Cytochrome P450 Gene Families: Role in Plant Secondary Metabolites Production and Plant Defense.

Cytochrome P450s (CYPs) are the most prominent family of enzymes involved in NADPH- and O2-dependent hydroxylation processes throughout all spheres of life. CYPs are crucial for the detoxification of xenobiotics in plants, insects, and other organisms. In addition to performing this function, CYPs serve as flexible catalysts and are essential for producing secondary metabolites, antioxidants, and phytohormones in higher plants. Numerous biotic and abiotic stresses frequently affect the growth and development of plants. They cause a dramatic decrease in crop yield and a deterioration in crop quality. Plants protect themselves against these stresses through different mechanisms, which are accomplished by the active participation of CYPs in several biosynthetic and detoxifying pathways. There are immense potentialities for using CYPs as a candidate for developing agricultural crop species resistant to biotic and abiotic stressors. This review provides an overview of the plant CYP families and their functions to plant secondary metabolite production and defense against different biotic and abiotic stresses.

Chromosome-level genome of black cutworm provides novel insights into polyphagy and seasonal migration in insects.

BACKGROUND: The black cutworm, Agrotis ipsilon, is a serious global underground pest. Its distinct phenotypic traits, especially its polyphagy and ability to migrate long distances, contribute to its widening distribution and increasing difficulty of control. However, knowledge about these traits is still limited. RESULTS: We generated a high-quality chromosome-level assembly of A. ipsilon using PacBio and Hi-C technology with a contig N50 length of ~ 6.7 Mb. Comparative genomic and transcriptomic analyses showed that detoxification-associated gene families were highly expanded and induced after insects fed on specific host plants. Knockout of genes that encoded two induced ABC transporters using CRISPR/Cas9 significantly reduced larval growth rate, consistent with their contribution to host adaptation. A comparative transcriptomic analysis between tethered-flight moths and migrating moths showed expression changes in the circadian rhythm gene AiCry2 involved in sensing photoperiod variations and may receipt magnetic fields accompanied by MagR and in genes that regulate the juvenile hormone pathway and energy metabolism, all involved in migration processes. CONCLUSIONS: This study provides valuable genomic resources for elucidating the mechanisms involved in moth migration and developing innovative control strategies.

Sf-FGFR and Sf-SR-C Are Not the Receptors for Vip3Aa to Exert Insecticidal Toxicity in Spodoptera frugiperda.

Vip3Aa is a novel insecticidal protein secreted by Bacillus thuringiensis (Bt) during its vegetative growth stages. It has high insecticidal activity against lepidopteran pests such as Spodoptera frugiperda, and has no cross-resistance with Cry insecticidal proteins. As a new type of insecticide, it plays an important role in controlling agricultural pests. However, the insecticidal mechanism of the Vip3Aa toxin, especially its definite receptors, have not been fully revealed. In this study, the previously reported Vip3Aa receptor genes Sf-FGFR and Sf-SR-C were knocked out separately using the CRISPR/Cas9 system. Bioassay results showed that the sensitivity of these two knockout strains to Vip3Aa were not significantly changed compared to that of the normal strain. The current results are not consistent with the previously reports that Sf-SR-C and Sf-FGFR were the receptors of Vip3Aa in vitro. This suggests that the Sf-SR-C and Sf-FGFR genes we tested may not be critical in the mode of action of Vip3Aa in vivo in Spodoptera frugiperda.

Two ABC transporters are differentially involved in the toxicity of two Bacillus thuringiensis Cry1 toxins to the invasive crop-pest Spodoptera frugiperda (J. E. Smith).

