Core RNAi Machinery and Sid1, a Component for Systemic RNAi, in the Hemipteran Insect, Aphis glycines.

Raman Bansal,Andy P Michel

doi:10.3390/ijms14023786

Abstract

RNA interference (RNAi) offers a novel tool to manage hemipteran pests. For the success of RNAi based pest control in the field, a robust and systemic RNAi response is a prerequisite. We identified and characterized major genes of the RNAi machinery, Dicer2 (Dcr2), Argonaute2 (Ago2), and R2d2 in Aphis glycines, a serious pest of soybean. The A. glycines genome encodes for at least one copy of Dcr2, R2d2 and Ago2. Comparative and molecular evolution analyses (dN/dS) showed that domain regions of encoded proteins are highly conserved, whereas linker (non-domain) regions are diversified. Sequence homology and phylogenetic analyses suggested that the RNAi machinery of A. glycines is more similar to that of Tribolium casteneum as compared to that of Drosophila melanogaster. We also characterized Sid1, a major gene implicated in the systemic response for RNAi-mediated gene knockdown. Through qPCR, Dcr2, R2d2, Ago2, and Sid1 were found to be expressed at similar levels in various tissues, but higher expression of Dcr2, R2d2, and Ago2 was seen in first and second instars. Characterization of RNAi pathway and Sid1 in A. glycines will provide the foundation of future work for controlling one of the most important insect pests of soybean in North America.

Highlights

Transgenic crops have emerged as an effective tool for global insect pest management
A search for major components of RNA interference (RNAi) machinery in an A. glycines transcriptomic database revealed that its genome encodes for at least one copy of Dcr2, R2d2 and Ago2 (Table 1) as well as the cDNA of a putative factor for systemic RNAi, systemic RNA interference deficient-1 (Sid1)
The proteins encoded by AyDcr2, AyR2d2, AyAgo2, and AySid1 have a high level of sequence similarity with corresponding proteins of various insects and, as expected, their closest matches were other Aphididae species, A. pisum or

Summary

Introduction

Transgenic crops have emerged as an effective tool for global insect pest management. Plants expressing cry-toxin encoding genes from the bacterium Bacillus thuringienesis (Bt) have achieved tremendous success for managing various lepidopteran and coleopteran pests [1,2]. The inefficacy of Bt toxins to hemipteran insects has been attributed to various factors [6,7,8,9,10,11] but the exact cause remains unclear, providing difficult challenges for managing this group of economically important crop pests. For RNAi-based crops to be successful, the target insect must express the RNAi pathway as well as a systemic response that leads to dsRNA processing and mRNA degradation in several tissues, not just the midgut where dsRNA uptake occurs [14]. In the absence of proper genetic machinery or systemic RNAi, the resulting knockdown will have either no effect or be localized (which may or may not cause mortality)

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Discussion

Conclusion