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PubMed1991

Strategies for expression of foreign genes in plants. Potential use of engineered viruses.

Advances in gene transfer techniques for higher plants have already permitted important achievements towards crop protection and improvement using recombinant DNA technology. Besides plant genetic engineering, the possible use of plant viruses to express foreign genes could be of considerable interest to plant biotechnology. However, insuring containment of engineered viruses for environmental use is an important safety issue that must be addressed.

Gene Expression Regulation↗
PubMed2002

Metabolic engineering of amino acids and storage proteins in plants.

Amino acid pathways are important targets for plant metabolic engineering. Since plants represent the major global food supply, large efforts are devoted to increasing the content of "essential" amino acids, which are absolutely required in human foods and animal feeds. Engineering of amino acids is also undertaken to improve plant growth and stress tolerance. Many of the pathways of amino acid metabolism in plants have been elucidated, and genes encoding most of the enzymes are now available. The expression of recombinant genes in transgenic plants, coupled with genetic and biochemical approaches, has contributed significantly to the understanding of regulatory networks of the metabolism of amino acids and their incorporation into proteins. This knowledge is now being extensively applied to metabolic engineering of crops, and this is reflected by a large patent literature. The problems of engineering plant amino acid metabolism, and ways to solve them, are discussed using the essential amino acids lysine and methionine as examples.

Amino Acids, Essential↗
PubMed2004

Biopharmaceuticals derived from genetically modified plants.

Modern biotechnology has resulted in a resurgence of interest in the production of new therapeutic agents using botanical sources. With nearly 500 biotechnology products approved or in development globally, and with production capacity limited, the need for efficient means of therapeutic protein production is apparent. Through genetic engineering, plants can now be used to produce pharmacologically active proteins, including mammalian antibodies, blood product substitutes, vaccines, hormones, cytokines, and a variety of other therapeutic agents. Efficient biopharmaceutical production in plants involves the proper selection of host plant and gene expression system, including a decision as to whether a food crop or a non-food crop is more appropriate. Product safety issues relevant to patients, pharmaceutical workers, and the general public must be addressed, and proper regulation and regulatory oversight must be in place prior to commercial plant-based biopharmaceutical production. Plant production of pharmaceuticals holds great potential, and may become an important production system for a variety of new biopharmaceutical products.

Antibodies↗
PubMed1993

Safety assessment of the neomycin phosphotransferase II (NPTII) protein.

Two approaches were used to assess the safety of the NPTII protein for human consumption using purified E. coli produced NPTII protein that was shown to be chemically and functionally equivalent to the NPTII protein produced in genetically engineered cotton seed, potato tubers and tomato fruit. The NPTII protein was shown, as expected, to degrade rapidly under simulated mammalian digestive conditions. An acute mouse gavage study confirmed that the NPTII protein caused no deleterious effects when administered by gavage at a cumulative target dosage of up to 5000 mg/kg of body weight. This dosage correlates to at least a million fold safety factor relative to the average daily consumption of potato or tomato, assuming all the potatoes or tomatoes consumed contained the NPTII protein. These results, along with previously published information, confirm that ingestion of genetically engineered plants expressing the NPTII protein poses no safety concerns.

Animals↗
PubMed2001

Resistance to an herbivore through engineered cyanogenic glucoside synthesis.

The entire pathway for synthesis of the tyrosine-derived cyanogenic glucoside dhurrin has been transferred from Sorghum bicolor to Arabidopsis thaliana. Here, we document that genetically engineered plants are able to synthesize and store large amounts of new natural products. The presence of dhurrin in the transgenic A. thaliana plants confers resistance to the flea beetle Phyllotreta nemorum, which is a natural pest of other members of the crucifer group, demonstrating the potential utility of cyanogenic glucosides in plant defense.

Animals↗
PubMed2006

Analytical strategies to investigate plant N-glycan profiles in the context of plant-made pharmaceuticals.

Plants are attractive hosts for the production of recombinant proteins. However, their inability to process authentic human N-glycan structures imposes a major limitation on their use as expression systems for therapeutic products. Several strategies have emerged to engineer plant N-glycans into human-compatible molecules. In this context, fast and reliable analytical strategies for the identification of plant N-glycan profiles have been developed to define the N-glycosylation pathways of crops, to monitor the production of plant-made pharmaceuticals and to assess in planta remodelling strategies.

Amino Acid Sequence↗
PubMed1994

Developmentally regulated cell death on expression of the fungal avirulence gene Avr9 in tomato seedlings carrying the disease-resistance gene Cf-9.

