ATP-Binding Cassette Subfamily a Member 2 Is a Functional Receptor for Bacillus thuringiensis Cry2A Toxins in Bombyx mori, But Not for Cry1A, Cry1C, Cry1D, Cry1F, or Cry9A Toxins
ATP-Binding Cassette Subfamily a Member 2 Is a Functional Receptor for Bacillus thuringiensis Cry2A Toxins in Bombyx mori, But Not for Cry1A, Cry1C, Cry1D, Cry1F, or Cry9A Toxins
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DOI：
10.3390/toxins12020104
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发表时间：
2020-02-01
期刊：
影响因子：
4.2
通讯作者：
Watanabe, Kenji
中科院分区：
文献类型：
--
作者：
Li, Xiaoyi;Miyamoto, Kazuhisa;Watanabe, Kenji
苏云金芽孢杆菌毒素是苏云金芽孢杆菌(Bt)产生的杀虫蛋白。它们在商业上被用来控制害虫，因为它们在特定的昆虫中非常活跃，对环境和人类健康无害。通过对Cry2A毒素抗性基因的分析，确定了编码ATP结合盒A亚家族成员2(ABCA2)的基因。然而，我们没有直接证据证明ABCA2在Cry2A毒素中的作用，也没有直接证据证明为什么Cry2A毒素的抗性不会与其他Cry毒素交叉。因此，我们进行了两个实验。首先，我们使用转录激活子样效应子-核酸酶(TALENS)编辑了家蚕ABCA2序列，并在饲料覆盖生物测定中证实了该序列的敏感性决定能力。C端缺失BmABCA2的菌株对Cry2A毒素表现出较强的特异性抗性，即使缺失1~3个氨基酸的菌株也表现出抗性。然而，C端半缺失的菌株对其他毒素没有交叉耐药性。其次，我们在HEK239T细胞中进行了细胞肿胀实验，证实了BmABCA2对Cry2A毒素的特异性。这些结果表明，BmABCA2是Cry2A毒素的功能性受体，依赖于BmABCA2缺乏的Cry2A抗性不会对Cry1A、Cry1Ca、Cry1Da、Cry1Fa或Cry9Aa毒素产生交叉抗性。
Cry toxins are insecticidal proteins produced by Bacillus thuringiensis (Bt). They are used commercially to control insect pests since they are very active in specific insects and are harmless to the environment and human health. The gene encoding ATP-binding cassette subfamily A member 2 (ABCA2) was identified in an analysis of Cry2A toxin resistance genes. However, we do not have direct evidence for the role of ABCA2 for Cry2A toxins or why Cry2A toxin resistance does not cross to other Cry toxins. Therefore, we performed two experiments. First, we edited the ABCA2 sequence in Bombyx mori using transcription activator-like effector-nucleases (TALENs) and confirmed the susceptibility-determining ability in a diet overlay bioassay. Strains with C-terminal half-deleted BmABCA2 showed strong and specific resistance to Cry2A toxins; even strains carrying a deletion of 1 to 3 amino acids showed resistance. However, the C-terminal half-deleted strains did not show cross-resistance to other toxins. Second, we conducted a cell swelling assay and confirmed the specific ability of BmABCA2 to Cry2A toxins in HEK239T cells. Those demonstrated that BmABCA2 is a functional receptor for Cry2A toxins and that BmABCA2 deficiency-dependent Cry2A resistance does not confer cross-resistance to Cry1A, Cry1Ca, Cry1Da, Cry1Fa or Cry9Aa toxins.