Navigation Links
Using plants to silence insect genes in a high-throughput manner
Date:2/2/2012

Natural toxins against herbivores

More than 200,000 insects species are herbivores. They depend on plants for food and have adapted their metabolism accordingly in the course of evolution to render plant defenses, such as the toxins plants produce to fend off herbivores, ineffective. The operating instructions of these detoxification processes are coded in different genes. Insects have evolved an enormous diversity of adaptation mechanisms; they colonize most habitats on this planet which makes them interesting research objects in ecological studies. Which insect species attack which plants species? Which toxins or signaling substances are involved? Has the insect species adapted to one specific plant species or is it a food generalist? Interesting for agriculture: Which genes allow particular pest insects, such as the pollen beetle Meligethes aeneus or the Western corn rootworm Diabrotica virgifera virgifera, to be so destructive to crop plants? Knowing these detoxification genes and switching them off with the consequence that plant toxins are no longer effective, is currently a research subject in plant breeding. First success stories have already been reported thanks to the use of RNAi technology.

Scientists at the Max Planck Institute for Chemical Ecology examined a well-known plant toxin: nicotine. Plants of the species Nicotiana attenuata (coyote tobacco) produce nicotine as a defensive substance against herbivores. However, it does not have any toxic effects on their worst enemy: larvae of the tobacco hornworm Manduca sexta. The insect is resistant against this alkaloid; genes that encode nicotine-catabolizing enzymes may be responsible for its resistance. These so called CYP genes are involved in the formation of cytochrome P450 enzymes; the expression of some of these genes is increased as soon as the insect larvae are exposed to nicotine in their food. Ian Baldwin and his team identified the DNA sequences of CYP genes in Manduca sexta and were able to switch off these genes using RNAi technology, but expressed in the plant.

Using plants to silence insect genes

RNA interference (RNAi) is triggered by the production of double-stranded RNA (dsRNA) comprising about 300 base pairs in the cells of tobacco plants. If larvae feed on these plant, the RNA is released in the insect gut. In the experiments, the dsRNA harbored the sequence of the insect gene,CYP6B46, a special cytochrome P450 oxidoreductase specific for Manduca sexta larvae. In a next step, the dsRNA was enzymatically broken down into smaller RNA segments; a special enzyme complex called RISC (RNA-induced silencing complex), which carries several of these RNA segments, specifically binds to the messenger RNA (mRNA) of the CYP6B46 gene and disassembles the mRNA in such a way that the cytochrome P450 enzyme cannot be produced anymore. "We were impressed by the high specificity of these RNAi experiments. The analysis of mRNA transcripts of closely related CYP6 genes revealed that only the CYP6B46 gene was silenced. This means that there was no collateral damage from the procedure: the gene silencing worked on only one targeted gene," says Ian Baldwin.

The use of additional CYP RNAi probes revealed further interesting results: Young caterpillars which had ingested dsRNA of the CYP4M3 gene gained significantly less weight within 14 days in comparison to larvae reared on control plants very likely a consequence of the nicotine and its toxic effect which had been restored by switching off the CYP gene. The RNAi experiments had been conducted using plant viral vectors. Unlike genetically transformed tobacco plants in which CYP dsRNA is produced constitutively, the virus vector-based technique provides dsRNA transiently produced in wildtype tobacco plants. Both methods worked well but the "plant virus-based dsRNA producing system" (VDPS) allows for a throughput of RNAi samples that is four times faster. Many unknown functions of different insect genes involved in the adaptation of insects to their environment can now be analyzed using the VDPS technique.

However, it is still unclear how the individual steps in the RNAi mechanism from producing dsRNA in the plant cell via their uptake in the insect gut to the silencing of the detoxification genes are accomplished to induce a maximum effect. One experiment provided some interesting information: If the enzymatic step which dices dsRNA into small fragments is inhibited in the experimental plants, the amount of transcripts of the detoxification gene was reduced even further. Therefore the plant mediated RNAi procedure may be more effective, if the caterpillars ingest complete dsRNA instead of smaller diced RNA segments. [JWK, AO]


'/>"/>
Contact: Ian T. Baldwin
baldwin@ice.mpg.de
49-364-157-1101
Max Planck Institute for Chemical Ecology
Source:Eurekalert  

Related biology news :

1. NJIT professor finds engineering technique to identify disease-causing genes
2. Research shows that the pill does not deserve its reputation for causing weight gain
3. Planetary "first family" discovered by astronomers using Gemini and Keck Observatories
4. Planetary first family discovered by astronomers using Gemini and Keck Observatories
5. Using water to understand human society, from the industrial revolution to global trade
6. ESA satellites focusing on the Arctic
7. Using the other guys toolkit: Similarities of pumping blood, oil examined
8. Bangladesh Voter Registration Project Now Using MegaMatcher Biometric Technology to Detect and Prevent Duplicate Registrations
9. City of Newark first in nation using cars to power grid
10. Warmer climate causing huge increase in tree mortality across the West
11. Cancer-causing gene discovery suggests new therapies
Post Your Comments:
*Name:
*Comment:
*Email:
Related Image:
Using plants to silence insect genes in a high-throughput manner
(Date:2/14/2017)... Feb. 14, 2017  Wake Forest Baptist Medical Center ... new chief executive officer (CEO). Freischlag joins the medical ... D. McConnell , M.D., who last year announced that ... Medical Center, after leading it since 2008.   ... of Wake Forest Baptist,s academic health system, which includes ...
(Date:2/13/2017)... , Feb. 13, 2017 Former 9/11 ... Senate Judiciary Committee, Janice Kephart of Identity ... President Donald Trump,s "Executive Order: Protecting the ... (Jan. 27, 2017):  "As President Trump,s ,Travel ... Circuit has now essentially banned the travel ban, it ...
(Date:2/9/2017)... LONDON , Feb. 9, 2017 The ... in-depth analysis of the biomass boiler market globally in ... sales of biomass boilers. The market for biomass boilers ... product type, end-user, application, and country/region. The market based ... agriculture & forest residues, biogas & energy crops, urban ...
Breaking Biology News(10 mins):
(Date:2/17/2017)... ... February 17, 2017 , ... Academy of ... the world’s leading maker of unmanned aircraft systems (UAS), are launching a joint ... use drones effectively, and support educational outreach efforts. , AMA and DJI will ...
(Date:2/16/2017)... Biotherapeutics (OTCMKTS: IMMG), an early-stage biotechnology company harnessing the power ... triple negative breast cancer (TNBC), announced today their completion of ... The YEi Start in France ... their business in France and Europe.  ... complete an intensive one week immersion in France ...
(Date:2/16/2017)... Research and Markets has announced ... report to their offering. ... The study ... delivery plasmids, chassis organisms, synthetic cells, production systems), enabling ... bioinformatics and specialty media) and enabled technologies (biofuels, chemicals, ...
(Date:2/16/2017)... , Feb. 16, 2017 UCHealth ( ... utilize LungDirect for pulmonary nodule patient management. In addition ... or a spot on the lung, UCHealth looks to ... manual data entry. Stephanie Brown, RN ... my nodule patients with an Excel spreadsheet, which was ...
Breaking Biology Technology: