The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon

doi:10.1016/S2095-3119(13)60427-X

Journal of Integrative Agriculture ›› 2013, Vol. 12 ›› Issue (9): 1598-1605.DOI: 10.1016/S2095-3119(13)60427-X

The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon

LU Qiong, CAO Guang-chun, ZHANG Li-li, LIANG Ge-mei, GAO Xi-wu, ZHANG Yong-jun , GUO Yu-yuan

1.State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, P.R.China
2.College of Agriculture and Biotechnology, China Agricultural University, Beijing 100193, P.R.China

收稿日期:2012-11-14 出版日期:2013-09-01 发布日期:2013-09-15
通讯作者: Correspondence ZHANG Yong-jun, Tel: +86-10-62815929, E-mail: yjzhang@ippcaas.cn; GUO Yu-yuan, Tel: +86-10-62815929, Fax: +86-10-62894786, E-mail: yuyuanguo@hotmail.com
基金资助:
This work was supported by the National 973 Program of China (2012CB114104), the National Natural Science Foundation of China (31071694, 31171858), and the International Joint Project between China and UK of National Natural Science Foundation of China and Royal Society (31111130203).

The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon

LU Qiong, CAO Guang-chun, ZHANG Li-li, LIANG Ge-mei, GAO Xi-wu, ZHANG Yong-jun , GUO Yu-yuan

1.State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, P.R.China
2.College of Agriculture and Biotechnology, China Agricultural University, Beijing 100193, P.R.China

Received:2012-11-14 Online:2013-09-01 Published:2013-09-15
Contact: Correspondence ZHANG Yong-jun, Tel: +86-10-62815929, E-mail: yjzhang@ippcaas.cn; GUO Yu-yuan, Tel: +86-10-62815929, Fax: +86-10-62894786, E-mail: yuyuanguo@hotmail.com
Supported by:
This work was supported by the National 973 Program of China (2012CB114104), the National Natural Science Foundation of China (31071694, 31171858), and the International Joint Project between China and UK of National Natural Science Foundation of China and Royal Society (31111130203).

摘要/Abstract

摘要： Cry toxins produced by Bacillus thuringiensis (Bt) are effective biological insecticides against certain insect species. However, there are potential risks of the evolved resistance of insects to Cry toxin owing to decreased binding of toxins to target sites in the brush border membranes of the larva midgut. The Cry toxins with different binding sites in the larval midgut have been considered to be a good combination to deploy in delaying resistance evolution. Bioassay results demonstrated that the toxicity of different Cry toxins ranked differently for each species. The toxicity ranking was Cry1Ac>Cry1Ab>Cry2Ab for Helicoverpa armigera, Cry1B>Cry1C>Cry2Ab for Spodoptera exigua, and Cry2Ab>Cry1B> Cry1C for S. litura. Only Cry2Ab was toxic to Agrotis ipsilon. Binding experiments were performed with 125I-Cry1Ab, 125ICry1Ac, 125I-Cry1B, 125I-Cry1C, 125I-Cry2Ab and the brush border membranes vesicles (BBMV) from H. armigera, S. exigua, S. litura and A. ipsilon. The binding of Cry1Ab and Cry1Ac was shown to be saturable by incubating with increasing concentrations of H. armigera BBMV (Kd=(45.00±2.01) nmol L-1 and (12.80±0.18) nmol L-1, respectively; Bmax=(54.95±1.79) ng and (55.44±0.91) ng, separately). The binding of Cry1B was shown to be saturable by incubating with increasing concentrations of S. exigua BBMV (Kd=(23.26±1.66) nmol L-1; Bmax=(65.37±1.87) ng). The binding of 125ICry toxins was shown to be non-saturable by incubating with increasing concentrations of S. litura and A. ipsilon BBMV. In contrast, Cry1B and Cry1C showed some combination with the BBMV of S. litura, and a certain amount of Cry2Ab could bind to the BBMV of A. ipsilon. These observations suggest that a future strategy could be devised for the focused combination of specific cry genes in transgenic crops to control target pests, widen the spectrum of insecticide effectiveness and postpone insect resistance evolution.

关键词: Cry toxin , toxicity , binding assays , 125I labeling , BBMV

Abstract: Cry toxins produced by Bacillus thuringiensis (Bt) are effective biological insecticides against certain insect species. However, there are potential risks of the evolved resistance of insects to Cry toxin owing to decreased binding of toxins to target sites in the brush border membranes of the larva midgut. The Cry toxins with different binding sites in the larval midgut have been considered to be a good combination to deploy in delaying resistance evolution. Bioassay results demonstrated that the toxicity of different Cry toxins ranked differently for each species. The toxicity ranking was Cry1Ac>Cry1Ab>Cry2Ab for Helicoverpa armigera, Cry1B>Cry1C>Cry2Ab for Spodoptera exigua, and Cry2Ab>Cry1B> Cry1C for S. litura. Only Cry2Ab was toxic to Agrotis ipsilon. Binding experiments were performed with 125I-Cry1Ab, 125ICry1Ac, 125I-Cry1B, 125I-Cry1C, 125I-Cry2Ab and the brush border membranes vesicles (BBMV) from H. armigera, S. exigua, S. litura and A. ipsilon. The binding of Cry1Ab and Cry1Ac was shown to be saturable by incubating with increasing concentrations of H. armigera BBMV (Kd=(45.00±2.01) nmol L-1 and (12.80±0.18) nmol L-1, respectively; Bmax=(54.95±1.79) ng and (55.44±0.91) ng, separately). The binding of Cry1B was shown to be saturable by incubating with increasing concentrations of S. exigua BBMV (Kd=(23.26±1.66) nmol L-1; Bmax=(65.37±1.87) ng). The binding of 125ICry toxins was shown to be non-saturable by incubating with increasing concentrations of S. litura and A. ipsilon BBMV. In contrast, Cry1B and Cry1C showed some combination with the BBMV of S. litura, and a certain amount of Cry2Ab could bind to the BBMV of A. ipsilon. These observations suggest that a future strategy could be devised for the focused combination of specific cry genes in transgenic crops to control target pests, widen the spectrum of insecticide effectiveness and postpone insect resistance evolution.

Key words: Cry toxin , toxicity , binding assays , 125I labeling , BBMV

LU Qiong, CAO Guang-chun, ZHANG Li-li, LIANG Ge-mei, GAO Xi-wu, ZHANG Yong-jun , GUO Yu-yuan. The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon[J]. Journal of Integrative Agriculture, 2013, 12(9): 1598-1605.

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The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon

The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon

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