A versatile contribution of both aminopeptidases N and ABC transporters to Bt Cry1Ac toxicity in the diamondback moth

BMC Biology

Mario Soberón¹  Alejandra Bravo¹  Xuguo Zhou²  Liuhong Zhu³  Zhaojiang Guo³  Youjun Zhang³  Le Guo³  Shaoli Wang³  Qingjun Wu³  Dan Sun⁴  Neil Crickmore⁵ 

[1] Departamento de Microbiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Apdo. Postal 510-3, 62250, Cuernavaca, Morelos, México;Department of Entomology, University of Kentucky, 40546-0091, Lexington, KY, USA;Department of Plant Protection, Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, 100081, Beijing, China;Department of Plant Protection, Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, 100081, Beijing, China;Guangdong Laboratory for Lingnan Modern Agriculture, 510642, Guangdong, China;School of Life Sciences, University of Sussex, BN1 9QE, Brighton, UK;

关键词: Bacillus thuringiensis;    Plutella xylostella;    CRISPR/Cas9;    ABC transporters;    GPI-anchored proteins;    Receptor redundancy;    Toxin action;   

DOI  :  10.1186/s12915-022-01226-1

来源: Springer

【 摘 要 】

BackgroundBiopesticides and transgenic crops based on Bacillus thuringiensis (Bt) toxins are extensively used to control insect pests, but the rapid evolution of insect resistance seriously threatens their effectiveness. Bt resistance is often polygenic and complex. Mutations that confer resistance occur in midgut proteins that act as cell surface receptors for the toxin, and it is thought they facilitate its assembly as a membrane-damaging pore. However, the mechanistic details of the action of Bt toxins remain controversial.ResultsWe have examined the contribution of two paralogous ABC transporters and two aminopeptidases N to Bt Cry1Ac toxicity in the diamondback moth, Plutella xylostella, using CRISPR/Cas9 to generate a series of homozygous polygenic knockout strains. A double-gene knockout strain, in which the two paralogous ABC transporters ABCC2 and ABCC3 were deleted, exhibited 4482-fold resistance to Cry1A toxin, significantly greater than that previously reported for single-gene knockouts and confirming the mutual functional redundancy of these ABC transporters in acting as toxin receptors in P. xylostella. A double-gene knockout strain in which APN1 and APN3a were deleted exhibited 1425-fold resistance to Cry1Ac toxin, providing the most direct evidence to date for these APN proteins acting as Cry1Ac toxin receptors, while also indicating their functional redundancy. Genetic crosses of the two double-gene knockouts yielded a hybrid strain in which all four receptor genes were deleted and this resulted in a > 34,000-fold resistance, indicating that while both types of receptor need to be present for the toxin to be fully effective, there is a level of functional redundancy between them. The highly resistant quadruple knockout strain was less fit than wild-type moths, but no fitness cost was detected in the double knockout strains.ConclusionOur results provide direct evidence that APN1 and APN3a are important for Cry1Ac toxicity. They support our overarching hypothesis of a versatile mode of action of Bt toxins, which can compensate for the absence of individual receptors, and are consistent with an interplay among diverse midgut receptors in the toxins’ mechanism of action in a super pest.

【 授权许可】

CC BY