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PubMed2006

Antibody-based metabolic engineering in plants.

Genetic engineering is a powerful tool for the manipulation of cellular metabolism and the development of plant varieties with enhanced biological and nutrional functions. Several strategies are available for the in vivo modulation of enzymatic activities, allowing metabolic flux to be directed towards desired biochemical products. Such strategies include the simultaneous expression and/or suppression of multiple genes encoding rate-limiting enzymes, ectopic expression of transcription factors, and the RNA-based inhibition of catabolic enzymes. As an alternative approach, recombinant antibodies expressed in plants have been used to inactivate or sequestrate specific host proteins or compounds, resulting in significant changes to metabolic pathways. The impact of this approach depends on prudent selection of the target antigen, careful antibody design, appropriate subcellular targeting and stable accumulation of the recombinant antibodies in planta. Here, we describe the current status of antibody-based metabolic engineering in plants, discuss procedures for the optimisation of this technology and consider the remaining challenges to its widespread use.

Antibodies↗
PubMed2005

Up-regulation of a H+-pyrophosphatase (H+-PPase) as a strategy to engineer drought-resistant crop plants.

Engineering drought -resistant crop plants is a critically important objective. Overexpression of the vacuolar H(+)-pyrophosphatase (H(+)-PPase) AVP1 in the model plant Arabidopsis thaliana results in enhanced performance under soil water deficits. Recent work demonstrates that AVP1 plays an important role in root development through the facilitation of auxin fluxes. With the objective of improving crop performance, we expressed AVP1 in a commercial cultivar of tomato. This approach resulted in (i) greater pyrophosphate-driven cation transport into root vacuolar fractions, (ii) increased root biomass, and (iii) enhanced recovery of plants from an episode of soil water deficit stress. More robust root systems allowed transgenic tomato plants to take up greater amounts of water during the imposed water deficit stress, resulting in a more favorable plant water status and less injury. This study documents a general strategy for improving drought resistance of crops.

Arabidopsis↗
PubMed2005

An alternative strategy for sustainable pest resistance in genetically enhanced crops.

Bacillus thuringiensis (Bt) crystal protein genes encode insecticidal delta-endotoxins that are widely used for the development of insect-resistant crops. In this article, we describe an alternative transgenic strategy that has the potential to generate broader and more sustainable levels of resistance against insect pests. Our strategy involves engineering plants with a fusion protein combining the delta-endotoxin Cry1Ac with the galactose-binding domain of the nontoxic ricin B-chain (RB). This fusion, designated BtRB, provides the toxin with additional, binding domains, thus increasing the potential number of interactions at the molecular level in target insects. Transgenic rice and maize plants engineered to express the fusion protein were significantly more toxic in insect bioassays than those containing the Bt gene alone. They were also resistant to a wider range of insects, including important pests that are not normally susceptible to Bt toxins. The potential impact of fusion genes such as BtRB in terms of crop improvement, resistance sustainability, and biosafety is discussed.

Analysis of Variance↗
PubMed1996

A survey of mortality at two automotive engine manufacturing plants.

