Skip to main content

Advertisement

SpringerLink
Log in

RNase MC2: a new Momordica charantia ribonuclease that induces apoptosis in breast cancer cells associated with activation of MAPKs and induction of caspase pathways

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

Abstract

Ribonucleases (RNases) are ubiquitously distributed nucleases that cleave RNA into smaller pieces. They are promising drugs for different cancers based on their concrete antitumor activities in vitro and in vivo. Here we report for the first time purification and characterization of a 14-kDa RNase, designated as RNase MC2, in the seeds of bitter gourd (Momordica charantia). RNase MC2 manifested potent RNA-cleavage activity toward baker’s yeast tRNA, tumor cell rRNA, and an absolute specificity for uridine. RNase MC2 demonstrated both cytostatic and cytotoxic activities against MCF-7 breast cancer cells. Treatment of MCF-7 cells with RNase MC2 caused nuclear damage (karyorrhexis, chromatin condensation, and DNA fragmentation), ultimately resulting in early/late apoptosis. Further molecular studies unveiled that RNase MC2 induced differential activation of MAPKs (p38, JNK and ERK) and Akt. On the other hand, RNase MC2 exposure activated caspase-8, caspase-9, caspase-7, increased the production of Bak and cleaved PARP, which in turn contributed to the apoptotic response. In conclusion, RNase MC2 is a potential agent which can be exploited in the worldwide fight against breast cancer.

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.

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

Similar content being viewed by others

Abbreviations

RNase MC2:

The second ribonuclease reported from Momordica charantia

MAPKs:

Mitogen-activated protein kinases

BG:

Bitter gourd (Momordica charantia)

References

  1. Deshpande RA, Shankar V (2002) Ribonucleases from T2 family. Crit Rev Microbiol 28:79–122

    Article  PubMed  CAS  Google Scholar 

  2. Cuchillo CM, Pares X, Guasch A, Barman T, Travers F, Nogues MV (1993) The role of 2′,3′-cyclic phosphodiesters in the bovine pancreatic ribonuclease A catalysed cleavage of RNA: intermediates or products? FEBS Lett 333:207–210

    Article  PubMed  CAS  Google Scholar 

  3. Thompson JE, Raines RT (1994) Value of general acid–base catalysis to ribonuclease A. J Am Chem Soc 116:5467–5468

    Article  PubMed  CAS  Google Scholar 

  4. Wilson CM (1975) Plant nucleases. Ann Rev Plant Physiol 26:187–208

    Article  CAS  Google Scholar 

  5. Aravind L, Koonin EV (2001) A natural classification of ribonucleases. Methods Enzymol 341:3–28

    Article  PubMed  CAS  Google Scholar 

  6. Fang EF, Ng TB (2011) Ribonucleases of different origins with a wide spectrum of medicinal applications. Biochim Biophys Acta Rev Cancer 1815:65–74

    CAS  Google Scholar 

  7. Rybak SM, Newton DL (1999) Natural and engineered cytotoxic ribonucleases: therapeutic potential. Exp Cell Res 253:325–335

    Article  PubMed  CAS  Google Scholar 

  8. Rutkoski TJ, Kink JA, Strong LE, Schilling CI, Raines RT (2010) Antitumor activity of ribonuclease multimers created by site-specific covalent tethering. Bioconj Chem 21:1691–1702

    Article  CAS  Google Scholar 

  9. Castro J, Ribo M, Navarro S, Nogues MV, Vilanova M, Benito A (2011) A human ribonuclease induces apoptosis associated with p21WAF1/CIP1 induction and JNK inactivation. BMC Cancer 11:9

    Article  PubMed  CAS  Google Scholar 

  10. Fang EF, Zhang CZ, Ng TB, Wong JH, Pan WL, Ye XJ et al. (2011) Momordica charantia lectin, a type II ribosome inactivating protein, exhibits antitumor activity toward human nasopharyngeal carcinoma cells in vitro and in vivo. Cancer Prev Res (Phila)

