Gemma Farré

Transgenic multivitamin corn through biofortification of endosperm with three vitamins representing three distinct metabolic pathways

Vitamin deficiency affects up to 50% of the worlds population, disproportionately impacting on developing countries where populations endure monotonous, cereal-rich diets. Transgenic plants offer an effective way to increase the vitamin content of staple crops, but thus far it has only been possible to enhance individual vitamins. We created elite inbred South African transgenic corn plants in which the levels of 3 vitamins were increased specifically in the endosperm through the simultaneous modification of 3 separate metabolic pathways. The transgenic kernels contained 169-fold the normal amount of β-carotene, 6-fold the normal amount of ascorbate, and double the normal amount of folate. Levels of engineered vitamins remained stable at least through to the T3 homozygous generation. This achievement, which vastly exceeds any realized thus far by conventional breeding alone, opens the way for the development of nutritionally complete cereals to benefit the worlds poorest people.

When more is better: multigene engineering in plants

The genomics revolution has taught us that a great deal of information can be derived from studying many genes or proteins at the same time. We are beginning to see this approach blossoming in applied research. Instead of attempting to generate useful transgenic plants by introducing single genes, we now see an increasing number of researchers embracing multigene transfer (MGT) as an approach to generate plants with more ambitious phenotypes. MGT allows researchers to achieve goals that were once impossible - the import of entire metabolic pathways, the expression of entire protein complexes, the development of transgenic crops simultaneously engineered to produce a spectrum of added-value compounds. The potential appears limitless.

The regulation of carotenoid pigmentation in flowers

Carotenoids fulfill many processes that are essential for normal growth and development in plants, but they are also responsible for the breathtaking variety of red-to-yellow colors we see in flowers and fruits. Although such visual diversity helps to attract pollinators and encourages herbivores to distribute seeds, humans also benefit from the aesthetic properties of flowers and an entire floriculture industry has developed on the basis that new and attractive varieties can be produced. Over the last decade, much has been learned about the impact of carotenoid metabolism on flower color development and the molecular basis of flower color. A number of different regulatory mechanisms have been described ranging from the transcriptional regulation of genes involved in carotenoid synthesis to the control of carotenoid storage in sink organs. This means we can now explain many of the natural colorful varieties we see around us and also engineer plants to produce flowers with novel and exciting varieties that are not provided by nature.

Promoter diversity in multigene transformation

Multigene transformation (MGT) is becoming routine in plant biotechnology as researchers seek to generate more complex and ambitious phenotypes in transgenic plants. Every nuclear transgene requires its own promoter, so when coordinated expression is required, the introduction of multiple genes leads inevitably to two opposing strategies: different promoters may be used for each transgene, or the same promoter may be used over and over again. In the former case, there may be a shortage of different promoters with matching activities, but repetitious promoter use may in some cases have a negative impact on transgene stability and expression. Using illustrative case studies, we discuss promoter deployment strategies in transgenic plants that increase the likelihood of successful and stable multiple transgene expression.

Nutritious crops producing multiple carotenoids – a metabolic balancing act

Plants and microbes produce multiple carotenoid pigments with important nutritional roles in animals. By unraveling the basis of carotenoid biosynthesis it has become possible to modulate the key metabolic steps in plants and thus increase the nutritional value of staple crops, such as rice (Oryza sativa), maize (Zea mays) and potato (Solanum tuberosum). Multigene engineering has been used to modify three different metabolic pathways simultaneously, producing maize seeds with higher levels of carotenoids, folate and ascorbate. This strategy may allow the development of nutritionally enhanced staples providing adequate amounts of several unrelated nutrients. By focusing on different steps in the carotenoid biosynthesis pathway, it is also possible to generate plants with enhanced levels of several nutritionally-beneficial carotenoid molecules simultaneously.

