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Asparagine synthetase is partially localized to the plasma membrane and upregulated by L-asparaginase in U937 cells

Journal of Huazhong University of Science and Technology [Medical Sciences] volume 31, pages 159–163 (2011)Cite this article

Summary

This study investigated the intracellular localization of asparagine synthetase (ASNS) in the relation with chemoresistance in leukemia. pIRES-GFP-ASNS-Flag/Neo expression vector was transiently tansfected into SK-N-MC cells and 297T cells respectively. Immunofluorescence and Western blot analysis were performed for cellular localization of ASNS respectively. U937 cells were treated with L-asparaginase for 48 h and examined for endogenous ASNS expression on plasma membrane by immunofluorescence staining. Immunofluorescence staining showed that the transiently expressed ASNS was partly localized on transfected-SK-N-MC cell surface. Moreover, Western blotting exhibited that ASNS expressed both in cytosol and on plasma membrane of transfected-293T cells. Immunofluorescence staining with anti-ASNS-specific monoclonal antibody revealed that endogenous ASNS was localized on the plasma membrane of U937 cells, except for its distribution in the cytosol. In addition, ASNS exhibited a higher expression on plasma membrane after treatment with L-asparaginase as compared with the untreated cells. It was concluded that the subcellular translocation of ASNS may play an important role in L-asparaginase resistance in leukemia cells.

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References

  1. Pui CH, Evans WE. Treatment of acute lymphoblastic leukemia. N Engl J Med, 2006,354(2):166–178

    PubMed  Article  CAS  Google Scholar 

  2. Zwaan CM, Kaspers GJ, Pieters R, et al. Cellular drug resistance profiles in childhood acute myeloid leukemia: differences between FAB types and comparison with acute lymphoblastic leukemia. Blood, 2000,96(8):2879–2886.

    PubMed  CAS  Google Scholar 

  3. Kaspers GJ, Veerman AJ, Pieters R, et al. In vitro cellular drug resistance and prognosis in newly diagnosed childhood acute lymphoblastic leukemia. Blood, 1997,90(7): 2723–2729

    PubMed  CAS  Google Scholar 

  4. Yamada S, Hongo T, Okada S, et al. Clinical relevance of in vitro chemoresistance in childhood acute myeloid leukemia. Leukemia, 2001,15(12):1892–1897

    PubMed  CAS  Google Scholar 

  5. Okada S, Hongo T, Yamada S, et al. In vitro efficacy of l-asparaginase in childhood acute myeloid leukaemia. Br J Haematol, 2003,123(5):802–809

    PubMed  Article  CAS  Google Scholar 

  6. Dubbers A, Wurthwein G, Muller HJ, et al. Asparagine synthetase activity in paediatric acute leukaemias: AML-M5 subtype shows lowest activity. Br J Haematol, 2000,109(2):427–429

    PubMed  Article  CAS  Google Scholar 

  7. Muller HJ, Boos J. Use of L-asparaginase in childhood ALL. Crit Rev Oncol Hematol, 1998,28(2):97–113

    PubMed  Article  CAS  Google Scholar 

  8. Miller HK, Salser JS, Balis ME. Amino acid levels following L-asparagine amidohydrolase (EC.3.5.1.1) therapy. Cancer Res, 1969,29(1):183–187

    PubMed  CAS  Google Scholar 

  9. Ohnuma T, Holland JF, Freeman A, et al. Biochemical and pharmacological studies wih asparaginase in man. Cancer Res, 1970,30(9):2297–2305

    PubMed  CAS  Google Scholar 

  10. Richards NG, Schuster SM. Mechanistic issues in asparagine synthetase catalysis. Adv Enzymol Relat Areas Mol Biol, 1998,72N:145–198

    Google Scholar 

  11. Li BS, Gu LJ, Luo CY, et al. The downregulation of asparagine synthetase expression can increase the sensitivity of cells resistant to L-asparaginase. Leukemia, 2006, 20(12):2199–2201