  11. Fang EF, Ng TB (2011) Bitter gourd (Momordica charantia) is a cornucopia of health: a review of its credited antidiabetic, anti-HIV, and antitumor properties. Curr Mol Med 11:417–436

    Article  PubMed  CAS  Google Scholar 

  12. Fang EF, Wong JH, Lin P, Ng TB (2010) Biochemical characterization of the RNA-hydrolytic activity of a pumpkin 2S albumin. FEBS Lett 584:4089–4096

    Article  PubMed  CAS  Google Scholar 

  13. Fang EF, Wong JH, Bah CS, Lin P, Tsao SW, Ng TB (2010) Bauhinia variegata var. variegata trypsin inhibitor: from isolation to potential medicinal applications. Biochem Biophys Res Commun 396:806–811

    Article  PubMed  CAS  Google Scholar 

  14. Fang EF, Wong JH, Ng TB (2010) Thermostable Kunitz trypsin inhibitor with cytokine inducing, antitumor and HIV-1 reverse transcriptase inhibitory activities from Korean large black soybeans. J Biosci Bioeng 109:211–217

    Article  PubMed  CAS  Google Scholar 

  15. Singh A, Ai Y, Kao TH (1991) Characterization of ribonuclease activity of three S-allele-associated proteins of Petunia inflata. Plant Physiol 96:61–68

    Article  PubMed  CAS  Google Scholar 

  16. Xing D, Ni S, Kennedy MA, Li QQ (2009) Identification of a plant-specific Zn2+-sensitive ribonuclease activity. Planta 230:819–825

    Article  PubMed  CAS  Google Scholar 

  17. Fang EF, Pan WL, Wong JH, Chan YS, Ye XJ, Ng TB (2011) A new Phaseolus vulgaris lectin induces selective toxicity on human liver carcinoma Hep G2 cells. Arch Toxicol 85:1551–1563

    Article  Google Scholar 

  18. Zhang CZ, Zhang HT, Chen GG, Lai PB (2011) Trichostatin A sensitizes HBx-expressing liver cancer cells to etoposide treatment. Apoptosis 16:683–695

    Article  PubMed  CAS  Google Scholar 

  19. Neumann GM, Condron R, Polya GM (1996) Purification and sequencing of napin-like protein small and large chains from Momordica charantia and Ricinus communis seeds and determination of sites phosphorylated by plant Ca(2+)-dependent protein kinase. Biochim Biophys Acta 1298:223–240

    Article  PubMed  CAS  Google Scholar 

  20. Vashishta A, Sahu T, Sharma A, Choudhary SK, Dixit A (2006) In vitro refolded napin-like protein of Momordica charantia expressed in Escherichia coli displays properties of native napin. Biochim Biophys Acta 1764:847–855

    PubMed  CAS  Google Scholar 

  21. Irwin SD, Keen JN, Findlay JB, Lord JM (1990) The Ricinus communis 2S albumin precursor: a single preproprotein may be processed into two different heterodimeric storage proteins. Mol Gen Genet 222:400–408

    Article  PubMed  CAS  Google Scholar 

  22. Mansouri A, Ridgway LD, Korapati AL, Zhang Q, Tian L, Wang Y et al (2003) Sustained activation of JNK/p38 MAPK pathways in response to cisplatin leads to Fas ligand induction and cell death in ovarian carcinoma cells. J Biol Chem 278:19245–19256

    Article  PubMed  CAS  Google Scholar 

  23. Lam SK, Ng TB (2010) First report of a haemagglutinin-induced apoptotic pathway in breast cancer cells. Biosci Rep 30:307–317

    Article  PubMed  CAS  Google Scholar 

  24. Riedl SJ, Shi Y (2004) Molecular mechanisms of caspase regulation during apoptosis. Natl Rev Mol Cell Biol 5:897–907

    Article  CAS  Google Scholar 

  25. Maddika S, Ande SR, Panigrahi S, Paranjothy T, Weglarczyk K, Zuse A et al (2007) Cell survival, cell death and cell cycle pathways are interconnected: implications for cancer therapy. Drug Resist Updat 10:13–29

