Plastid metabolism is critical in both photoautotrophic and heterotrophic plant cells. In chloroplasts, fructose-1,6-bisphosphate aldolase (FBA) catalyses the formation of both fructose 1,6-bisphosphate and sedoheptulose 1,7-bisphosphate within the Calvin-Benson cycle. Three Arabidopsis genes, AtFBA1-AtFBA3, encode plastidial isoforms of FBA, but the contributions of each isoform is unknown. Phylogenetic analysis indicates that FBA1 and FBA2 derive from a recently duplicated gene, while FBA3 is a more ancient paralog. The fba1 mutants are phenotypically indistinguishable from the wild type, while both fba2 and fba3 have reduced growth. We show that FBA2 is the major isoform in leaves, contributing most of the measurable activity. Partial redundancy with FBA1 allows both single mutants to survive, but combining both mutations is lethal, indicating a block of photoautotrophy. In contrast, FBA3 is expressed predominantly in heterotrophic tissues, especially the leaf and root vasculature, but not in the leaf mesophyll. We show that the loss of FBA3 affects plastidial glycolytic metabolism of the root, potentially limiting the biosynthesis of essential compounds such as amino acids. However, grafting experiments suggest that fba3 is dysfunctional in leaf phloem transport, and we suggest that a block in photoassimilate export from leaves causes the buildup of high carbohydrate levels and retarded growth.

The carbon isotopic composition (δ 13 C) of foliage is often used as proxy for plant performance. However, the effect of NO3vs. NH4 + supply on δ 13 C of leaf metabolites and respired CO2 is largely unknown. We supplied tobacco plants with a gradient of NO3to NH4 + concentration ratios and determined gas exchange variables, concentrations and δ 13 C of TCA cycle intermediates, δ 13 C 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 NO3decreased, activities of all measured enzymes decreased. δ 13 C of CO2 and organic acids closely co-varied and were more positive under NO3supply, suggesting organic acids as potential substrates for respiration. Together with estimates of intra-molecular 13 C enrichment in malate, we conclude that a change in the anaplerotic reaction of TCA cycle possibly contributes to 13 C enrichment in organic acids and respired CO2 under NO3supply. Thus, the effect of NO3vs. NH4 + on δ 13 C is highly relevant, particularly if δ 13 C of leaf metabolites or respiration is used as proxy for plant performance.Key words: ammonium (NH4 + ), nitrate (NO3 -), nitrate reductase (NR), phosphoenolpyruvate carboxylase (PEPC), compound specific carbon isotope analysis (CSIA), stable carbon isotopes.

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