A golden era—pro-vitamin A enhancement in diverse crops

Numerous crops have been bred or engineered to increase carotenoid levels in an effort to develop novel strategies that address vitamin A deficiency in the developing world. The pioneering work in rice (not covered in this review) has been followed up in many additional crops, some of which are staples like rice whereas others are luxury products whose impact on food security is likely to be marginal. This review surveys the progress that has been made in carotenoid breeding and metabolic engineering, focusing on β-carotene enhancement in crops other than rice. We ask if these efforts have the potential to address vitamin A deficiency in developing countries by comparing bioavailable pro-vitamin A levels in wild type and enhanced crops to determine whether nutritional requirements can be met without the consumption of unrealistic amounts of food. The potential impact of carotenoid enhancement should therefore be judged against benchmarks that include the importance of particular crops in terms of global food security, the amount of bioavailable β-carotene, and the amount of food that must be consumed to achieve the reference daily intake of vitamin A.

Biofortification of plants with altered antioxidant content and composition: genetic engineering strategies

Antioxidants are protective molecules that neutralize reactive oxygen species and prevent oxidative damage to cellular components such as membranes, proteins and nucleic acids, therefore reducing the rate of cell death and hence the effects of ageing and ageing-related diseases. The fortification of food with antioxidants represents an overlap between two diverse environments, namely fortification of staple foods with essential nutrients that happen to have antioxidant properties (e.g. vitamins C and E) and the fortification of luxury foods with health-promoting but non-essential antioxidants such as flavonoids as part of the nutraceuticals/functional foods industry. Although processed foods can be artificially fortified with vitamins, minerals and nutraceuticals, a more sustainable approach is to introduce the traits for such health-promoting compounds at source, an approach known as biofortification. Regardless of the target compound, the same challenges arise when considering the biofortification of plants with antioxidants, that is the need to modulate endogenous metabolic pathways to increase the production of specific antioxidants without affecting plant growth and development and without collateral effects on other metabolic pathways. These challenges become even more intricate as we move from the engineering of individual pathways to several pathways simultaneously. In this review, we consider the state of the art in antioxidant biofortification and discuss the challenges that remain to be overcome in the development of nutritionally complete and health-promoting functional foods.

Standards for plant synthetic biology: a common syntax for exchange of DNA parts

Inventors in the field of mechanical and electronic engineering can access multitudes of components and, thanks to standardization, parts from different manufacturers can be used in combination with each other. The introduction of BioBrick standards for the assembly of characterized DNA sequences was a landmark in microbial engineering, shaping the field of synthetic biology. Here, we describe a standard for Type IIS restriction endonuclease-mediated assembly, defining a common syntax of 12 fusion sites to enable the facile assembly of eukaryotic transcriptional units. This standard has been developed and agreed by representatives and leaders of the international plant science and synthetic biology communities, including inventors, developers and adopters of Type IIS cloning methods. Our vision is of an extensive catalogue of standardized, characterized DNA parts that will accelerate plant bioengineering.

Engineering Complex Metabolic Pathways in Plants

Metabolic engineering can be used to modulate endogenous metabolic pathways in plants or introduce new metabolic capabilities in order to increase the production of a desirable compound or reduce the accumulation of an undesirable one. In practice, there are several major challenges that need to be overcome, such as gaining enough knowledge about the endogenous pathways to understand the best intervention points, identifying and sourcing the most suitable metabolic genes, expressing those genes in such a way as to produce a functional enzyme in a heterologous background, and, finally, achieving the accumulation of target compounds without harming the host plant. This article discusses the strategies that have been developed to engineer complex metabolic pathways in plants, focusing on recent technological developments that allow the most significant bottlenecks to be overcome.

Nutritionally enhanced crops and food security : scientific achievements versus political expediency

Genetic engineering (GE) is one of a raft of strategies that can be used to tackle malnutrition. Recent scientific advances have shown that multiple deficiencies can be tackled simultaneously using engineered plant varieties containing high levels of different minerals and organic nutrients. However, the impact of this progress is being diluted by the unwillingness of politicians to see beyond immediate popular support, favoring political expediency over controversial but potentially life-saving decisions based on rational scientific evidence.