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Expression of recombinant and mosaic Cry1Ac receptors from Helicoverpa armigera and their influences on the cytotoxicity of activated Cry1Ac to Spodoptera litura Sl-HP cells

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Abstract

Bacillus thuringiensis (Bt) toxin receptors play important roles in the killing of pests, and investigation on characterization of the receptors is essential for utilization of Bt and management of insect resistance. Here, recombinant and mosaic receptors of Bt Cry1Ac toxin from Helicoverpa armigera were expressed in Spodoptera litura Sl-HP cells and their influences on cytotoxicity of activated Cry1Ac toxin were investigated. When H. armigera aminopeptidase N1 (APN1), alkaline phosphatase 2 (ALP2) and cadherin fused with or without GFP tag were, respectively, expressed in Sl-HP cells, live cell-immunofluorescence staining detection revealed that the quantity of the toxin binding to cadherin or cadherin-GFP was much more than that binding to ALP2 and APN1 or their fusion proteins with GFP, and only the cadherin- or cadherin-GFP-expressing cells showed aberrant cell morphology after the treatment of the toxin at low concentrations. ALP2 and APN1 fused with or without GFP tag did not significantly enhance the cadherin-mediated cytotoxicity of the toxin. The mosaic ALP-TBR-GFP-GPI was located on cell membrane, but did not bind to the toxin. The mosaic truncated cadherin-GFP-GPI was not located on cell membrane even if the signal peptide was sustained. The concentrations of the toxin resulting in swelling of 50 % cells for noncadherin-expressing Sl-HP cells and cadherin-expressing Hi5 cells were 5.08 and 9.50 µg/ml within 1 h, respectively. Taken together, our data have indicated that the binding affinity of ALP2 and APN1 to activated Cry1Ac toxin is much weaker than that of cadherin and both ALP2 and APN1 do not enhance the cytotoxicity of the toxin even though cadherin is co-expressed, and the mosaic receptor of ALP2 inserted with cadherin toxin binding domain does not mediate cytotoxicity of the toxin. In addition, the noncadherin-expressing Sl-HP cells are more susceptible to activated Cry1Ac than the cadherin-expressing Hi5 cells.

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Acknowledgments

The work was supported by the National Natural Science Foundation of China (Grant Numbers: 31071739 and 31372260).

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Authors and Affiliations

  1. School of Life Sciences, Central China Normal University, Wuhan, 430079, People’s Republic of China

    Peng Xu, Mayira Islam, Fei He, Jianxin Peng, Huazhu Hong & Kaiyu Liu

  2. State Key Laboratory of Plant Disease and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing, 100193, People’s Republic of China

    Yutao Xiao & Chenxi Liu

  3. Department of Biological Sciences and Biotechnology, Wuhan Bioengineering Institute, Wuhan, 430415, People’s Republic of China

    Yi Li

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  1. Peng Xu
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  2. Mayira Islam
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  3. Yutao Xiao
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  4. Fei He
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  6. Jianxin Peng
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Correspondence to Chenxi Liu or Kaiyu Liu.

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Peng Xu and Mayira Islam have contributed equally to this work.

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Xu, P., Islam, M., Xiao, Y. et al. Expression of recombinant and mosaic Cry1Ac receptors from Helicoverpa armigera and their influences on the cytotoxicity of activated Cry1Ac to Spodoptera litura Sl-HP cells. Cytotechnology 68, 481–496 (2016). https://doi.org/10.1007/s10616-014-9801-5

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  • DOI: https://doi.org/10.1007/s10616-014-9801-5

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