Plastidic Phosphoenolpyruvate: Investigations on its role in plant growth and development

Phosphoenolpyruvate (PEP) plays an essential role in plant metabolism. In catabolic direction, it delivers ATP and pyruvate by the action of pyruvate kinase, which can be fed into mitochondrial respiration. PEP and pyruvate also represent essential precursors for anabolism i.e. PEP is the precursor for the synthesis of aromatic amino acids, secondary plant products and pyruvate is important for the production of fatty acids, branched-chain amino acids or isoprenoids via the mevalonate-independent way. These pathways are exclusively localized to the plastid stroma. PEP may be imported into the plastids via a PEP/phosphate translocator (PPT) of the inner envelope membrane or it may also be generated inside the stroma by complete plastid glycolysis starting from hexose phosphates. Glycolysis as the main route for PEP production involves the enzymatic sequence of 3-phosphoglycerate to PEP conversion catalyzed by phosphoglyceromutase (PGyM) and enolase (ENO). However, biochemical studies indicate that chloroplasts and a number of non-green plastids lack ENO and/or PGyM. The present study shows for the first time, the identification and functional characterization of the putative plastid localized ENO (ENO1) of Arabidopsis thaliana. The effect of a deficiency in PEP provision to plastids was investigated with the aid of double mutants defective in ENO1 and PPT1. Crosses between different alleles of eno1 and the chlorophyll a/b binding protein underexpressed (cue1) mutant defective in PPT1 could not be obtained as double homozygous lines due to partial lethality of the gametophytes and constraints in sporophyte development, including a largely reduced oil production in the seeds, thus highlighting the essential role of PEP in plastids. The heterozygous eno1 mutants in the homozygous cue1 background (cue1/eno1[+/-]) revealed an aberrant segregation pattern, retarded vegetative growth and disturbed flower development attributed to a reduced ENO1 gene dosage. Constitutive overexpression of ENO1 could rescue the reticulate leaf and the stunted root phenotype of cue1 and alleviate the diminished seed and oil production in the mutant. In contrast, wild-type plants overexpressing ENO1 alone, could not improve seed quality and oil content