ME: Interagency Announcement of Opportunities in Metabolic Engineering: Metabolic Engineering of Plant Vitamin C Biosynthesis for Improved Nutrition and Health

Virginia Polytechnic Institute And State University, Blacksburg VA

Investigators

Abstract

Humans, unlike most animals, cannot synthesize vitamin C and thus have an absolute dietary requirement for this nutrient. Plants are the primary source of dietary vitamin C, which is key to several metabolic functions including collagen synthesis. Because vitamin C is a good anti-oxidant it can also be used as a natural preservative. Fresh produce with higher levels of vitamin C should therefore have improved shelf-life and consumer appeal. Additionally, increasing vitamin C in plants would result in higher intake per portion of fresh fruit or vegetables that would have a positive impact on human health. Toward this goal we will engineer the model plant Arabidopsis thaliana for higher steady state levels of vitamin C. Arabidopsis, while not part of human diet, is an essential model because it is the only plant with a fully sequenced genome. Unlike animals, the vitamin C biosynthetic metabolic network in plants is at present still not completely known. A pathway for plant systems has been proposed however, a parallel pathway, possibly similar to the animal pathway, may also operate in plants. Evidence for this pathway comes from experiments in which a cDNA encoding L-gulono-g-lactone oxidase (GLOase), the terminal enzyme in the animal pathway was expressed in tobacco and lettuce, which resulted in a 5-7 fold increase in vitamin C levels. Specific aims of the project include metabolically engineering normal and vitamin C-deficient Arabidopsis mutants to express GLOase to determine if any of the plant pathway intermediates directly supply GLOase or its precursors. Genes encoding vitamin C pathway(s) enzymes will be identified in activation tagged Arabidopsis lines by ozone screening and vitamin C analysis to fill in remaining gaps in the pathway(s). The outcomes of this project are anticipated to result in (a) Arabidopsis transformants with larger pools of vitamin C than the wild type, (b) identification of plant genes related to metabolic steps of vitamin C synthesis, and (c) further knowledge of which steps of the animal pathway also operate in plants.