Abstract
Ingestion of double stranded RNA (dsRNA) has been previously demonstrated to be effective in triggering RNA interference (RNAi) in western corn rootworm (WCR, Diabrotica virgifera virgifera LeConte), providing potential novel opportunities for insect pest control. The putative Snf7 homolog of WCR (DvSnf7) has previously been shown to be an effective RNAi target for insect control, as DvSnf7 RNAi leads to lethality of WCR larvae. Snf7 functions as a part of the ESCRT (Endosomal Sorting Complex Required for Transport) pathway which plays a crucial role in cellular housekeeping by internalization, transport, sorting and lysosomal degradation of transmembrane proteins. To understand the effects that lead to death of WCR larvae by DvSnf7 RNAi, we examined some of the distinct cellular processes associated with ESCRT functions such as de-ubiquitination of proteins and autophagy. Our data indicate that ubiquitinated proteins accumulate in DvSnf7 dsRNA-fed larval tissues and that the autophagy process seems to be impaired. These findings suggest that the malfunctioning of these cellular processes in both midgut and fat body tissues triggered by DvSnf7 RNAi were the main effects leading to the death of WCR. This study also illustrates that Snf7 is an essential gene in WCR and its functions are consistent with biological functions described for other eukaryotes.
Highlights
Gene silencing by RNA interference (RNAi) has been used as a powerful tool for studying gene function in a variety of organisms
We have demonstrated that feeding of double-stranded RNA (dsRNA) of a putative western corn rootworm (WCR) Snf7 homolog (DvSnf7) to larvae caused severe stunting after five days of exposure followed by the death of the larvae
In order to gain a better understanding of the cellular processes that lead to mortality of WCR larvae after ingestion of Snf7 dsRNA, we investigated the functions of Snf7 by observing changes in the endosomalautophagic pathway at cellular level after DvSnf7 dsRNA treatment of WCR larvae
Summary
Gene silencing by RNA interference (RNAi) has been used as a powerful tool for studying gene function in a variety of organisms. We mapped the endosomal protein sorting activity in the midguts and fat bodies of DvSnf7 and GFP dsRNA-fed larvae using an anti-ubiquitin antibody and immunohistochemistry (Fig. 2). No fluorescent signals or puncta were observed in the midguts and fat bodies of either DvSnf7 or GFP dsRNA-fed larvae in the absence of primary anti-ubiquitin antibody (negative control; data not shown).
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