Plant defense responses are induced when the products of disease-resistance genes and pathogen avirulence genes interact. We report here the effects of expressing the Cladosporium fulvum avirulence Avr9 gene product in a tomato line containing the Cf-9 disease-resistance gene. A synthetic Avr9 gene was constructed to produce constitutive high-level expression of AVR9 peptide in the plant apoplast. Avr9 expression in Cf-9-containing tomato lines is lethal, but cell death is developmentally regulated, in that necrosis is not visible until 10 days after planting seed. Plant lines lacking Cf-9 and expressing Avr9 remain healthy. The synthetic Avr9 gene exhibited the same specificity of action as the authentic C. fulvum Avr9 gene. Our results have significant implications for strategies using the gene combination Avr9/Cf-9 to engineer plants with enhanced disease resistance.

Amino Acid Sequence↗
PubMed1996

Novel approaches to vaccine development against HBV.

This review summarizes studies from my laboratory based on two novel approaches towards vaccine development against hepatitis B surface antigen. The first approach deals with the ability of a monoclonal anti-idiotypic antibody to functionally mimic the a determinant of HBsAg. The second approach exploits the strategy of genetically engineering plants so that they can be used as inexpensive alternatives to fermentation systems for the production of candidate vaccine antigens.

Amino Acid Sequence↗
PubMed2000

Integration of advanced process control and full-scale dynamic simulation

In the process control industry, multivariable model predictive controller and dynamic simulation for operator training are usually available in separate packages. It is very difficult for the operators and plant engineers to find good tools for them to get trained in multivariable advanced process control. This paper presents a system, which integrates the advanced process control and full-scale dynamic simulation. The advanced process control uses multivariable model predictive control techniques. The model used in the predictive control algorithms is generated from the dynamic simulated process. The advanced process controller can control the simulated plant directly, or through a DCS system to control the simulated plant. The combined system provides an excellent environment for training operators in process operation with multivariable advanced process control. The same environment is also very useful for engineers in designing and tuning the advanced process controllers, and in testing communication between the advanced process controller and the DCS systems, or the other type of process control systems.

Journal Article↗
PubMed2002

New Insights into the Functions of Cytokinins in Plant Development.

Recent breakthroughs in cytokinin research have shed new light on the role of cytokinin in plant development. Loss-of-function mutants of a cytokinin receptor reveal a role for the hormone in establishment of the vasculature during embryonic development. Cytokinin controls the number of early cell divisions via a two-component signaling system. Genetically engineered plants that have a reduced cytokinin content demonstrate the regulatory role of the hormone in control of meristem activity and organ growth during postembryonic development, with opposite roles in roots and shoots. There is increasing evidence from work with transgenic plants and mutant analysis that cytokinins do not perform the previously proposed function as a root-derived signal for the regulation of shoot branching. Root-borne cytokinins might serve as a long-range signal controling other processes at distant sites, such as responding to nutritional status, particularly nitrogen availability.

Journal Article↗
PubMed2000

Enhanced phosphorus uptake in transgenic tobacco plants that overproduce citrate.

Phosphorus (P) is one of the most important nutrients limiting agricultural production worldwide. In acid and alkaline soils, which make up over 70% of the world's arable land, P forms insoluble compounds that are not available for plant use. To reduce P deficiencies and ensure plant productivity, nearly 30 million tons of P fertilizer are applied every year. Up to 80% of the applied P fertilizer is lost because it becomes immobile and unavailable for plant uptake. Therefore, the development of novel plant varieties more efficient in the use of P represents the best alternative to reduce the use of P fertilizers and achieve a more sustainable agriculture. We show here that the ability to use insoluble P compounds can be significantly enhanced by engineering plants to produce more organic acids. Our results show that when compared to the controls, citrate-overproducing plants yield more leaf and fruit biomass when grown under P-limiting conditions and require less P fertilizer to achieve optimal growth.

Biological Transport↗
PubMed2001

Plant concepts for mineral acquisition and allocation.

Plant biotechnology is expected to make a major contribution to the steady increase of crop production in the near future. The improvement of mineral assimilation has to meet the challenges of reducing fertilizer application in developed countries, preserving the environment, enabling sustainable agriculture management and generating low-input crops with increased performance in areas where soil infertility limits productivity. Natural genetic resources and engineered plants will help to achieve the implementation of traits for improved mineral assimilation.

Crops, Agricultural↗
PubMed2006

Genetic modulation of ethylene biosynthesis and signaling in plants.

With the isolation and characterization of the key enzymes and proteins, and the corresponding genes, involved in ethylene biosynthesis and sensing it has become possible to manipulate plant ethylene levels and thereby alter a wide range of physiological processes. The phytohormone ethylene is an essential signaling molecule that affects a large number of physiological processes; plants deprived of ethylene do not grow and develop normally. In a search for flexible on-off ethylene control, scientists have used inducible organ- and tissue-specific promoters to drive expression of different transgenes. Here, the various strategies that have been used to genetically engineer plants with decreased ethylene biosynthesis and sensitivity are reviewed and discussed.