    Article  PubMed  CAS  Google Scholar 

  26. Korsmeyer SJ, Wei MC, Saito M, Weiler S, Oh KJ, Schlesinger PH (2000) Pro-apoptotic cascade activates BID, which oligomerizes BAK or BAX into pores that result in the release of cytochrome c. Cell Death Differ 7:1166–1173

    Article  PubMed  CAS  Google Scholar 

  27. Numata T, Suzuki A, Kakuta Y, Kimura K, Yao M, Tanaka I et al (2003) Crystal structures of the ribonuclease MC1 mutants N71T and N71S in complex with 5′-GMP: structural basis for alterations in substrate specificity. Biochemistry 42:5270–5278

    Article  PubMed  CAS  Google Scholar 

  28. Schmitz KH, Speck RM (2010) Risks and benefits of physical activity among breast cancer survivors who have completed treatment. Womens Health (Lond Engl) 6:221–238

    Google Scholar 

  29. Tseng HH, Yu YL, Chen YL, Chen JH, Chou CL, Kuo TY et al (2011) RC-RNase-induced cell death in estrogen receptor positive breast tumors through down-regulation of Bcl-2 and estrogen receptor. Oncol Rep 25:849–853

    Article  PubMed  CAS  Google Scholar 

  30. Chang CH, Gupta P, Michel R, Loo M, Wang Y, Cardillo TM et al (2010) Ranpirnase (frog RNase) targeted with a humanized, internalizing, anti-Trop-2 antibody has potent cytotoxicity against diverse epithelial cancer cells. Mol Cancer Ther 9:2276–2286

    Article  PubMed  CAS  Google Scholar 

  31. Balandin TG, Edelweiss E, Andronova NV, Treshalina EM, Sapozhnikov AM, Deyev SM (2011) Antitumor activity and toxicity of anti-HER2 immunoRNase scFv 4D5-dibarnase in mice bearing human breast cancer xenografts. Invest New Drugs 29:22–32

    Article  PubMed  CAS  Google Scholar 

  32. Dhanasekaran DN, Johnson GL (2007) MAPKs: function, regulation, role in cancer and therapeutic targeting. Oncogene 26:3097–3099

    Article  PubMed  CAS  Google Scholar 

  33. Dasmahapatra G, Lembersky D, Kramer L, Fisher RI, Friedberg J, Dent P et al (2010) The pan-HDAC inhibitor vorinostat potentiates the activity of the proteasome inhibitor carfilzomib in human DLBCL cells in vitro and in vivo. Blood 115:4478–4487

    Article  PubMed  CAS  Google Scholar 

  34. Chen J, Rusnak M, Luedtke RR, Sidhu A (2004) D1 dopamine receptor mediates dopamine-induced cytotoxicity via the ERK signal cascade. J Biol Chem 279:39317–39330

    Article  PubMed  CAS  Google Scholar 

  35. Wilhelm SM, Carter C, Tang L, Wilkie D, McNabola A, Rong H et al (2004) BAY 43-9006 exhibits broad spectrum oral antitumor activity and targets the RAF/MEK/ERK pathway and receptor tyrosine kinases involved in tumor progression and angiogenesis. Cancer Res 64:7099–7109

    Article  PubMed  CAS  Google Scholar 

  36. Iordanov MS, Wong J, Newton DL, Rybak SM, Bright RK, Flavell RA et al (2000) Differential requirement for the stress-activated protein kinase/c-Jun NH(2)-terminal kinase in RNAdamage-induced apoptosis in primary and in immortalized fibroblasts. Mol Cell Biol Res Commun 4:122–128

    Article  PubMed  CAS  Google Scholar 

  37. Song G, Ouyang G, Bao S (2005) The activation of Akt/PKB signaling pathway and cell survival. J Cell Mol Med 9:59–71

    Article  PubMed  CAS  Google Scholar 

  38. Meier F, Busch S, Lasithiotakis K, Kulms D, Garbe C, Maczey E et al (2007) Combined targeting of MAPK and AKT signalling pathways is a promising strategy for melanoma treatment. Br J Dermatol 156:1204–1213

