This Small Business Technology Transfer (STTR) Phase I project proposes to improve intrinsic crop yield through metabolic engineering of photosynthetic pathways. Genes encoding five rate-limiting enzymes for photosynthesis and starch synthesis will be co-expressed constitutively or in a cell-specific manner in a model C4 plant species. Multiple metabolic pathways will be altered to simultaneously relieve multiple rate-limiting steps for carbon assimilation. The effects of enzyme accumulation on photosynthetic performance, plant growth, and biomass accumulation will be assayed, and optimal combinations of enzymes and relative enzyme levels will be determined. The effects of constitutive enzyme accumulation will be compared with cell-specific enzyme accumulation to determine whether a targeted expression profile can deliver a more substantial improvement in carbon assimilation rates and plant growth than constitutive enzyme accumulation. The results of this work will identify key rate-limiting enzymes in the photosynthetic machinery, optimal enzyme expression profiles, and optimal enzyme concentrations to improve photosynthetic performance, carbon assimilation, and yield.
The broader impact/commercial potential of this project, if successful, will be to identify novel ways to improve crop yields in a range of both food and non-food crops. Photosynthetic engineering for improved carbon assimilation is a promising, but underexplored, method for improving intrinsic crop yield. Traditional plant biotechnology approaches have sought to protect yield, e.g. through insect resistance and herbicide tolerance. Utilizing synthetic biology to engineer primary metabolism and increase intrinsic crop yields will work in tandem with existing yield-protecting technologies. Modeling and initial proof-of-concept studies have confirmed that photosynthetic pathways can be engineered for greater efficiency, resulting in yield improvements. The results of this work will significantly enhance our understanding of the rate-limiting steps of photosynthetic carbon assimilation, providing insight into the most promising reactions and metabolic pathways for these engineering approaches. Translating the results of this Phase I work to crop plants will result in improved crop harvests without expanding the agricultural footprint, translating to enormous commercial benefits to the agricultural, food, and energy sectors.
Facing increasing demand for food and feed as a result of the growing global population, rising incomes and standards of living, and agriculture-intensive biofuels policies, a primary goal for agriculture is the generation of higher-yielding crop varieties that can increase production without expanding agriculture’s footprint. While breeding programs have produced significant increases in crop yield, the rate of yield increase is not sufficient to meet the growing demand for food and feed in a number of crops. Novel approaches are therefore required in order to supply food and feed for the world’s growing population. An approach that has gained support among the scientific community in recent years is the use of rational engineering approaches to optimize photosynthetic carbon metabolism. By improving on photosynthetic carbon metabolism, carbon dioxide capture by plants will be accelerated in order to provide additional carbon for their growth. Of particular importance is the C4 photosynthetic pathway that is used by many high-yielding crop plants including corn (maize), sugarcane, and sorghum. While this photosynthetic pathway can provide advantages relative to C3 photosynthesis in many environments, optimization of the C4 photosynthetic pathway has the potential to greatly increase carbon assimilation to improve crop sustainability, growth, and ultimately yield. Manipulation of the C4 photosynthetic pathway requires complex metabolic engineering approaches that have rarely been used in higher plants. This project used plant transformation constructs engineered to provide overexpression of five genes encoding rate-limiting enzymes in the metabolic pathways of photosynthesis. These genes were expressed from different promoters to provide expression of these genes in all of the plants’ cells or only in those cells in which transgene expression was predicted to be beneficial. Setaria viridis, an emerging model grass species that shares many genes and physiological similarities with C4 crop plants, was transformed with the constructs that were built for this work. The resulting transgenic lines were characterized at the molecular, biochemical, and phenotypic levels in order to determine the effects of transgene expression on plant growth. A number of unexpected phenotypes not predicted by existing models of C4 photosynthesis were uncovered, underscoring the need for more experiments to understand the effects of altering the levels of photosynthetic enzymes on C4 plant growth and biochemistry. The results of this work will have implications for basic as well as applied research programs focused on understanding and improving on photosynthetic carbon metabolism.