Mortality at two engine plants was analyzed using proportional mortality and logistic regression models of mortality odds ratios to expand previous observations of increased cancers of the stomach, pancreas, and bladder, and cirrhosis of the liver among workers exposed to machining fluids. Causes of death and work histories were available for 1,870 decendents. There was a significant excess of deaths coded as diabetes for white men in both plants (PMR = 25/16.7 = 1.5, 95% CI = 1.02, 2.20), and a deficit of respiratory diseases. Black men had fewer than expected diabetes deaths and more emphysema deaths. Elevated PMRs for cancers of the stomach, pancreas, prostate, bladder, and kidney were not statistically significant in plantwide populations. However, stomach cancer mortality increased with duration in camshaft and crankshaft production at Plant 1 (OR = 5.1, 95% CI = 1.6, 17; at mean duration of exposed cases), and among tool room workers (OR = 6.3, 95% CI = 1.3, 31), but these results were based on five cases. Nitrosamines were probably present in camshaft and crankshaft grinding at Plant 1. Pancreas cancer risk increased among workers at both plants ever employed in inspection (OR = 2.5, 16), in machining with straight oil (OR = 3.6, 95% CI = 1.04, 12), or in skilled trades (OR = 2.9, 95% CI = 1.1, 7.5). Lung cancer increased in cylinder head machining (OR = 3.9, 95% CI = 1.4, 11), millwright work (OR = 3.8, 95% CI = 1.6, 9.0), and in Plant 2 generally (OR = 1.45, 95% CI = 0.97, 2.2). Potential lung carcinogens included heat treatment emissions, chlorinated oils, and coal tar fumes (millwrights). Bladder cancer increased with duration among workers grinding in straight oil MF (OR = 3.0, 95% CI = 1.15, 7.8) and in machining/heat-treat operations (OR = 2.9, 95% CI = 1.14, 7.2).

Automobiles↗
PubMed2007

Functionality of the beta/six site-specific recombination system in tobacco and Arabidopsis: a novel tool for genetic engineering of plant genomes.

The beta recombinase is a member of the prokaryotic site-specific serine recombinases (invertase/resolvase family), which in the presence of a DNA bending cofactor can catalyse DNA deletions between two directly oriented 90-bp six recombination sites. We have examined here whether the beta recombinase can be expressed in plants and whether it displays in planta its specific catalytic activity excising DNA sequences that are flanked by six sites. In plant protoplasts, the enzyme could be expressed as a GFP-beta recombinase fusion which can localise to the cell nucleus. Beta recombinase stably expressed in tobacco plants can catalyse deletion of a spacer region that is flanked by directly oriented six sites and has been placed between promoter and a GUS reporter gene (preventing GUS expression). In transient transformation experiments, beta recombinase-mediated elimination of the spacer results in transcriptional induction of the GUS gene. Similarly, beta recombinase in stably double-transformed Arabidopsis plants deletes specifically the spacer region of a reporter construct that has been incorporated into the genome. In the segregating T1 generation, plants were identified that contain exclusively the recombined reporter construct. In summary, our results demonstrate that functional / recombinase can be expressed in plants and that the enzyme is suitable to precisely eliminate undesired sequences from plant genomes. Therefore, the beta/six recombination system (and presumably related recombinases) may become an attractive tool for plant genetic engineering.

Amino Acid Sequence↗
PubMed1994

Replicase-mediated resistance: a novel type of virus resistance in transgenic plants?

Genetically engineered plants expressing either intact or mutant forms of the virus-encoded replicase subunit are resistant to infection by the virus from which the transgene was obtained. In many instances, the resistance is very effective and will be useful in the field. However, some unexpected features of the resistance in these transgenic plants indicate that simple models for the mechanism of replicase-mediated resistance do not apply.

Genes, Viral↗
PubMed2003

Improving plant genetic engineering by manipulating the host.

Agrobacterium-mediated transformation is a major technique for the genetic engineering of plants. However, there are many economically important crop and tree species that remain highly recalcitrant to Agrobacterium infection. Although attempts have been made to "improve" transformation by altering the bacterium, future successes might come from manipulation of the plant. Recent studies that identified several plant genes involved in Agrobacterium-mediated transformation, and their over-expression in currently transformable species, suggest that this approach holds great promise for improving the transformation of recalcitrant, but agronomically important, crops.

DNA, Bacterial↗
PubMed1992

[Genetic engineering in plants].

Until recent years, plant genetic was involved in heredity studies through the analysis of segregations in progenies after crossing. New potentiality arose as genetic tools with the use of dissociated plant elements, transforming and cultivating them in vitro. When plants are regenerated from manipulated tissues, new structures of varieties (clones) new genotypes (transgenic plants) and new regulations of genes expression (vitrovariants) open new ways for plant genetic engineering. Progressively these technological tools are integrated in the methods of plant breeding. Yet all possible consequences of these new types of heredity and of these new genetic structures must be evaluated. As first priority the analysis of possible incidences in the field of food, nutrition and health gives the basis for diagnostics and organisations aiming to avoid the release of genotypes which could have unwanted effects.