BACKGROUND: The fall armyworm Spodoptera frugiperda is a major agricultural pest that has invaded the East Hemisphere since 2016, generating a serious threat to food security worldwide including Africa and Asia. The Cry toxins produced by Bacillus thuringiensis (Bt) have been shown to be effective against this insect pest. In different insect ABC transporters (ABCC2 or ABCC3) have been shown to be involved as receptors of some Cry1 toxins. Here we analyzed the role of SfABCC2 and SfABCC3 in the toxicity of Cry1Fa and Cry1Ab toxins in this insect pest. RESULTS: Two S. frugiperda SfABCC2 and SfABCC3 knockout strains, coding for potential functional Bt receptors, were created using CRISPR/Cas9 genome editing system. Both knockout strains showed resistance to both Cry1Fa and Cry1Ab toxins compared with the susceptible strain. SfABCC2 knockout strain showed higher resistance to both Cry toxins than SfABCC3 knockout strain, suggesting a major role of SfABCC2 in the mode of action of these Cry toxins. In addition, expression of SfABCC2 and SfABCC3 genes in Trichoplusia ni Hi5 cells also increased the susceptibility to Cry1Ab and Cry1Fa toxins, in agreement with the genome editing results. The double knockout of SfABCC2 and SfABCC3 strain was not viable in contrast to other lepidopteran species. Furthermore, we report here that SfABCC2 or SfABCC3 knockout strains increased their susceptibility to abamectin and spinosad insecticides. CONCLUSION: We provide functional evidence that in S. frugiperda these two ABCC transporters serve as receptors of Bt Cry1Fa and Cry1Ab toxins. © 2020 Society of Chemical Industry.

Genetic structure and insecticide resistance characteristics of fall armyworm populations invading China.

The rapid wide-scale spread of fall armyworm (Spodoptera frugiperda) has caused serious crop losses globally. However, differences in the genetic background of subpopulations and the mechanisms of rapid adaptation behind the invasion are still not well understood. Here we report the assembly of a 390.38-Mb chromosome-level genome of fall armyworm derived from south-central Africa using Pacific Bioscience (PacBio) and Hi-C sequencing technologies, with scaffold N50 of 12.9 Mb and containing 22,260 annotated protein-coding genes. Genome-wide resequencing of 103 samples and strain identification were conducted to reveal the genetic background of fall armyworm populations in China. Analysis of genes related to pesticide- and Bacillus thuringiensis (Bt) resistance showed that the risk of fall armyworm developing resistance to conventional pesticides is very high. Laboratory bioassay results showed that insects invading China carry resistance to organophosphate and pyrethroid pesticides, but are sensitive to genetically modified maize expressing the Bt toxin Cry1Ab in field experiments. Additionally, two mitochondrial fragments were found to be inserted into the nuclear genome, with the insertion event occurring after the differentiation of the two strains. This study represents a valuable advance toward improving management strategies for fall armyworm.

Down-regulation of lysosomal protein ABCB6 increases gossypol susceptibility in Helicoverpa armigera.

Cotton bollworm (Helicoverpa armigera) is the major insect herbivore of cotton plants. As its larvae feed and grow on cotton, H. armigera can likely tolerate gossypol, the main defense metabolite produced by cotton plants, through detoxification and sequestration mechanisms. Recent reports have shown that various P450 monooxygenases and UDP-glycosyltransferases in H. armigera are involved in gossypol detoxification, while the roles of ABC transporters, another gene family widely associated with metabolite detoxification, remain to be elucidated. Here, we show that ingestion of gossypol-infused artificial diet and cotton leaves significantly induced the expression of HaABCB6 in H. armigera larvae. Knockdown and knockout of HaABCB6 increased sensitivity of H. armigera larvae to gossypol. Moreover, HaABCB6-GFP fusion protein was localized on lysosomes in Hi5 cells and its overexpression significantly enhanced gossypol tolerance in vitro. These experimental results strongly support that HaABCB6 plays an important role in gossypol detoxification by H. armigera.

Bacillus thuringiensis vegetative insecticidal protein family Vip3A and mode of action against pest Lepidoptera.