    PubMed  Article  CAS  Google Scholar 

  12. Appel IM, den Boer ML, Meijerink JP, et al. Up-regulation of asparagine synthetase expression is not linked to the clinical response L-asparaginase in pediatric acute lymphoblastic leukemia. Blood, 2006,107(11): 4244–4249

    PubMed  Article  CAS  Google Scholar 

  13. Holleman A, den Boer ML, Kazemier KM, et al. Resistance to different classes of drugs is associated with impaired apoptosis in childhood acute lymphoblastic leukemia. Blood, 2003,102(13):4541–4546

    PubMed  Article  CAS  Google Scholar 

  14. Leslie M, Case MC, Hall AG, et al. Expression levels of asparagine synthetase in blasts from children and adults with acute lymphoblastic leukaemia. Br J Haematol, 2006,132(6):740–742

    PubMed  Article  CAS  Google Scholar 

  15. Ando M, Sugimoto K, Kitoh T, et al. Selective apoptosis of natural killer-cell tumours by L-asparaginase, Br J Haematol, 2005,130(6):860–868

    PubMed  Article  CAS  Google Scholar 

  16. Aslanian AM, Kilberg MS. Multiple adaptive mechanisms affect asparagine synthetase substrate availability in asparaginase-resistant MOLT-4 human leukaemia cells. Biochem J, 2001,358(Pt 1):59–67

    PubMed  Article  CAS  Google Scholar 

  17. Haskell CM, Canellos GP. L-asparaginase resistance in human leukemia—asparagine synthetase. Biochem Pharmacol, 1969,18(10):2578–2580

    PubMed  Article  CAS  Google Scholar 

  18. Kiriyama Y, Kubota M, Takimoto T, et al. Biochemical characterization of U937 cells resistant to L-asparaginase: the role of asparagine synthetase. Leukemia, 1989,3(4): 294–297

    PubMed  CAS  Google Scholar 

  19. Codegoni AM, Biondi A, Conter V, et al. Human monocytic leukemia expresses low levels of asparagine synthase and is potentially sensitive to L-asparaginase. Leukemia, 1995,9(2):360–361

    PubMed  CAS  Google Scholar 

  20. Barbosa-Tessmann IP, Chen C, Zhong C, et al. Activation of the human asparagine synthetase gene by the amino acid response and the endoplasmic reticulum stress response pathways occurs by common genomic elements. J Biol Chem, 2000,275(35):26976–26985

    PubMed  CAS  Google Scholar 

  21. Jousse C, Averous J, Bruhat A, et al. Amino acids as regulators of gene expression: molecular mechanisms. Biochem Biophys Res Commun, 2004,313(2):447–452

    PubMed  Article  CAS  Google Scholar 

  22. Fafournoux P, Bruhat A, Jousse C. Amino acid regulation of gene expression. Biochem J, 2000,351(Pt 1N):1–12

    PubMed  Article  CAS  Google Scholar 

  23. Hutson RG, Kitoh T, Moraga Amador DA, et al. Amino acid control of asparagine synthetase: relation to asparaginase resistance in human leukemia cells. Am J Physiol, 1997,272(5 Pt 1):C1691–C1699

    PubMed  CAS  Google Scholar 

  24. Guerrini L, Gong SS, Mangasarian K, et al. Cis- and trans-acting elements involved in amino acid regulation of asparagine synthetase gene expression. Mol Cell Biol, 1993,13(6):3202–3212

    PubMed  CAS  Google Scholar 

  25. Greco A, Gong SS, Ittmann M, et al. Organization and expression of the cell cycle gene, ts11, that encodes asparagine synthetase. Mol Cell Biol, 1989,9 (6):2350–2359

    PubMed  CAS  Google Scholar 

  26. Hongo S, Sakagami H, Sato T. Decrease in asparagine synthetase activity during cell differentiation of mouse and human leukemia cell lines. Leukemia, 1990,4(10): 708–711

