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MicroRNA-128 downregulates Bax and induces apoptosis in human embryonic kidney cells

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Abstract

MicroRNAs (miRNAs) are short ~21-nt non-coding RNA molecules that have been shown to regulate a number of biological processes. Previous reports have shown that overexpression of miR-128 in glioma cells inhibited cell proliferation. Literature also suggests that miR-128 negatively regulates prostate cancer cell invasion. Here, we show that overexpression of hsa-miR-128, a brain-enriched microRNA, induces apoptosis in HEK293T cells as elucidated by apoptosis assay, cell cycle changes, loss of mitochondrial membrane potential and multicaspase assay. By in silico analysis, we identified a putative target site within the 3′ untranslated region (UTR) of Bax, a proapoptotic member of the apoptosis pathway. We found that ectopic expression of hsa-miR-128 suppressed a luciferase reporter containing the Bax-3′ UTR and reduced the levels of Bax in HEK293T cells. Taken together, our study demonstrates that overexpression of hsa-miR-128 not only induces apoptosis in HEK293T cells but also is an endogenous regulator of Bax protein.

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Abbreviations

DHE:

Dihydroethidium

ROS:

Reactive oxygen species

DiOC6:

3,3′ Dihexyloxacarbocyanine iodide

Δψm :

Mitochondrial membrane potential

SR-VAD-FMK:

Sulforhodamine-valyl-alanyl-aspartyl-fluoromethyl-ketone

endoG:

Endonuclease G

References

  1. Pillai RS (2005) MicroRNA function: multiple mechanisms for a tiny RNA? RNA 11:1753–1761

    Article  PubMed  CAS  Google Scholar 

  2. Zamore PD, Haley B (2005) Ribo-gnome: the big world of small RNAs. Science 309:1519–1524

    Article  PubMed  CAS  Google Scholar 

  3. Yu Z, Jian Z, Shen SH, Purisima E, Wang E (2007) Global analysis of microRNA target gene expression reveals that miRNA targets are lower expressed in mature mouse and Drosophila tissues than in the embryos. Nucleic Acids Res 35:152–164

    Article  PubMed  CAS  Google Scholar 

  4. Karp X, Ambros V (2005) Developmental biology. Encountering microRNAs in cell fate signaling. Science 310:1288–1289

    Article  PubMed  CAS  Google Scholar 

  5. Chen CZ, Li L, Lodish HF, Bartel DP (2004) MicroRNAs modulate hematopoietic lineage differentiation. Science 303:83–86

    Article  PubMed  CAS  Google Scholar 

  6. Cheng AM, Byrom MW, Shelton J, Ford LP (2005) Antisense inhibition of human miRNAs and indications for an involvement of miRNA in cell growth and apoptosis. Nucleic Acids Res 33:1290–1297

    Article  PubMed  CAS  Google Scholar 

  7. Lu S, Sun YH, Shi R, Clark C, Li L, Chiang VL (2005) Novel and mechanical stressresponsive MicroRNAs in Populus trichocarpa that are absent from Arabidopsis. Plant Cell 17:2186–2203

    Article  PubMed  CAS  Google Scholar 

  8. Garzon R, Calin GA, Croce CM (2009) MicroRNAs in Cancer. Annu Rev Med 60:167–179

    Article  PubMed  CAS  Google Scholar 

  9. Ciafrè SA, Galardi S, Mangiola A, Ferracin M, Liu CG, Sabatino G, Negrini M, Maira G, Croce CM, Farace MG (2005) Extensive modulation of a set of microRNAs in primary glioblastoma. Biochem Biophys Res Commun 334:1351–1358

    Article  PubMed  Google Scholar 

  10. Myatt SS, Wang J, Monteiro LJ, Christian M, Ho KK, Fusi L, Dina RE, Brosens JJ, Ghaem-Maghami S, Lam EW (2010) Definition of microRNAs that repress expression of the tumor suppressor gene FOXO1 in endometrial cancer. Cancer Res 70:367–377

