Ran Involved in the Development and Reproduction Is a Potential Target for RNA-Interference-Based Pest Management in Nilaparvata lugens

Abstract

Ran (RanGTPase) in insects participates in the 20-hydroxyecdysone signal transduction pathway in which downstream genes, FTZ-F1, Krüppel-homolog 1 (Kr-h1) and vitellogenin, are involved. A putative Ran gene (NlRan) was cloned from Nilaparvata lugens, a destructive phloem-feeding pest of rice. NlRan has the typical Ran primary structure features that are conserved in insects. NlRan showed higher mRNA abundance immediately after molting and peaked in newly emerged female adults. Among the examined tissues ovary had the highest transcript level, followed by fat body, midgut and integument, and legs. Three days after dsNlRan injection the NlRan mRNA abundance in the third-, fourth-, and fifth-instar nymphs was decreased by 94.3%, 98.4% and 97.0%, respectively. NlFTZ-F1 expression levels in treated third- and fourth-instar nymphs were reduced by 89.3% and 23.8%, respectively. In contrast, NlKr-h1 mRNA levels were up-regulated by 67.5 and 1.5 folds, respectively. NlRan knockdown significantly decreased the body weights, delayed development, and killed >85% of the nymphs at day seven. Two apparent phenotypic defects were observed: (1) Extended body form, and failed to molt; (2) The cuticle at the notum was split open but cannot completely shed off. The newly emerged female adults from dsNlRan injected fifth-instar nymphs showed lower levels of NlRan and vitellogenin, lower weight gain and honeydew excretion comparing with the blank control, and no offspring. Those results suggest that NlRan encodes a functional protein that was involved in development and reproduction. The study established proof of concept that NlRan could serve as a target for dsRNA-based pesticides for N. lugens control.

Ras-like family small GTPases genes in Nilaparvata lugens: Identification, phylogenetic analysis, gene expression and function in nymphal development

“…The putative start codon was preceded by an in-frame stop codon in all this twenty-nine sequences. Among the twenty-nine genes, one Ran gene has been described previously [ 32 ]. The cDNA of NlRSGs contains an entire ORF range from 540 to 845bp in length.…”

Double-Stranded RNAs in Plant Protection Against Pathogenic Organisms and Viruses in Agriculture

“…The efficiency of RNA interference induction by exogenous dsRNAs also depends on their optimal (sufficiently high) concentration, which in practical applications requires the production of large amounts of dsRNA [73][74][75]. In RNA interference experiments, dsRNAs have been produced in vitro by bidirectional transcription using T7 polymerase [76,77].…”

“…Treatment of leaves with artificially synthesized dsRNAs targeted at the genes involved in insect development significantly increases mortality and inhibits insect growth [88][89][90][91][92]. This effect can be achieved by irrigating the roots of plants by dsRNA, which leads to effective suppression of the target gene and abnormal development of insect pests [74,76,93]. RNA interference using exogenous dsRNAs can be used for a very wide range of insect genes.…”

A Genome-Wide Identification and Analysis of the Basic Helix-Loop-Helix Transcription Factors in Brown Planthopper, Nilaparvata lugens

“…The sequencing of N. lugens genome aids the identification of genes that are involved in molting, reproduction and wing development as potential targets for RNA-interference-based management [21,22,23]. bHLHs, as important transcription factors, could be effective targets of RNA interference (RNAi).…”