EFFECT OF LIGHT AND NO3- ON WHEAT LEAF PHOSPHOENOLPYRUVATE CARBOXYLASE ACTIVITY - EVIDENCE FOR COVALENT MODULATION OF THE C3 ENZYME

Phosphoenolpyruvate carboxylase (PEPcase) activity was studied in excised leaves of wheat (Triticum aestivum L.) in the dark and in the light, in presence of either N-free (low-NO3- leaves) or 40 millimolar KNO3 (high-NO3- leaves) nutrient solutions. PEPcase activity increased to 2.7-fold higher than that measured in dark-adapted tissue (control) during the first 60 minutes and continued to increase more slowly to 3.8-fold that of the control. This level was reached after 200 minutes exposure of the leaves to light and high NO3-. In contrast, the lower rate of increase recorded for low-NO3- leaves ceased after 60 minutes of exposure to light at 2.3-fold the control level. The short-term NO3- effect increased linearly with the level of NO3- uptake. In immunoprecipitation experiments, the antibody concentration for PEPcase precipitation increased with the protein extracts from the different treatments in the order: control, illuminated low-NO3- leaves, illuminated high-NO3- leaves. This order also applied with regard to a decreasing sensitivity to malate and an increasing stimulation by okadaic acid (an inhibitor of P-protein phosphatases). Following these studies P-32 labeling experiments were carried out in vivo. These showed that the light-induced change in the properties of the PEPcase was due to an alteration in the phosphorylation state of the protein and that this effect was enhanced in high-NO3- conditions. Based on the responses of PEPcase and sucrose phosphate synthase in wheat leaves to light and NO3-, an interpretation of the role of NO3- as either an inhibitor of P-protein phosphatase(s) or activator of protein kinase(s) is inferred. In the presence of NO3-, the phosphorylation state of both PEPcase and sucrose phosphate synthase is increased. This causes activation of the former enzyme and inhibition of the latter. We suggest that NO3- modulates the relative protein kinase/protein phosphatase ratio to favor increased phosphorylation of both enzymes in order to redirect carbon flow away from sucrose synthesis and toward amino acid synthesis.