Vip3A proteins are widely used for controlling pest Lepidoptera. Different binding sites with different receptors in the insect midgut membrane and lack of cross-resistance with crystal (Cry) proteins enhance their applicability, as both single proteins and proteins pyramided with Cry proteins in transgenic Bt crops. Vip3A proteins are effective but there is relatively little information about their structure, function, activation, specificity, and mode of action. In addition, the mechanism of insect resistance to these proteins is unknown. Phylogenetic analysis and multiple sequence alignment showed that Vip3A proteins are genetically distant from Cry proteins. The mode of action and insecticidal activity of Vip3A proteins are discussed in this review. This review also provides detailed information about the Vip3A protein family that may aid in the design of more efficient pest management strategies in response to insect resistance to insecticidal proteins. © 2020 Society of Chemical Industry.

Insect ATP-Binding Cassette (ABC) Transporters: Roles in Xenobiotic Detoxification and Bt Insecticidal Activity.

ATP-binding cassette (ABC) transporters, a large class of transmembrane proteins, are widely found in organisms and play an important role in the transport of xenobiotics. Insect ABC transporters are involved in insecticide detoxification and Bacillus thuringiensis (Bt) toxin perforation. The complete ABC transporter is composed of two hydrophobic transmembrane domains (TMDs) and two nucleotide binding domains (NBDs). Conformational changes that are needed for their action are mediated by ATP hydrolysis. According to the similarity among their sequences and organization of conserved ATP-binding cassette domains, insect ABC transporters have been divided into eight subfamilies (ABCA-ABCH). This review describes the functions and mechanisms of ABC transporters in insecticide detoxification, plant toxic secondary metabolites transport and insecticidal activity of Bt toxin. With improved understanding of the role and mechanisms of ABC transporter in resistance to insecticides and Bt toxins, we can identify valuable target sites for developing new strategies to control pests and manage resistance and achieve green pest control.

Transcriptional response of ATP-binding cassette (ABC) transporters to insecticides in the cotton bollworm, Helicoverpa armigera.

When any living organism is frequently exposed to any drugs or toxic substances, they evolve different detoxification mechanism to confront with toxicants during absorption and metabolism. Likewise, the insects have evolved detoxification mechanisms as they are frequently exposed to different toxic secondary plant metabolites and commercial insecticides. ABC transporter superfamily is one of the largest and ubiquitous group of proteins which play an important role in phase III of the detoxification process. However, knowledge about this gene family remains largely unknown. To help fill this gap, we have identified a total of 54 ABC transporters in the Helicoverpa armigera genome which are classified into eight subfamilies (A-H) by phylogenetic analysis. The temporal and spatial expression profiles of these 54 ABC transporters throughout H. armigera development stages and seven tissues and their responses to five different insecticides, were investigated using RNA-seq analysis. Furthermore, the mRNA expression of eight selected genes in different tissues and six genes responses to insecticides were confirmed by the quantitative real-time PCR (RT-qPCR). Moreover, H. armigera become more sensitive to abamectin and indoxacarb when P-gp was inhibited. These results provide a foundation for further studies of ABCs in H. armigera.

Comparative Proteomics of Peritrophic Matrix Provides an Insight into its Role in Cry1Ac Resistance of Cotton Bollworm Helicoverpa armigera.

Crystalline (Cry) proteins from Bacillus thuringiensis (Bt) are widely used in sprays and transgenic crops to control insect pests, but the evolution of insect resistance threatens their long-term use. Different resistance mechanisms have been identified, but some have not been completely elucidated. Here, the transcriptome of the midgut and proteome of the peritrophic matrix (PM) were comparatively analyzed to identify potential mechanism of resistance to Cry1Ac in laboratory-selected strain XJ10 of Helicoverpa armigera. This strain had a 146-fold resistance to Cry1Ac protoxin and 45-fold resistance to Cry1Ac activated toxin compared with XJ strain. The mRNA and protein levels for several trypsin genes were downregulated in XJ10 compared to the susceptible strain XJ. Furthermore, 215 proteins of the PM were identified, and nearly all had corresponding mRNAs in the midgut. These results provide new insights that the PM may participate in Bt resistance.