Biotechnology↗
PubMed1992

Novel applications of the ubiquitin-dependent proteolytic pathway in plant genetic engineering.

One goal of plant genetic engineering is the manipulation of protein levels within crop plants. New insights into the ubiquitin-dependent proteolytic pathway provide potential novel ways of enhancing levels of desired proteins by synthesizing them as ubiquitin fusions, and reducing levels of undesired proteins by selective protein degradation. As a result, the ubiquitin pathway should become a useful tool for many aspects of plant biotechnology.

Amino Acid Sequence↗
PubMed2002

Plant biotechnology--genetic engineering to enhance plant salt tolerance.

Plants not only provide food to humans and animals, but also provide a large number of non-food products of industrial and chemical importance. Moreover, they have the ability to purify the air, soil and water on the earth. Various trials to genetically improve the potential of plants are actively in progress. Salt-tolerance would be an especially important ability to bestow upon plants for agricultural and industrial purposes, because high salinity conditions are ubiquitous on earth and represent major barriers to growth. Enhancement of resistance against both hyper-osmotic stress and Na+ toxity is necessary for successful molecular breeding of salt tolerant plants. Introduction of genes for osmolyte bio-synthesis is useful to increase hyperosmotic tolerance of plant cells. It is introduced in this review that genetically engineered ectoine synthesis results in increased hyperosmotic tolerance of tobacco cells. High concentrations of Na+ reduce cellular activity by interfering with vital Na+-sensitive enzymes and by affecting K+ transport. Understanding the regulation of K+ and Na+ homeostasis is thus indispensable for enhancement of plant Na+ tolerance. My research group is investigating the Na+ efflux activity of the yeast Na+-ATPase (Ena1) when installed in the plasma membrane of plant cells, and the rice K+-Na+ co-transporters (HKT) that contribute to the regulation of K+ and Na+ uptake in root cells.

Journal Article↗
PubMed2005

Engineering of plant natural product pathways.

Although many important and valuable traits are associated with plant natural products, engineering natural product pathways for plant improvement has often been limited by a lack of understanding of their biochemistry, and by the need for coordinate regulation of multiple gene activities. New approaches are facilitating both the discovery of genes that encode natural products and pathway engineering. Notable successes have been reported in altering complex pathways to improve plant quality and resistance to biotic and abiotic stresses.

Biological Factors↗
PubMed2005

[Research progress in genetic engineering of plant secondary metabolism].

Secondary metabolism plays an important role in plant life as well as the interaction between plants and environmental factors. Many secondary metabolites derived from plants have been used for the production of medicines, dyes, insecticides, food flavors, fragrances and so on. With increasingly comprehensive understanding of the plant metabolic networks, great progress has been made in the genetic improvement of plant secondary metabolic pathways through gene engineering. Strategies for the genetic engineering of plant secondary metabolism include: (1) enabling the host plant to accumulate a novel desirable compound by transformation of single/multiple enzyme gene (s) or a whole metabolic pathway; (2) decreasing target gene expression or inhibiting competitive metabolic pathway to achieve metabolic flux towards higher production of particular molecules through antisense RNA and RNA interference technologies; (3) effectively manipulating the transcription factors responsible for the metabolic regulation at multiple steps in a given pathway so as to have a great synthesis of the target bio-chemicals. Basing on author's research work on flavonoid synthesis mechanism in soybean seed and its gene engineering, recent progress in the engineering of plant secondary metabolism involved in the synthesis of anthocyanins, flavonoids, alkaloids, terpenoids, benzoic acid derivatives etc are reviewed.

Gene Expression Regulation, Plant↗