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Double-Stranded RNA-Degrading Enzymes Reduce the Efficiency of RNA Interference in Plutella xylostella.

ABSTRACT: DsRNA-degrading enzymes (dsRNases) have been recognized as important factors in reducing RNA interference (RNAi) efficiency in different insect species. However, dsRNases in Plutella xylostella are still unknown. We identified the full-length cDNAs of PxdsRNase1, PxdsRNase2, PxdsRNase3, and PxdsRNase4. Gene expression profile showed that PxdsRNase1 was mainly expressed in the hemolymph; and that PxdsRNase2 and PxdsRNase3 were mainly expressed in the intestinal tract. The expression of PxCht (Chitinase of P. xylostella) in P. xylostella larvae injected with the mixture of dsPxCht (dsRNA of PxCht) and dsPxdsRNase1 (dsRNA of PxdsRNase1), dsPxdsRNase2 (dsRNA of PxdsRNase2), or dsPxdsRNase3 (dsRNA of PxdsRNase3) was significantly higher than that in the larvae injected with the mixture of dsGFP (dsRNA of green fluorescent protein gene, GFP) and dsPxCht; the transcription level of PxCht in the larvae feeding on the mixture of dsPxCht and dsPxdsRNase1, dsPxdsRNase2, or dsPxdsRNase3 was significantly higher than that in the larvae feeding on the mixture of dsPxCht and dsGFP. The recombinant protein of PxdsRNase1 degraded dsRNA rapidly, PxdsRNase3 cleaved dsRNA without complete degradation, and PxdsRNase2 could not degrade dsRNA in vitro. These results suggested that PxdsRNases1, PxdsRNases2, and PxdsRNases3 were involved in the dsRNA degradation to reduce RNAi efficiency with different mechanisms.

SUBMITTER: Chen JZ

PROVIDER: S-EPMC8396913 | biostudies-literature | 2021 Aug

REPOSITORIES: biostudies-literature

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Double-Stranded RNA-Degrading Enzymes Reduce the Efficiency of RNA Interference in Plutella xylostella.

Chen Jin-Zhi JZ Jiang Ying-Xia YX Li Miao-Wen MW Li Jian-Wen JW Zha Ben-Hu BH Yang Guang G

Insects 20210809 8

DsRNA-degrading enzymes (dsRNases) have been recognized as important factors in reducing RNA interference (RNAi) efficiency in different insect species. However, dsRNases in Plutella xylostella are still unknown. We identified the full-length cDNAs of PxdsRNase1, PxdsRNase2, PxdsRNase3, and PxdsRNase4. Gene expression profile showed that PxdsRNase1 was mainly expressed in the hemolymph; and that PxdsRNase2 and PxdsRNase3 were mainly express ...[more]

PMID: 34442278

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Project description:Ecdysteroids play an essential role in controlling insect development and reproduction. Their pathway is regulated by a group of enzymes called Halloween gene proteins. The relationship between the Halloween genes and ecdysteroid synthesis has yet to be clearly understood in diamondback moth, Plutella xylostella (L.), a worldwide Lepidoptera pest attacking cruciferous crops and wild plants. In this study, complete sequences for six Halloween genes, neverland (nvd), shroud (sro), spook (spo), phantom (phm), disembodied (dib), shadow (sad), and shade (shd), were identified. Phylogenetic analysis revealed a strong conservation in insects, including Halloween genes of P. xylostella that was clustered with all other Lepidoptera species. Three Halloween genes, dib, sad, and shd were highly expressed in the adult stage, while nvd and spo were highly expressed in the egg and pupal stages, respectively. Five Halloween genes were highly expressed specifically in the prothorax, which is the major site of ecdysone production. However, shd was expressed predominantly in the fat body to convert ecdysone into 20-hydroxyecdysone. RNAi-based knockdown of sad, which is involved in the last step of ecdysone biosynthesis, significantly reduced the 20E titer and resulted in a longer developmental duration and lower pupation of fourth-instar larvae, as well as caused shorter ovarioles and fewer fully developed eggs of P. xylostella. Furthermore, after the knockdown of sad, the expression levels of Vg and VgR genes were significantly decreased by 77.1 and 53.0%. Meanwhile, the number of eggs laid after 3 days was significantly reduced in sad knockdown females. These results suggest that Halloween genes may play a critical role in the biosynthesis of ecdysteroids and be involved in the development and reproduction of P. xylostella. Our work provides a solid basis for understanding the functional importance of these genes, which will help to screening potential genes for pest management of P. xylostella.

Dataset Information

Double-Stranded RNA-Degrading Enzymes Reduce the Efficiency of RNA Interference in Plutella xylostella.

Publications

Double-Stranded RNA-Degrading Enzymes Reduce the Efficiency of RNA Interference in <i>Plutella xylostella</i>.

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