Skip to main content
SpringerLink
Log in

Nm23-H1 Protein Binds to APE1 at AP Sites and Stimulates AP Endonuclease Activity Following Ionizing Radiation of the Human Lung Cancer A549 Cells

  • Original Paper
  • Published:
Cell Biochemistry and Biophysics Aims and scope Submit manuscript

Abstract

Non-metastatic protein-23 homolog-1 (Nm23-H1) is a multifunctional protein with DNase and histidine protein kinase activities. Human apurinic endonuclease-1 (APE1) is the AP endonuclease DNA base excision repair (BER) enzyme involved in several important cellular functions. Since the relationship between Nm23-H1 and APE1 proteins is unclear, we evaluated their interaction at different time points after irradiating human lung cancer A549 cells with X-rays. We found that Nm23-H1 and APE1 overexpression was induced by irradiation in a dose- and time-dependent manner. Subcellular distribution pattern of both proteins was reversed after irradiation. After irradiation, APE1 that initially showed nuclear localization was gradually increased in the cytoplasm, whereas Nm23-H1 that mainly showed cytoplasmic localization was gradually increased in the nuclei of A549 cells. Nm23-H1 and APE1 interaction was demonstrated by His-pull-down and co-immunoprecipitation assays. The presence of Nm23-H1/APE1 complex in X-ray-irradiated A549 cells was also detected by DNA affinity precipitation analysis of a DNA fragment containing an AP site. Although the AP endonuclease activity of Nm23-H1 was too weak to be detected, the AP endonuclease activity of APE1 was increased with the enhanced Nm23-H1 expression. In conclusion, our data point to a mechanism by which Nm23-H1 protects cells against oxidative stress through the engagement of DNA BER enzyme APE1.

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

References

  1. Jin, L., Liu, G., Zhang, C. H., et al. (2009). Nm23–H1 regulates the proliferation and differentiation of the human chronic myeloid leukemia K562 cell line: A functional proteomics study. Life Sciences, 84, 458–467.

    Article  PubMed  CAS  Google Scholar 

  2. Jung, H., Seong, H. A., & Ha, H. (2007). Nm23–H1 tumor suppressor and its interacting partner STRAP activate p53 function. The Journal of Biological Chemistry, 282, 35293–35307.

    Article  PubMed  CAS  Google Scholar 

  3. Biggs, J., Hersperger, E., Steeg, P. S., Liotta, L. A., & Shearn, A. A. (1990). Drosophila gene that is homologous to a mammalian gene associated with tumor metastasis codes for a nucleoside diphosphate kinase. Cell, 63, 933–940.

    Article  PubMed  CAS  Google Scholar 

  4. Hartsough, M. T., Morrison, D. K., Salerno, M., et al. (2002). Nm23–H1 metastasis suppressor of Ras via a histidine protein kinase pathway. The Journal of Biological Chemistry, 277, 32389–32399.

    Article  PubMed  CAS  Google Scholar 

  5. Bosnar, M. H., Dubravcˇic′, K., Bago, R., & Pavelic′, J. (2008). Head and neck tumor cells exhibit altered proliferation upon overexpression of nm23 genes. Croatica Chemica Acta, 81, 183–189.

    CAS  Google Scholar 

  6. Cipollini, G., Berti, A., Fiore, L., et al. (1997). Down-regulation of the nm23–H1 gene inhibits cell proliferation. International Journal of Cancer, 73, 297–302.

    Article  CAS  Google Scholar 

  7. Kimura, N., Shimada, N., Fukuda, M., et al. (2000). Regulation of cellular functions by nucleoside diphosphate kinases in mammals. Journal of Bioenergetics and Biomembranes, 32, 309–315.

    Article  PubMed  CAS  Google Scholar 

  8. Parlanti, E., Locatelli, G., Maga, G., & Dogliotti, E. (2007). Human base excision repair complex is physically associated to DNA replication and cell cycle regulatory proteins. Nucleic Acids Research, 35, 1569–1577.

    Article  PubMed  CAS  Google Scholar 

  9. Wilson, D. M., 3rd, & Bohr, V. A. (2007). The mechanics of base excision repair, and its relationship to aging and disease. DNA Repair, 6, 544–559.

    Article  PubMed  CAS  Google Scholar 

  10. Evans, A. R., Limp-Foster, M., & Kelley, M. R. (2000). Going APE over ref-1. Mutation Research, 461, 83–108.

    PubMed  CAS  Google Scholar 

  11. Chowdhury, D., Beresford, P. J., Zhu, P., et al. (2006). The exonuclease TREX1 is in the SET complex and acts in concert with NM23–H1 to degrade DNA during granzyme A-mediated cell death. Molecular Cell, 23, 133–142.

