Environmental RNAi in herbivorous insects.

Environmental RNAi (eRNAi) is a sequence-specific regulation of endogenous gene expression in a receptive organism by exogenous double-stranded RNA (dsRNA). Although demonstrated under artificial dietary conditions and via transgenic plant presentations in several herbivorous insects, the magnitude and consequence of exogenous dsRNA uptake and the role of eRNAi remains unknown under natural insect living conditions. Our analysis of coleopteran insects sensitive to eRNAi fed on wild-type plants revealed uptake of plant endogenous long dsRNAs, but not small RNAs.

Establishing an in vivo assay system to identify components involved in environmental RNA interference in the western corn rootworm.

The discovery of environmental RNA interference (RNAi), in which gene expression is suppressed via feeding with double-stranded RNA (dsRNA) molecules, opened the door to the practical application of RNAi-based techniques in crop pest management. The western corn rootworm (WCR, Diabrotica virgifera virgifera) is one of the most devastating corn pests in North America. Interestingly, WCR displays a robust environmental RNAi response, raising the possibility of applying an RNAi-based pest management strategy to this pest.

Genetically engineered crops: from idea to product.

Genetically engineered crops were first commercialized in 1994 and since then have been rapidly adopted, enabling growers to more effectively manage pests and increase crop productivity while ensuring food, feed, and environmental safety. The development of these crops is complex and based on rigorous science that must be well coordinated to create a plant with desired beneficial phenotypes. This article describes the general process by which a genetically engineered crop is developed from an initial concept to a commercialized product.

Ultrastructural changes caused by Snf7 RNAi in larval enterocytes of western corn rootworm (Diabrotica virgifera virgifera Le Conte).

The high sensitivity to oral RNA interference (RNAi) of western corn rootworm (WCR, Diabrotica virgifera virgifera Le Conte) provides a novel tool for pest control. Previous studies have shown that RNAi of DvSnf7, an essential cellular component of endosomal sorting complex required for transport (ESCRT), caused deficiencies in protein de-ubiquitination and autophagy, leading to WCR death. Here we investigated the detailed mechanism leading to larval death by analyzing the ultrastructural changes in midgut enterocytes of WCR treated with double-stranded RNA (ds-DvSnf7).

Quantification of transgene-derived double-stranded RNA in plants using the QuantiGene nucleic acid detection platform.

The expanding use of RNA interference (RNAi) in agricultural biotechnology necessitates tools for characterizing and quantifying double-stranded RNA (dsRNA)-containing transcripts that are expressed in transgenic plants. We sought to detect and quantify such transcripts in transgenic maize lines engineered to control western corn rootworm (Diabrotica virgifera virgifera LeConte) via overexpression of an inverted repeat sequence bearing a portion of the putative corn rootworm orthologue of yeast Snf7 (DvSnf7), an essential component of insect cell receptor sorting. A quantitative assay was developed to detect DvSnf7 sense strand-containing dsRNA transcripts that is based on the QuantiGene Plex 2.

Physiological and cellular responses caused by RNAi- mediated suppression of Snf7 orthologue in western corn rootworm (Diabrotica virgifera virgifera) larvae.

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.

Characterizing the mechanism of action of double-stranded RNA activity against western corn rootworm (Diabrotica virgifera virgifera LeConte).

RNA interference (RNAi) has previously been shown to be effective in western corn rootworm (WCR, Diabrotica virgifera virgifera LeConte) larvae via oral delivery of synthetic double-stranded RNA (dsRNA) in an artificial diet bioassay, as well as by ingestion of transgenic corn plant tissues engineered to express dsRNA. Although the RNAi machinery components appear to be conserved in Coleopteran insects, the key steps in this process have not been reported for WCR. Here we characterized the sequence of events that result in mortality after ingestion of a dsRNA designed against WCR larvae.

Hypovirus papain-like protease p29 suppresses RNA silencing in the natural fungal host and in a heterologous plant system.

Virulence-attenuating hypoviruses of the species Cryphonectria hypovirus 1 (CHV1) encode a papain-like protease, p29, that shares similarities with the potyvirus-encoded suppressor of RNA silencing HC-Pro. We now report that hypovirus CHV1-EP713-encoded p29 can suppress RNA silencing in the natural host, the chestnut blight fungus Cryphonectria parasitica. Hairpin RNA-triggered silencing was suppressed in C.

Evidence for a role of the regulator of G-protein signaling protein CPRGS-1 in Galpha subunit CPG-1-mediated regulation of fungal virulence, conidiation, and hydrophobin synthesis in the chestnut blight fungus Cryphonectria parasitica.

We previously reported that the chestnut blight fungus Cryphonectria parasitica expresses at least three G-protein alpha subunits and that Galpha subunit CPG-1 is essential for regulated growth, pigmentation, sporulation, and virulence. We now report the cloning and characterization of a C. parasitica regulator of G-protein signaling (RGS) protein, CPRGS-1.