    PubMed  CAS  Google Scholar 

  27. Richards NG, Kilberg MS. Asparagine synthetase chemotherapy. Annu Rev Biochem, 2006,75N:629–654

    Article  Google Scholar 

  28. Fine BM, Kaspers GJ, Ho M, et al. A genome-wide view of the in vitro response to l-asparaginase in acute lymphoblastic leukemia. Cancer Res. 2005,65(1):291–299

    PubMed  CAS  Google Scholar 

  29. Palacin M, Estevez R, Bertran J, et al. Molecular biology of mammalian plasma membrane amino acid transporters. Physiol Rev, 1998,78(4):969–1054

    PubMed  CAS  Google Scholar 

  30. Iwamoto S, Mihara K, Downing JR, et al. Mesenchymal cells regulate the response of acute lymphoblastic leukemia cells to asparaginase. J Clin Invest, 2007,117(4): 1049–1057

    PubMed  Article  CAS  Google Scholar 

  31. Rubartelli A, Sitia R. Interleukin 1 beta and thioredoxin are secreted through a novel pathway of secretion. Biochem Soc Trans, 1991,19(2):255–259

    PubMed  CAS  Google Scholar 

  32. Rubartelli A, Bajetto A, Allavena G, et al. Secretion of thioredoxin by normal and neoplastic cells through a leaderless secretory pathway. J Biol Chem, 1992,267(34): 24161–24164

    PubMed  CAS  Google Scholar 

  33. Mignatti P, Morimoto T, Rifkin DB. Basic fibroblast growth factor, a protein devoid of secretory signal sequence, is released by cells via a pathway independent of the endoplasmic reticulum-Golgi complex. J Cell Physiol, 1992,151(1):81–93

    PubMed  Article  CAS  Google Scholar 

  34. Mehul B, Hughes RC. Plasma membrane targetting, vesicular budding and release of galectin 3 from the cytoplasm of mammalian cells during secretion. J Cell Sci, 1997,110( Pt10):1169–1178

    PubMed  CAS  Google Scholar 

  35. Chang HC, Samaniego F, Nair BC, et al. HIV-1 Tat protein exits from cells via a leaderless secretory pathway and binds to extracellular matrix-associated heparan sulfate proteoglycans through its basic region. AIDS, 1997,11(12): 1421–1431

    PubMed  Article  CAS  Google Scholar 

  36. Tanudji M, Hevi S, Chuck SL. Improperly folded green fluorescent protein is secreted via a non-classical pathway. J Cell Sci, 2002,115(Pt 19):3849–3857

    PubMed  Article  CAS  Google Scholar 

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Affiliations

  1. Department of Hematology/Oncology, Shanghai Children’s Medical Center, Shanghai Jiaotong University School of Medicine, Shanghai, 200127, China

    Yingyi He, Benshang Li, Changying Luo, Shuhong Shen, Jing Chen, Huiliang Xue, Jingyan Tang & Longjun Gu

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  1. Yingyi He
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  2. Benshang Li
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  3. Changying Luo
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  4. Shuhong Shen
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  5. Jing Chen
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  6. Huiliang Xue
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  7. Jingyan Tang
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  8. Longjun Gu
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Corresponding author

Correspondence to Longjun Gu.

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The two authors contributed equally to this work.

This project was supported by grants from the National Natural Science Foundation of China (Nos. 30672271 and 81000227).

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He, Y., Li, B., Luo, C. et al. Asparagine synthetase is partially localized to the plasma membrane and upregulated by L-asparaginase in U937 cells. J. Huazhong Univ. Sci. Technol. [Med. Sci.] 31, 159–163 (2011). https://doi.org/10.1007/s11596-011-0243-4

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  • DOI: https://doi.org/10.1007/s11596-011-0243-4

Key words

  • asparagine synthetase
  • L-asparaginase
  • acute myeloid leukemia
  • drug resistance
  • plasma membrane