    Article  PubMed  CAS  Google Scholar 

  39. Kinkade CW, Castillo-Martin M, Puzio-Kuter A, Yan J, Foster TH, Gao H et al (2008) Targeting AKT/mTOR and ERK MAPK signaling inhibits hormone-refractory prostate cancer in a preclinical mouse model. J Clin Invest 118:3051–3064

    PubMed  CAS  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  41. Bailey K, Cook HW, McMaster CR (2005) The phospholipid scramblase PLSCR1 increases UV induced apoptosis primarily through the augmentation of the intrinsic apoptotic pathway and independent of direct phosphorylation by protein kinase C delta. Biochim Biophys Acta 1733:199–209

    PubMed  CAS  Google Scholar 

  42. Semenov DV, Aronov PA, Kuligina EV, Potapenko MO, Richter VA (2004) Oligonucleosome DNA fragmentation of caspase 3 deficient MCF-7 cells in palmitate-induced apoptosis. Nucleosides Nucleotides Nucleic Acids 23:831–836

    Article  PubMed  CAS  Google Scholar 

  43. Mc Gee MM, Hyland E, Campiani G, Ramunno A, Nacci V, Zisterer DM (2002) Caspase-3 is not essential for DNA fragmentation in MCF-7 cells during apoptosis induced by the pyrrolo-1,5-benzoxazepine, PBOX-6. FEBS Lett 515:66–70

    Article  PubMed  CAS  Google Scholar 

  44. Fernandes-Alnemri T, Takahashi A, Armstrong R, Krebs J, Fritz L, Tomaselli KJ et al (1995) Mch3, a novel human apoptotic cysteine protease highly related to CPP32. Cancer Res 55:6045–6052

    PubMed  CAS  Google Scholar 

  45. Fang EF, Ng TB (2011) Ribonucleases of different origins with a wide spectrum of medicinal applications. Biochim Biophys Acta 1815:65–74

    PubMed  CAS  Google Scholar 

  46. Hu CC, Tang CH, Wang JJ (2001) Caspase activation in response to cytotoxic Rana catesbeiana ribonuclease in MCF-7 cells. FEBS Lett 503:65–68

    Article  PubMed  CAS  Google Scholar 

  47. Zong WX, Li C, Hatzivassiliou G, Lindsten T, Yu QC, Yuan J et al (2003) Bax and Bak can localize to the endoplasmic reticulum to initiate apoptosis. J Cell Biol 162:59–69

    Article  PubMed  CAS  Google Scholar 

  48. Parkash A, Ng TB, Tso WW (2002) Purification and characterization of charantin, a napin-like ribosome-inactivating peptide from bitter gourd (Momordica charantia) seeds. J Pept Res 59:197–202

    Article  PubMed  CAS  Google Scholar 

Download references

Conflicts of interest

The authors declare that they have no conflicts of interest.

Author information

Authors and Affiliations

  1. School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong

    Evandro Fei Fang & Tzi Bun Ng

  2. State Key Laboratory of Oncology in Southern China, Sun Yat-Sen University Cancer Center, Guangzhou, China

    Chris Zhi Yi Zhang

  3. Department of Pathology, Sun Yat-Sen University Cancer Center, Guangzhou, China

    Chris Zhi Yi Zhang

  4. School of Life Sciences, Faculty of Science, The Chinese University of Hong Kong, Shatin, Hong Kong

    Wing Ping Fong

  5. Choh Ming Li Basic Medical Sciences Building, The Chinese University of Hong Kong, Shatin, Hong Kong

    Tzi Bun Ng

Authors
  1. Evandro Fei Fang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Chris Zhi Yi Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Wing Ping Fong
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Tzi Bun Ng
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Tzi Bun Ng.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Fang, E.F., Zhang, C.Z.Y., Fong, W.P. et al. RNase MC2: a new Momordica charantia ribonuclease that induces apoptosis in breast cancer cells associated with activation of MAPKs and induction of caspase pathways. Apoptosis 17, 377–387 (2012). https://doi.org/10.1007/s10495-011-0684-z

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10495-011-0684-z

Keywords

Search

Navigation