    Article  PubMed  CAS  Google Scholar 

  11. Evangelisti C, Florian MC, Massimi I, Dominici C, Giannini G, Galardi S, Buè MC, Massalini S, McDowell HP, Messi E, Gulino A, Farace MG, Ciafrè SA (2009) MiR-128 up-regulation inhibits Reelin and DCX expression and reduces neuroblastoma cell motility and invasiveness. FASEB J 23:4276–4287

    Article  PubMed  CAS  Google Scholar 

  12. Zhang Y, Chao T, Li R, Liu W, Chen Y, Yan X, Gong Y, Yin B, Liu W, Qiang B, Zhao J, Yuan J, Peng X (2009) MicroRNA-128 inhibits glioma cells proliferation by targeting transcription factor E2F3a. J Mol Med 87:43–51

    Article  PubMed  CAS  Google Scholar 

  13. Khan AP, Poisson LM, Bhat VB, Fermin D, Zhao R, Kalyana-Sundaram S, Michailidis G, Nesvizhskii AI, Omenn GS, Chinnaiyan AM, Sreekumar A (2010) Quantitative proteomic profiling of prostate cancer reveals a role for miR-128 in prostate cancer. Mol Cell Proteomics 9:298–312

    Article  PubMed  CAS  Google Scholar 

  14. Cory S, Adams JM (2002) The Bcl2 family: regulators of the cellular life-or-death switch. Nat Rev Cancer 2:647–656

    Article  PubMed  CAS  Google Scholar 

  15. Ott M, Norberg E, Zhivotovsky B, Orrenius S (2009) Mitochondrial targeting of tBid/BAX protein: a role for the TOM complex? Cell Death Differ 16:1075–1082

    Article  PubMed  CAS  Google Scholar 

  16. Khanna N, Jayaram HN, Singh N (2004) Benzamide riboside induced mitochondrial mediated apoptosis in human lung cancer H520 cells. Life Sci 75:179–190

    Article  PubMed  CAS  Google Scholar 

  17. Su YT, Chang HL, Shyue SK, Hsu SL (2005) Emodin induces apoptosis in human lung adenocarcinoma cells through a reactive oxygen species-dependent mitochondrial signaling pathway. Biochem Pharmacol 70:229–241

    Article  PubMed  CAS  Google Scholar 

  18. John B, Enright AJ, Aravin A, Tuschl T, Sander C, Marks DS (2004) Human microRNA targets. PLoS Biol 2:e363

    Article  PubMed  Google Scholar 

  19. Rehmsmeier M, Steffen P, Hochsmann M, Giegerich R (2004) Fast and effective prediction of microRNA/target duplexes. RNA 10:1507–1517

    Article  PubMed  CAS  Google Scholar 

  20. Lewis BP, Shih IH, Jones-Rhoades MW, Bartel DP, Burge CB (2003) Prediction of mammalian microRNA targets. Cell 115:787–798

    Article  PubMed  CAS  Google Scholar 

  21. Pfaffl MW (2001) A new mathematical model for relative quantification in realtime RT-PCR. Nucleic Acid Res 29:2002–2007

    Article  Google Scholar 

  22. Radovic N, Cucic S, Altarac S (2008) Molecular aspects of apoptosis. Acta Med Croatica 62:249–256

    PubMed  CAS  Google Scholar 

  23. Orrenius S (2007) Reactive oxygen species in mitochondria-mediated cell death. Drug Metab Rev 39:443–455

    Article  PubMed  CAS  Google Scholar 

  24. Wei MC, Zong WX, Cheng EH, Lindsten T, Panoutsakopoulou V, Ross AJ, Roth KA, MacGregor GR, Thompson CB, Korsmeyer SJ (2001) Proapoptotic BAX and BAK: a requisite gateway to mitochondrial dysfunction and death. Science 292:727–730

