Gluconeogenesis and nitrogen metabolism in maize.

Two pathways can be used by gluconeogenesis in plants: one employs phosphoenolpyruvate carboxykinase (PEPCK) and the other pyruvate orthophosphate dikinase (PPDK). The occurrence-location of these enzymes was determined in developing kernels of maize. PPDK was much more abundant than PEPCK in extracts of whole kernels. However, their location within the kernel was different. PPDK was particularly abundant in the peripheral endosperm (in which alanine is abundant), whereas PEPCK was localised in the pedicel and basal endosperm transfer cells (where asparagine is metabolised). The abundance of these enzymes was also determined in maize roots where there was a massive increase in abundance of PEPCK and a small increase in abundance of PPDK when they were fed ammonium; PEPCK was located in the pericycle and various cell types associated with the vasculature. On the other hand, there was a large increase in abundance of PPDK in roots subjected to anoxia (which induces an accumulation of alanine), whereas the abundance of PEPCK was decreased. These results show: firstly, that gluconeogenesis can potentially occur in many different tissues of maize. Secondly, within one organ PPDK can be abundant in some tissues and PEPCK in others. Thirdly, the abundance of PPDK and PEPCK is often associated with the metabolism of certain nitrogenous compounds and can be dramatically altered by factors related to nitrogen metabolism. In maize roots and developing kernels PPDK was associated with alanine metabolism. By contrast, the presence of PEPCK in maize roots and kernels was associated with either ammonium or asparagine metabolism. We propose that gluconeogenesis is often a component of a widespread mechanism that is used in coordinating the import/mobilisation of nitrogenous compounds with their utilisation. Further, potentially component of this mechanism may have provided building blocks that were used in the evolution of processes such as C4 photosynthesis, Crassulacean acid metabolism, stomatal metabolism and the biochemical pH stat.

Phosphoenolpyruvate carboxykinase and gluconeogenesis in grape pericarp.

Glycolysis from sugars is necessary at all stages of development of grape pericarp, and this raises the question as to why gluconeogenesis from malate occurs. Phosphoenolpyruvate carboxykinase (PEPCK) is required for gluconeogenesis in grape pericarp. In this study we determined the abundance of PEPCK protein and activity in different parts of grape pericarp during its development. Both PEPCK protein and activity were present throughout development, however, in both the skin and the flesh their abundance increased greatly at the start of ripening. This coincided with the onset of the decrease in the malate content of the berry. The location of PEPCK in the pericarp at different stages of development was determined using both immunohistochemistry and dissection. We provide a possible explanation for the occurrence of gluconeogenesis in grape pericarp.

Phosphoenolpyruvate carboxykinase in cucumber plants is increased both by ammonium and by acidification, and is present in the phloem.

In cucumber ( Cucumis sativus L.), phosphoenolpyruvate carboxykinase (PEPCK) was shown by activity measurements and immunoblots to be present in leaves, stems, roots, flowers, fruit and seed. However, immunolocalisation showed that it was present only in certain cell types. PEPCK was present in the companion cells of the adaxial phloem of minor veins, the adaxial and abaxial phloem of larger veins, the internal and external phloem of vascular bundles in petioles and stems, the phloem in roots and the extra-fascicular phloem in leaves, cotyledons, petioles and stems. Immunohistochemical evidence suggests that both the extra-fascicular phloem and the adaxial phloem are involved in the transport of amino acids. In roots and stems, the abundance of PEPCK was greatly increased by watering plants with a solution of ammonium chloride at low, but not at high pH. PEPCK also increased in leaves, but not roots or stems, of seedlings grown in an atmosphere containing 5% CO(2), and in roots and stems of seedlings watered with butyric acid. All these treatments are known to lower the pH of plant cells. Amino acid metabolism in the phloem may produce an excess of carbon skeletons, pH perturbations and an imbalance in the production/utilisation of NADH. This raises the possibility that PEPCK may function in the conversion of these carbon skeletons to PEP, which, depending on the energy requirements of the phloem, is subsequently utilised by either gluconeogenesis or the Krebs cycle, which both consume protons.