Molecular farming and metabolic engineering promise a new generation of high-tech crops.
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The cotton boll weevil, Anthonomus grandis, is an economically important pest of cotton in tropical and subtropical areas of several countries in the Americas, causing severe losses due to their damage in cotton floral buds. Enzymatic assays using gut extracts from larval and adult boll weevil have demonstrated the presence of digestive serine proteinase-like activities. Furthermore, in vitro assays showed that soybean Kunitz trypsin inhibitor (SKTI) was able to inhibit these enzymes. Previously, in vivo effects of black-eyed pea trypsin chymotrypsin inhibitor (BTCI) have been demonstrated towards the boll weevil pest. Here, when neonate larvae were reared on an artificial diet containing SKTI at three different concentrations, a reduction of larval weight of up to 64% was observed for highest SKTI concentration 500 microM. The presence of SKTI caused an increase in mortality and severe deformities of larvae, pupae and adult insects. This work therefore represents the first observation of a Kunitz trypsin inhibitor active in vivo and in vitro against A. grandis. Bioassays suggested that SKTI could be used as a tool in engineering crop plants, which might exhibit increased resistance against cotton boll weevil.
Chloroplast genetic engineering offers several unique advantages, including high-level transgene expression, multi-gene engineering in a single transformation event and transgene containment by maternal inheritance, as well as a lack of gene silencing, position and pleiotropic effects and undesirable foreign DNA. More than 40 transgenes have been stably integrated and expressed using the tobacco chloroplast genome to confer desired agronomic traits or express high levels of vaccine antigens and biopharmaceuticals. Despite such significant progress, this technology has not been extended to major crops. However, highly efficient soybean, carrot and cotton plastid transformation has recently been accomplished through somatic embryogenesis using species-specific chloroplast vectors. This review focuses on recent exciting developments in this field and offers directions for further research and development.
Voltage-gated sodium channels are a major target for toxins and insecticides due to their central role in excitability, but due to the conservation of these channels in Animalia most insecticides do not distinguish between those of insects and mammals, thereby imposing risks to humans and livestock. Evidently, as long as modern agriculture depends heavily on the use of insecticides there is a great need for new substances capable of differentiating between sodium channel subtypes. Such substances exist in venomous animals, but ways for their exploitation have not yet been developed due to problems associated with manufacturing, degradation, and delivery to the target channels. Engineering of plants for expression of anti-insect toxins or use of natural vectors that express toxins near their target site (e.g. baculoviruses) are still problematic and raise public concern. In this problematic reality a rational approach might be to learn from nature how to design highly selective anti-insect compounds preferably in the form of peptidomimetics. This is a complex task that requires the elucidation of the face of interaction between insect-selective toxins and their sodium channel receptor sites. This review delineates current progress in: (i) elucidation of the bioactive surfaces of scorpion beta-toxins, especially the excitatory and depressant groups, which show high preference for insects and bind insect sodium channels with high affinity; (ii) studies of the mode of interaction of scorpion beta-toxins with receptor site-4 on voltage-gated sodium channels; and (iii) clarification of channel elements that constitute receptor site-4. This information may be useful in future attempts to mimic the bioactive surface of the toxins for the design of anti-insect selective peptidomimetics.
The cotton boll weevil Anthonomus grandis (Boheman) is one of the major pests of cotton (Gossypium hirsutum L.) in tropical and sub-tropical areas of the New World. This feeds on cotton floral fruits and buds causing severe crop losses. Digestion in the boll weevil is facilitated by high levels of serine proteinases, which are responsible for the almost all proteolytic activity. Aiming to reduce the proteolytic activity, the inhibitory effects of black-eyed pea trypsin/chymotrypsin inhibitor (BTCI), towards trypsin and chymotrypsin from bovine pancreas and from midguts of A. grandis larvae and adult insects were analyzed. BTCI, purified from Vigna unguiculata (L.) seeds, was highly active against different trypsin-like proteinases studied and moderately active against the digestive chymotrypsin of adult insects. Nevertheless, no inhibitory activity was observed against chymotrypsin from A. grandis larval guts. To test the BTCI efficiency in vivo, neonate larvae were reared on artificial diet containing BTCI at 10, 50 and 100 microM. A reduction of larval weight of up to approximately 54% at the highest BTCI concentration was observed. At this concentration, the insect mortality was 65%. This work constitutes the first observation of a Bowman-Birk type inhibitor active in vitro and in vivo toward the cotton boll weevil A. grandis. The results of bioassays strongly suggest that BTCI may have potential as a transgene protein for use in engineered crop plants modified for heightened resistance to the cotton boll weevil.
Growing interest in biorational pesticides has placed the Bacillus thuringiensis insecticidal crystal proteins at the forefront of pesticides for plant genetic engineering. The development of improvement pesticides, both in enhanced activity and broader host range, depends on an understanding of its mechanism of action. This review presents a complete overview of the bacterium and the group of insecticidal proteins known as Cry proteins or delta-endotoxins. The molecular mode of action is described in detail, including the mapping of receptor binding sites by site-directed mutagenesis, the known receptors, and the ion-channel activity of the toxins.
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Gene silencing through the increased degradation of mRNA appears to represent a novel cellular pathway that is functional in a broad range of organisms. Recent work has established a role for RNA silencing in host antiviral defense and transposon silencing, suggesting a potential application in plant functional genomics.
The treatment of environmental pollution by microorganisms is a promising technology. Various genetic approaches have been developed and used to optimize the enzymes, metabolic pathways and organisms relevant for biodegradation. New information on the metabolic routes and bottlenecks of degradation is still accumulating, enlarging the available toolbox. With molecular methods allowing the characterization of microbial community structure and activities, the performance of microorganisms under in situ conditions and in concert with the indigenous microflora will become predictable.
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Plants are an important source of essential nutrients and health-beneficial components that are crucial for human life. Because the intake of these phytochemicals is not always adequate, the resources of plant biotechnology are being used to enhance the nutritional quality of our plant-based food supply. Various improvement strategies are feasible, depending on whether the phytochemical target is a major or minor constituent. Recent efforts in gene discovery and functional genomics are providing the necessary understanding to develop and evaluate different approaches to manipulate phytochemical composition.
Trees represent a unique life form of upmost importance for mankind, as these organisms have developed a perennial lifestyle that produces the majority of terrestrial biomass. The difference between trees and annual plants is one of the main arguments behind the effort to sequence the entire genome of the poplar tree. This initiative is being backed up with a full-scale functional genomics effort on trees that will set a completely new agenda for forest research.
The legal battles over GM crop trials in Europe are hotting up, reports Nigel Williams.
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