediates cytochrome c release from mitochondria in response to activation of cell surface death receptors. Cell 94:481–490

    Article  PubMed  CAS  Google Scholar 

  35. Desagher S, Osen-Sand A, Nichols A, Eskes R, Montessuit S, Lauper S et al (1999) Bid-induced conformational change of Bax is responsible for mitochondrial cytochrome c release during apoptosis. J Cell Biol 144:891–901

    Article  PubMed  CAS  Google Scholar 

  36. Eskes R, Desagher S, Antonsson B, Martinou JC (2000) Bid induces the oligomerization and insertion of Bax into the outer mitochondrial membrane. Mol Cell Biol 20:929–935

    Article  PubMed  CAS  Google Scholar 

  37. Li P, Nijhawan D, Budihardjo I, Srinivasula SM, Ahmad M, Alnemri ES et al (1997) Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell 91:479–489

    Article  PubMed  CAS  Google Scholar 

  38. Jiang X, Wang X (2000) Cytochrome c promotes caspase-9 activation by inducing nucleotide binding to Apaf-1. J Biol Chem 275:31199–31203

    Article  PubMed  CAS  Google Scholar 

  39. Hsu TC, Wu WJ, Chen MC, Tsay GJ (2004) Human parvovirus B19 non-structural protein (NS1) induces apoptosis through mitochondria cell death pathway in COS-7 cells. Scand J Infect Dis 36:570–577

    Article  PubMed  CAS  Google Scholar 

  40. Poole BD, Zhou J, Grote A, Schiffenbauer A, Naides SJ (2006) Apoptosis of liver-derived cells induced by parvovirus B19 nonstructural protein. J Virol 80:4114–4121

    Article  PubMed  CAS  Google Scholar 

  41. Olsson M, Vakifahmetoglu H, Abruzzo PM, Högstrand K, Grandien A, Zhivotovsky B (2009) DISC-mediated activation of caspase-2 in DNA damage-induced apoptosis. Oncogene 28:1949–1959

    Article  PubMed  CAS  Google Scholar 

  42. Robertson JD, Enoksson M, Suomela M, Zhivotovsky B, Orrenius S (2002) Caspase-2 acts upstream of mitochondria to promote cytochrome c release during etoposide-induced apoptosis. J Biol Chem 277:29803–29809

    Article  PubMed  CAS  Google Scholar 

  43. Panaretakis T, Laane E, Pokrovskaja K, Björklund AC, Moustakas A, Zhivotovsky B et al (2005) Doxorubicin requires the sequential activation of caspase-2, protein kinase Cdelta, and c-Jun NH2-terminal kinase to induce apoptosis. Mol Biol Cell 16:3821–3831

    Article  PubMed  CAS  Google Scholar 

  44. Arnaudeau C, Lundin C, Helleday T (2001) DNA double-strand breaks associated with replication forks are predominantly repaired by homologous recombination involving an exchange mechanism in mammalian cells. J Mol Biol 307:1235–1245

    Article  PubMed  CAS  Google Scholar 

  45. Roos WP, Kaina B (2006) DNA damage-induced cell death by apoptosis. Trends Mol Med 12:440–450

    Article  PubMed  CAS  Google Scholar 

  46. Hsieh YH, Su IJ, Wang HC, Chang WW, Lei HY, Lai MD et al (2004) Pre-S mutant surface antigens in chronic hepatitis B virus infection induce oxidative stress and DNA damage. Carcinogenesis 25:2023–2032

    Article  PubMed  CAS  Google Scholar 

  47. Mori K (2000) Tripartite management of unfolded proteins in the endoplasmic reticulum. Cell 101:451–454

    Article  PubMed  CAS  Google Scholar 

  48. Harding HP, Zhang Y, Bertolotti A, Zeng H, Ron D (2000) Perk is essential for translational regulation and cell survival during the unfolded protein response. Mol Cell 5:897–904

    Article  PubMed  CAS  Google Scholar 

  49. Oyadomari S, Mori M (2004) Roles of CHOP/GADD153 in endoplasmic reticulum stress. Cell Death Differ 11:381–389

    Article  PubMed  CAS  Google Scholar 

  50. Szegezdi E, Logue SE, Gorman AM, Samali A (2006) Mediators of endoplasmic reticulum stress-induced apoptosis. EMBO Rep 7:880–885

    Article  PubMed  CAS  Google Scholar 

  51. Hitomi J, Katayama T, Eguchi Y, Kudo T, Taniguchi M, Koyama Y (2004) Involvement of caspase-4 in endoplasmic reticulum stress-induced apoptosis and Abeta-induced cell death. J Cell Biol 165:347–356

    Article  PubMed  CAS  Google Scholar 

  52. Kim SJ, Zhang Z, Hitomi E, Lee YC, Mukherjee AB (2006) Endoplasmic reticulum stress-induced caspase-4 activation mediates apoptosis and neurodegeneration in INCL. Hum Mol Genet 15:1826–1834

    Article  PubMed  CAS  Google Scholar 

  53. Krumschnabel G, Sohm B, Bock F, Manzl C, Villunger A (2009) The enigma of caspase-2: the laymen’s view. Cell Death Differ 16:195–207

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgment

This study was supported by the National Nuclear R&D Program of the Ministry of Education, Science, and Technology (MEST) of the Republic of Korea.

Author information

Authors and Affiliations

  1. Division of Radiation Cancer Research, Korea Institute of Radiological & Medical Sciences, 215-4 Gongneung-dong, Nowon-Gu, Seoul, 139-706, Korea

    Byeong Mo Kim, Eun Jong Han & Sung Hee Hong

  2. Department of Chemistry, College of National Science, Kongju National University, Gongju, Chungnam, 314-701, Korea

    Ambadas B. Rode & In Seok Hong

Authors
  1. Byeong Mo Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ambadas B. Rode
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Eun Jong Han
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. In Seok Hong
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sung Hee Hong
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Sung Hee Hong.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kim, B.M., Rode, A.B., Han, E.J. et al. 5-Phenylselenyl- and 5-methylselenyl-methyl-2′-deoxyuridine induce oxidative stress, DNA damage, and caspase-2-dependent apoptosis in cancer cells. Apoptosis 17, 200–216 (2012). https://doi.org/10.1007/s10495-011-0665-2

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10495-011-0665-2

Keywords

Search

Navigation