A metabolomic study of alfalfa plants genetically engineered to overproduce a key enzyme in nitrogen metabolism, the cytosolic form of glutamine synthetase

In this study we have conclusively demonstrated that increasing GS₁ activity using the CaMV 35S promoter to drive the soybean GS ₁ gene (Gmglnb1) without its 3'UTR in alfalfa is accompanied by increase in GS activity and produces unique phenotypic, physiological, and metabolic features different from control plants. Differences include long internodes, early flowering and a small increase in biomass. At the physiological level, we see an increase in chlorophyll content but little increase in photosynthetic rates. The transformants showed an increase in total amino acids and protein content accounting for improved nitrogen use efficiency. There was no change in the elemental carbon content and in fact a drop in starch levels.;Analysis of the polar metabolite extract of the leaves of control and transgenic plants by GC/MS, showed a definite change in metabolite profile between the two classes of plants. The majority of carbohydrates, phosphate sugars, and organic acids detected in this study showed a decrease in concentration in the transgenic plants. All the evidence indicates that the unregulated expression of GS₁ in alfalfa resulting in increased GS₁ activity causes a reduction in carbon containing metabolites and may be the cause for limited increase in plant biomass. To elucidate altered biochemical networks in transformants, we looked at metabolite correlation matrices and their network topology.;We propose from the evidence found in this work that although GS ₁ overexpressers have a positive impact on NUE, any further improvement in NUE along with increase in biomass can be attained by co-expressing GS ₁ with a gene for a key enzyme in sucrose synthesis. One such enzyme sucrose phosphate synthase (SPS), increases the synthesis of carbon skeletons and substrates for energy production.