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Breast Cancer Research and Treatment

, Volume 116, Issue 1, pp 91–102 | Cite as

Expression of TNF-α leader sequence renders MCF-7 tumor cells resistant to the cytotoxicity of soluble TNF-α

  • Dan Yan
  • Nalin Qin
  • Hailong Zhang
  • Tao Liu
  • Mingxia Yu
  • Xiaodan Jiang
  • Wei Feng
  • Jing Wang
  • Bingjiao Yin
  • Tao Zhang
  • Muxiang ZhouEmail author
  • Zhuoya LiEmail author
Preclinical Study

Abstract

Transmembrane TNF-α (tmTNF-α) contains a leader sequence (LS) that can be phosphorylated and cleaved at its cytoplasmic portion, inducing IL-12 production. We observed that the breast cancer cell line MDA-MB-231 expressing transmembrane TNF-α (tmTNF-α) at high level was resistant to soluble TNF-α (sTNF-α)-induced cytotoxicity, accompanied by constitutive NF-κB activation. In contrast, MCF-7 cells expressing tmTNF-α at very low level were sensitive to sTNF-α-induced cell death and had no detectable NF-κB activation. Consistently, siRNA-mediated tmTNF-α knockdown blocked NF-κB activation and rendered MDA-MB-231 sensitive. To test our hypothesis that TNF-LS may play an important role in determining the sensitivity of tumor cells to sTNF-α, we stably transfected MCF-7 cells with TNF-LS. We found that transfection of TNF-LS or wild-type TNF-α containing LS constitutively activated NF-κB and conferred the cytotoxic resistance of MCF-7 cells, while transfection of a mutant tmTNF-α lacking the cytoplasmic segment of LS neither activated NF-κB nor affected the sensitivity. However, NF-κB inhibitor PDTC suppressed NF-κB activation and reconstituted sensitivity of TNF-LS/MCF-7 cells. To check whether TNF-LS is required to be cleaved or internalized for NF-κB activation to occur, we used signal peptide peptidase inhibitor (Z-LL)2-ketone and receptor internalization inhibitor MDC to treat cells. Interestingly, both inhibitors increased TNF-LS expression on the cell surface and enhanced NF-κB activation. These results indicate that membrane-anchored TNF-LS contributes to constitutive activation of NF-κB and resistance to sTNF-α-induced cell death. Therefore, TNF-LS appears to be responsible for tmTNF-α-induced resistance in the breast cancer cells.

Keywords

Transmembrane TNF-α TNF-leader sequence Cytotoxicity NF-κB Breast cancer 

Notes

Acknowledgements

We thank Dr. Malcolm Kennedy for editing the manuscript. This work was supported by the National Natural Science Foundation of China (30670421), National Key Basic Research Program of China from the Ministry of Science and Technology of the People’s Republic of China (2004AA215162).

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Copyright information

© Springer Science+Business Media, LLC. 2008

Authors and Affiliations

  • Dan Yan
    • 1
  • Nalin Qin
    • 1
  • Hailong Zhang
    • 1
  • Tao Liu
    • 1
  • Mingxia Yu
    • 1
  • Xiaodan Jiang
    • 1
  • Wei Feng
    • 1
  • Jing Wang
    • 1
  • Bingjiao Yin
    • 1
  • Tao Zhang
    • 1
  • Muxiang Zhou
    • 2
    Email author
  • Zhuoya Li
    • 1
    Email author
  1. 1.Department of Immunology, Tongji Medical CollegeHuazhong University of Science and TechnologyWuhanPeople’s Republic of China
  2. 2.Division of Pediatric Hematology/Oncology/BMTEmory University School of MedicineAtlantaUSA

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