    Article  PubMed  CAS  Google Scholar 

  12. Fan, Z., Beresford, P. J., Zhang, D., et al. (2003). Cleaving the oxidative repair protein Ape1 enhances cell death mediated by granzyme A. Nature Immunology, 4, 145–153.

    Article  PubMed  CAS  Google Scholar 

  13. Yan, N., Cherepanov, P., Daigle, J. E., Engelman, A., & Lieberman, J. (2009). The SET complex acts as a barrier to autointegration of HIV-1. PLoS Pathogens, 5, e1000327.

    Article  PubMed  Google Scholar 

  14. Chowdhury, D., & Lieberman, J. (2008). Death by a thousand cuts: Granzyme pathways of programmed cell death. Annual Review of Immunology, 26, 389–420.

    Article  PubMed  CAS  Google Scholar 

  15. Zhao, Y., Seefeldt, T., Chen, W., et al. (2009). Increase in thiol oxidative stress via glutathione reductase inhibition as a novel approach to enhance cancer sensitivity to X-ray irradiation. Free Radical Biology and Medicine, 47, 176–183.

    Article  PubMed  CAS  Google Scholar 

  16. Zhou, Q., Yang, X., Zhu, D., et al. (2007). Double mutant P96S/S120G of Nm23–H1 abrogates its NDPK activity and motility-suppressive ability. Biochemical and Biophysical Research Communications, 356, 348–353.

    Article  PubMed  CAS  Google Scholar 

  17. Dai, N., Wang, D., Li, M. X., et al. (2009). Construction, purification and functional characterization of hAPE1 and fusion protein. Acta Academiae Medicinae Militaris Tertiae, 31, 1123–1126.

    CAS  Google Scholar 

  18. Li, M. X., Wang, D., Zhong, Z. Y., et al. (2008). Targeting truncated APE1 in mitochondria enhances cell survival after oxidative stress. Free Radical Biology and Medicine, 45, 592–601.

    Article  PubMed  CAS  Google Scholar 

  19. Wang, D., Xiang, D. B., Yang, X. Q., et al. (2009). APE1 overexpression is associated with cisplatin resistance in non-small cell lung cancer and targeted inhibition of APE1 enhances the activity of cisplatin in A549 cells. Lung Cancer, 66, 298–304.

    Article  PubMed  Google Scholar 

  20. Xiang, D. B., Chen, Z. T., Wang, D., et al. (2008). Chimeric adenoviral vector Ad5/F35-mediated APE1 siRNA enhances sensitivity of human colorectal cancer cells to radiotherapy in vitro and in vivo. Cancer Gene Therapy, 15, 625–635.

    Article  PubMed  CAS  Google Scholar 

  21. Wang, Y. F., Chen, J. Y., Chang, S. Y., et al. (2008). Nm23–H1 expression of metastatic tumors in the lymph nodes is a prognostic indicator of oral squamous cell carcinoma. International Journal of Cancer, 122, 377–386.

    Article  CAS  Google Scholar 

  22. Neyton, S., Lespinasse, F., Lahaye, F., Staccini, P., Paquis-Flucklinger, V., & Santucci-Darmanin, S. (2007). CRM1-dependent nuclear export and dimerization with hMSH5 contribute to the regulation of hMSH4 subcellular localization. Experimental Cell Research, 313, 3680–3693.

    Article  PubMed  CAS  Google Scholar 

  23. Fan, Z., Beresford, P. J., Oh, D. Y., Zhang, D., & Lieberman, J. (2003). Tumor suppressor NM23–H1 is a granzyme A-activated DNase during CTL-mediated apoptosis and the nucleosome assembly protein SET is its inhibitor. Cell, 112, 659–672.

    Article  PubMed  CAS  Google Scholar 

  24. Wong, H. K., Muftuoglu, M., Beck, G., Imam, S. Z., Bohr, V. A., & Wilson, D. M., 3rd. (2007). Cockayne syndrome B protein stimulates apurinic endonuclease 1 activity and protects against agents that introduce base excision repair intermediates. Nucleic Acids Research, 35, 4103–4113.

    Article  PubMed  CAS  Google Scholar 

  25. Belzile, J. P., Choudhury, S. A., Cournoyer, D., & Chow, T. Y. (2006). Targeting DNA repair proteins: A promising avenue for cancer gene therapy. Current Gene Therapy, 6, 111–123.

