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Ren X.-L.,Nanjing Agricultural University | Chen R.-R.,Nanjing Agricultural University | Zhang Y.,Nanjing Agricultural University | Ma Y.,Institute of Cotton Research of CAAS State Key Laboratory of Cotton Biology | And 4 more authors.
Applied and Environmental Microbiology | Year: 2013

Crystal toxin Cry1Ca from Bacillus thuringiensis has an insecticidal spectrum encompassing lepidopteran insects that are tolerant to current commercially used B. thuringiensis crops (Bt crops) expressing Cry1A toxins and may be useful as a potential bioinsecticide. The mode of action of Cry1A is fairly well understood. However, whether Cry1Ca interacts with the same receptor proteins as Cry1A remains unproven. In the present paper, we first cloned a cadherin-like gene, SeCad1b, from Spodoptera exigua (relatively susceptible to Cry1Ca). SeCad1b was highly expressed in the larval gut but scarcely detected in fat body, Malpighian tubules, and remaining carcass. Second, we bacterially expressed truncated cadherin rSeCad1bp and its interspecific homologue rHaBtRp from Helicoverpa armigera (more sensitive to Cry1Ac) containing the putative toxin-binding regions. Competitive binding assays showed that both Cry1Ca and Cry1Ac could bind to rSeCad1bp and rHaBtRp, and they did not compete with each other. Third, Cry1Ca ingestion killed larvae and decreased the weight of surviving larvae. Dietary introduction of SeCad1b double-stranded RNA (dsRNA) reduced approximately 80% of the target mRNA and partially alleviated the negative effect of Cry1Ca on larval survival and growth. Lastly, rSeCad1bp and rHaBtRp differentially enhanced the negative effects of Cry1Ca and Cry1Ac on the larval mortalities and growth of S. exigua and H. armigera. Thus, we provide the first lines of evidence to suggest that SeCad1b from S. exigua is a functional receptor of Cry1Ca. ©2013, American Society for Microbiology. Source

Chen R.-R.,Nanjing Agricultural University | Ren X.-L.,Nanjing Agricultural University | Han Z.-J.,Nanjing Agricultural University | Mu L.-L.,Nanjing Agricultural University | And 3 more authors.
Archives of Insect Biochemistry and Physiology | Year: 2014

In S. exigua, ingestion of Cry1Ac reduces larval growth, shortens lifespan, and decreases copulation and oviposition of the adults. Cadherin-like protein SeCad1b in S. exigua has recently been published. Here, we tested whether SeCad1b mediates the negative effects of Cry1Ac. We identified three potential Cry toxin binding regions in SeCad1b, i.e., 879EIAIQITDTNN889, 1357SLLTVTI1363, and 1436GVISLNFQ1443. We expressed and purified a truncated cadherin, rSeCad1bp, and its interspecific homologue, rHaBtRp, from H. armigera that contain the putative toxin binding regions. Using a toxin overlay assay, we found that rSeCad1bp specifically binds to biotinylated Cry1Ac in a dose-dependent manner. We also discovered that an addition of rSeCad1bp and rHaBtRp enhances the suppression of larval growth by Cry1Ac, although rSeCad1bp is less suppressive than rHaBtRp. Finally, RNA interference-mediated knockdown of SeCad1b reduced approximately 80% of the target gene and significantly alleviated the negative effect of CrylAc on larval growth. We infer that the S. exigua SeCad1b is a functional receptor of Cry1Ac. © 2014 Wiley Periodicals, Inc. Source

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