Nitrate and ammonium differ in their impact on δ13C of plant metabolites and respired CO2 from tobacco leaves.

The carbon isotopic composition (δ13C) of foliage is often used as proxy for plant performance. However, the effect of N O 3 - vs. N H 4 + supply on δ13C of leaf metabolites and respired CO2 is largely unknown. We supplied tobacco plants with a gradient of N O 3 - to N H 4 + concentration ratios and determined gas exchange variables, concentrations and δ13C of tricarboxylic acid (TCA) cycle intermediates, δ13C of dark-respired CO2, and activities of key enzymes nitrate reductase, malic enzyme and phosphoenolpyruvate carboxylase. Net assimilation rate, dry biomass and concentrations of organic acids and starch decreased along the gradient. In contrast, respiration rates, concentrations of intercellular CO2, soluble sugars and amino acids increased. As N O 3 - decreased, activities of all measured enzymes decreased. δ13C of CO2 and organic acids closely co-varied and were more positive under N O 3 - supply, suggesting organic acids as potential substrates for respiration. Together with estimates of intra-molecular 13C enrichment in malate, we conclude that a change in the anaplerotic reaction of the TCA cycle possibly contributes to 13C enrichment in organic acids and respired CO2 under N O 3 - supply. Thus, the effect of N O 3 - vs. N H 4 + on δ13C is highly relevant, particularly if δ13C of leaf metabolites or respiration is used as proxy for plant performance.

Increasing the carbohydrate storage capacity of plants by engineering a glycogen-like polymer pool in the cytosol.

Global demand for higher crop yields and for more efficient utilization of agricultural products will grow over the next decades. Here, we present a new concept for boosting the carbohydrate content of plants, by channeling photosynthetically fixed carbon into a newly engineered glucose polymer pool. We transiently expressed the starch/glycogen synthases from either Saccharomyces cerevisiae or Cyanidioschyzon merolae, together with the starch branching enzyme from C. merolae, in the cytosol of Nicotiana benthamiana leaves. This effectively built a UDP-glucose-dependent glycogen biosynthesis pathway. Glycogen synthesis was observed with Transmission Electron Microscopy, and the polymer structure was further analyzed. Within three days of enzyme expression, glycogen content of the leaf was 5-10 times higher than the starch levels of the control. Further, the leaves produced less starch and sucrose, which are normally the carbohydrate end-products of photosynthesis. We conclude that after enzyme expression, the newly fixed carbohydrates were routed into the new glycogen sink and trapped. Our approach allows carbohydrates to be efficiently stored in a new subcellular compartment, thus increasing the value of vegetative crop tissues for biofuel production or animal feed. The method also opens new potential for increasing the sink strength of heterotrophic tissues.

Loss of starch granule initiation has a deleterious effect on the growth of arabidopsis plants due to an accumulation of ADP-glucose.

STARCH SYNTHASE4 (SS4) is required for proper starch granule initiation in Arabidopsis (Arabidopsis thaliana), although SS3 can partially replace its function. Unlike other starch-deficient mutants, ss4 and ss3/ss4 mutants grow poorly even under long-day conditions. They have less chlorophyll and carotenoids than the wild type and lower maximal rates of photosynthesis. There is evidence of photooxidative damage of the photosynthetic apparatus in the mutants from chlorophyll a fluorescence parameters and their high levels of malondialdehyde. Metabolite profiling revealed that ss3/ss4 accumulates over 170 times more ADP-glucose (Glc) than wild-type plants. Restricting ADP-Glc synthesis, by introducing mutations in the plastidial phosphoglucomutase (pgm1) or the small subunit of ADP-Glc pyrophosphorylase (aps1), largely restored photosynthetic capacity and growth in pgm1/ss3/ss4 and aps1/ss3/ss4 triple mutants. It is proposed that the accumulation of ADP-Glc in the ss3/ss4 mutant sequesters a large part of the plastidial pools of adenine nucleotides, which limits photophosphorylation, leading to photooxidative stress, causing the chlorotic and stunted growth phenotypes of the plants.