Ethylenes↗
PubMed2001

Over-expression of a cDNA for human ornithine decarboxylase in transgenic rice plants alters the polyamine pool in a tissue-specific manner.

We investigated how over-expression of a cDNA for human ornithine decarboxylase (odc) affects the polyamine pools in transgenic rice. We further investigated tissue-specific expression patterns and product accumulation levels of the transgene driven by either constitutive or seed-specific promoters. Our results indicate that: (1) whereas the expression of a heterologous arginine decarboxylase (adc) cDNA in rice resulted in increased putrescine and spermine levels only in seeds, plants engineered to express odc cDNA exhibited significant changes in the levels of all three major polyamines in seeds and also in vegetative tissues (leaves and roots); (2) there was no linear correlation between odc mRNA levels, ODC enzyme activity and polyamine accumulation, suggesting that control of the polyamine pathway in plants is more complex than in mammalian systems; (3) ODC activity and polyamine changes varied in different tissues, indicating that the pathway is regulated in a tissue-specific manner. Our results suggest that ODC rather than ADC is responsible for the regulation of putrescine synthesis in plants.

Carboxy-Lyases↗
PubMed2004

Recent advances in chlorophyll biosynthesis and breakdown in higher plants.

Chlorophyll (Chl) has unique and essential roles in photosynthetic light-harvesting and energy transduction, but its biosynthesis, accumulation and degradation is also associated with chloroplast development, photomorphogenesis and chloroplast-nuclear signaling. Biochemical analyses of the enzymatic steps paved the way to the identification of their encoding genes. Thus, important progress has been made in the recent elucidation of almost all genes involved in Chl biosynthesis and breakdown. In addition, analysis of mutants mainly in Arabidopsis , genetically engineered plants and the application of photo-reactive herbicides contributed to the genetic and regulatory characterization of the formation and breakdown of Chl. This review highlights recent progress in Chl metabolism indicating highly regulated pathways from the synthesis of precursors to Chl and its degradation to intermediates, which are not longer photochemically active.

Arabidopsis↗
PubMed1988

Analysis of the mechanism of protection in transgenic plants expressing the potato virus X coat protein or its antisense RNA.

Transgenic tobacco plants engineered to express either the potato virus X (PVX) coat protein (CP+) or the antisense coat protein transcript (CP-antisense) were protected from infection by PVX, as indicated by reduced lesion numbers on inoculated leaves, delay or absence of systemic symptom development and reduction in virus accumulation in both inoculated and systemic leaves. The extent of protection observed in CP+ plants primarily depended upon the level of expression of the coat protein. Plants expressing antisense RNA were protected only at low inoculum concentrations. The extent of this protection was even lower than that observed in plants expressing low levels of CP. In contrast to previous reports for plants expressing tobacco mosaic virus or alfalfa mosaic virus CP, inoculation of plants expressing high levels of PVX CP with PVX RNA did not overcome the protection. Specifically, lesion numbers on inoculated leaves and PVX levels on inoculated and systemtic leaves of the CP+ plants were reduced to a similar extent in both virus and RNA inoculated plants. Although these results do not rule out that CP-mediated protection involves inhibition of uncoating of the challenge virus, they suggest that PVX CP (or its RNA) can moderate early events in RNA infection by a different mechanism.

Journal Article↗
PubMed2001

The role of chlorophyll b in photosynthesis: hypothesis.

BACKGROUND: The physico-chemical properties of chlorophylls b and c have been known for decades. Yet the mechanisms by which these secondary chlorophylls support assembly and accumulation of light-harvesting complexes in vivo have not been resolved. PRESENTATION: Biosynthetic modifications that introduce electronegative groups on the periphery of the chlorophyll molecule withdraw electrons from the pyrrole nitrogens and thus reduce their basicity. Consequently, the tendency of the central Mg to form coordination bonds with electron pairs in exogenous ligands, a reflection of its Lewis acid properties, is increased. Our hypothesis states that the stronger coordination bonds between the Mg atom in chlorophyll b and chlorophyll c and amino acid sidechain ligands in chlorophyll a/b- and a/c-binding apoproteins, respectively, enhance their import into the chloroplast and assembly of light-harvesting complexes. TESTING: Several apoproteins of light-harvesting complexes, in particular, the major protein Lhcb1, are not detectable in leaves of chlorophyll b-less plants. A direct test of the hypothesis--with positive selection--is expression, in mutant plants that synthesize only chlorophyll a, of forms of Lhcb1 in which weak ligands are replaced with stronger Lewis bases. IMPLICATIONS: The mechanistic explanation for the effects of deficiencies in chlorophyll b or c points to the need for further research on manipulation of coordination bonds between these chlorophylls and chlorophyll-binding proteins. Understanding these interactions will possibly lead to engineering plants to expand their light-harvesting antenna and ultimately their productivity.

Amino Acid Sequence↗