Nitrate regulation of the oxidative pentose phosphate pathway in maize (Zea mays L.) root plastids: induction of 6-phosphogluconate dehydrogenase activity, protein and transcript levels

We examined the effect of nitrate on the expression of the NADPH producing enzymes of the oxidative pentose phosphate pathway, glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH) in maize seedlings (Zea mays L. W64A×A182E). In extracts of 5 day old maize roots and leaves treated with 10 mM KNO3, G6PDH and 6PGDH activities increased by 44 and 53%, respectively, relative to untreated roots. In isolated plastids from KNO3 treated roots, G6PDH and 6PGDH specific activities were more than 25and 12-fold higher than in the untreated control. Western blot analysis showed higher levels of 6PGDH protein in root plastid extracts from KNO3 treated plants. The data suggest that KNO3 specifically affects the plastidic forms of G6PDH and 6PGDH. Three classes of 6PGDH cDNA were identified in maize roots. Of these, one cDNA hybridized with a transcript that accumulated rapidly and transiently in response to low concentrations of external nitrate. The accumulation of this transcript was not affected by pretreating plants with 50 mg/ml cycloheximide, which was previously shown to inhibit cytoplasmic protein synthesis in maize roots by more than 85% (Gowri et al., Plant Mol. Biol. 26 (1998) 679). Neither NH4 + nor K treatment affected transcript accumulation. The data indicate coordinated regulation of genes and enzymes required for NO3 − assimilation and NADPH production in root plastids. © 1998 Elsevier Science Ireland Ltd. All rights reserved. Abbre6iations: 6PGDH, 6-phosphogluconate dehydrogenase; CHX, cycloheximide; DTT, dithiothreitol; Fd, ferredoxin; FNR, ferredoxin NADP oxidoreductase; G6PDH, glucose-6-phosphate dehydrogenase; GS, glutamine synthetase; GOGAT, glutamate synthase; NiR, nitrite reductase; NR, nitrate reductase; OPPP, oxidative pentose phosphate pathway. * Corresponding author. Present address: USDA, ARS, Department of Plant Pathology, OARDC, Wooster, OH 44691, USA; tel.: +1 33