    Article  PubMed  CAS  Google Scholar 

  26. Ding, J., Miao, Z. H., Meng, L. H., & Geng, M. Y. (2006). Emerging cancer therapeutic opportunities target DNA-repair systems. Trends in Pharmacological Sciences, 27, 338–344.

    Article  PubMed  CAS  Google Scholar 

  27. Madhusudan, S., & Middleton, M. R. (2005). The emerging role of DNA repair proteins as predictive, prognostic and therapeutic targets in cancer. Cancer Treatment Review, 31, 603–617.

    Article  CAS  Google Scholar 

  28. Kaetzel, D. M., Zhang, Q., Yang, M., McCorkle, J. R., Ma, D., & Craven, R. J. (2006). Potential roles of 3′-5′ exonuclease activity of NM23–H1 in DNA repair and malignant progression. Journal of Bioenergetics and Biomembranes, 38, 163–167.

    Article  PubMed  CAS  Google Scholar 

  29. Lu, Q., Zhang, X., Almaula, N., Mathews, C. K., & Inouye, M. (1995). The gene for nucleoside diphosphate kinase functions as a mutator gene in Escherichia coli. Journal of Molecular Biology, 254, 337–341.

    Article  PubMed  CAS  Google Scholar 

  30. Fabbro, M., & Henderson, B. R. (2003). Regulation of tumor suppressors by nuclear-cytoplasmic shuttling. Experimental Cell Research, 282, 59–69.

    Article  PubMed  CAS  Google Scholar 

  31. Henderson, B. R. (2005). Regulation of BRCA1, BRCA2 and BARD1 intracellular trafficking. BioEssays, 27, 884–893.

    Article  PubMed  CAS  Google Scholar 

  32. Simeonov, A., Kulkarni, A., Dorjsuren, D., et al. (2009). Identification and characterization of inhibitors of human apurinic/apyrimidinic endonuclease APE1. PLoS One, 4, e5740.

    Article  PubMed  Google Scholar 

  33. Ma, D., McCorkle, J. R., & Kaetzel, D. M. (2004). The metastasis suppressor NM23–H1 possesses 3′-5′ exonuclease activity. The Journal of Biological Chemistry, 279, 18073–18084.

    Article  PubMed  CAS  Google Scholar 

  34. Postel, E. H. (2003). Multiple biochemical activities of Nm23/NDP kinase in gene regulation. Journal of Bioenergetics and Biomembranes, 35, 31–40.

    Article  PubMed  CAS  Google Scholar 

  35. Zou, Y., Crowley, D. J., & Van Houten, B. (1998). Involvement of molecular chaperonins in nucleotide excision repair: Dna K leads to increased thermal stability of UvrA, catalytic UvrB loading, enhanced repair and increased UV resistance. The Journal of Biological Chemistry, 273, 12887–12892.

    Article  PubMed  CAS  Google Scholar 

  36. Chow, K. C., & Tung, W. L. (2000). Magnetic field exposure enhances DNA repair through the induction of DnaK/J synthesis. FEBS Letters, 478, 133–136.

    Article  PubMed  CAS  Google Scholar 

  37. Wilson, D. M., 3rd, Takeshita, M., & Demple, B. (1997). Abasic site binding by the human apurinic endonuclease, Ape, and determination of the DNA contact sites. Nucleic Acids Research, 25, 933–939.

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank the Chinese National Natural Science Foundation (grants #30801367, #30973457).

Author information

Authors and Affiliations

  1. Cancer Center, Institute of Surgical Research, Daping Hospital, Third Military Medical University, Chongqing, 400042, China

    Zhi-Min Zhang, Xue-Qin Yang, Dong Wang, Ge Wang, Zhen-Zhou Yang, Yi Qing, Zhi-Xiang Yang, Meng-Xia Li & De-Bing Xiang

Authors
  1. Zhi-Min Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xue-Qin Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Dong Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ge Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Zhen-Zhou Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yi Qing
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Zhi-Xiang Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Meng-Xia Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. De-Bing Xiang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Xue-Qin Yang or Dong Wang.

Additional information

Zhi-Min Zhang and Xue-Qin Yang contributed equally to this work as co-first authors.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zhang, ZM., Yang, XQ., Wang, D. et al. Nm23-H1 Protein Binds to APE1 at AP Sites and Stimulates AP Endonuclease Activity Following Ionizing Radiation of the Human Lung Cancer A549 Cells. Cell Biochem Biophys 61, 561–572 (2011). https://doi.org/10.1007/s12013-011-9238-9

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12013-011-9238-9

Keywords

Search

Navigation