    Article  PubMed  CAS  Google Scholar 

  25. Godlewski J, Nowicki MO, Bronisz A, Williams S, Otsuki A, Nuovo G, Raychaudhury A, Newton HB, Chiocca EA, Lawler S (2008) Targeting of the Bmi-1 oncogene/stem cell renewal factor by MicroRNA-128 inhibits glioma proliferation and self-renewal. Cancer Res 68:9125–9130

    Article  PubMed  CAS  Google Scholar 

  26. Cui JG, Zhao Y, Sethi P, Li YY, Mahta A, Culicchia F, Lukiw WJ (2009) Micro-RNA-128 (miRNA-128) down-regulation in glioblastoma targets ARP5 (ANGPTL6), Bmi-1 and E2F-3a, key regulators of brain cell proliferation. J Neurooncol. doi:10.1007/s11060-009-0077-0

  27. Weiss GJ, Bemis LT, Nakajima E, Sugita M, Birks DK, Robinson WA, Varella-Garcia M, Bunn PA Jr, Haney J, Helfrich BA, Kato H, Hirsch FR, Franklin WA (2008) EGFR regulation by microRNA in lung cancer: correlation with clinical response and survival to gefitinib and EGFR expression in cell lines. Ann Oncol 19:1053–1059

    Article  PubMed  CAS  Google Scholar 

  28. Andreas G (2003) Introduction to apoptosis. Aporeview 1–26

  29. Andre N, Rome A, Carre M (2006) Antimitochondrial agents: a new class of anticancer agents. Arch Pediatr 13:69–75

    Article  PubMed  CAS  Google Scholar 

  30. Bustamante J, Caldas Lopes E, Garcia M, Di Libero E, Alvarez E, Hajos SE (2004) Disruption of mitochondrial membrane potential during apoptosis induced by PSC 833 and CsA in multidrug-resistant lymphoid leukemia. Toxicol Appl Pharmacol 199:44–51

    Article  PubMed  CAS  Google Scholar 

  31. Lewis J, Oyler GA, Ueno K, Fannjiang YR, Chau BN, Vornov J, Korsmeyer SJ, Zou S, Hardwick JM (1999) Inhibition of virus-induced neuronal apoptosis by BAX protein. Nat Med 5:832–835

    Article  PubMed  CAS  Google Scholar 

  32. Zhang L, Yu J, Park BH, Kinzler KW, Vogelstein B (2000) Role of BAX in the apoptotic response to anticancer agents. Science 290:989–992

    Article  PubMed  CAS  Google Scholar 

  33. Pearson AS, Spitz FR, Swisher SG, Kataoka M, Sarkiss MG, Meyn RE, McDonnell TJ, Cristiano RJ, Roth JA (2000) Up-regulation of the proapoptotic mediators BAX and Bak after Adenovirus-mediated p53 gene transfer in lung cancer cells. Clin Cancer Res 6:887–890

    PubMed  CAS  Google Scholar 

  34. Sethi P, Lukiw WJ (2009) Micro-RNA abundance and stability in human brain: specific alterations in Alzheimer’s disease temporal lobe neocortex. Neurosci Lett 459:100–104

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

This work was supported by Grants NWP0036 from the Council of Scientific and industrial research (CSIR). Y.K.A. was supported with fellowship from CSIR. The authors thank Prof. Samir K. Brahmachari for project conceptualization. We also acknowledge Anita Goel, Ravindresh Chhabra, Richa Singh, Richa Dubey and Shruti Chowdhari for their help.

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Correspondence to Neeru Saini.

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Adlakha, Y.K., Saini, N. MicroRNA-128 downregulates Bax and induces apoptosis in human embryonic kidney cells. Cell. Mol. Life Sci. 68, 1415–1428 (2011). https://doi.org/10.1007/s00018-010-0528-y

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  • DOI: https://doi.org/10.1007/s00018-010-0528-y

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