Drought-induced effects on nitrate reductase activity and mRNA and on the coordination of nitrogen and carbon metabolism in maize leaves

Abstract

Maize (Zea mays L.) plants were grown to the nine-leaf stage. Despite a saturating N supply, the youngest mature leaves (seventh position on the stem) contained little NO3- reserve. Droughted plants (deprived of nutrient solution) showed changes in foliar enzyme activities, mRNA accumulation, photosynthesis, and carbohydrate and amino acid contents. Total leaf water potential and CO2 assimilation rates, measured 3 h into the photoperiod, decreased 3 d after the onset of drought. Starch, glucose, fructose, and amino acids, but not sucrose (Suc), accumulated in the leaves of droughted plants. Maximal extractable phosphoenolpyruvate carboxylase activities increased slightly during water deficit, whereas the sensitivity of this enzyme to the inhibitor malate decreased. Maximal extractable Suc phosphate synthase activities decreased as a result of water stress, and there was an increase in the sensitivity to the inhibitor orthophosphate. A correlation between maximal extractable foliar nitrate reductase (NR) activity and the rate of CO2 assimilation was observed. The NR activation state and maximal extractable NR activity declined rapidly in response to drought. Photosynthesis and NR activity recovered rapidly when nutrient solution was restored at this point. The decrease in maximal extractable NR activity was accompanied by a decrease in NR transcripts, whereas Suc phosphate synthase and phosphoenolpyruvate carboxylase mRNAs were much less affected. The coordination of N and C metabolism is retained during drought conditions via modulation of the activities of Suc phosphate synthase and NR commensurate with the prevailing rate of photosynthesis.

Figure 1

Total leaf water potential of maize plants continuously irrigated (▪) or subjected to 8 d of water deprivation (▴). The water potential of detached leaves was measured with a Scholander pressure chamber. Each given value represents the mean of three replicates. se is indicated for each value.

Figure 2

The rate of net photosynthetic CO₂ assimilation (μmol m⁻² s⁻¹) by leaves of maize plants continuously irrigated (▪), subjected to 8 d of water deprivation (▴), or deprived of water for 3 d and then rewatered to soil capacity (○). Each value represents the mean of three replicate experiments. se is indicated for each value.

Figure 3

The maximal extractable activities of NR (I), PEPCase (II), or SPS (III) of leaves of maize plants continuously irrigated (shaded bars) or exposed to 1 week of water deprivation (black bars). Total extractable NR activity (μmol mg⁻¹ chlorophyll [Chl] h⁻¹) was assayed in the presence of 5 mm EDTA (A) or in the presence of 10 mm Mg²⁺, allowing calculation of the activation state (D). Total extractable PEPCase activities (mmol mg⁻¹ Chl h⁻¹) were measured in the absence of malate (B). In E, each given value represents the ratio of total extractable PEPCase activity and the respective PEPCase activity assayed in the presence of 3 mm malate. Maximal extractable SPS activities (μmol mg⁻¹ Chl h⁻¹) are given in C, and F shows the ratio of maximal catalytic SPS activity to that measured under conditions of limiting substrates in the presence of Pi. Each value represents the mean of three replicate experiments. se is indicated for each value.

Figure 4

Maximal extractable NR activities in leaves from droughted plants (▴), water-replete controls (▪), and plants rewatered 3 d after the onset of water deprivation (○). Chl, Chlorophyll. Results are means ± se.

Figure 5

The relationship between the CO₂ assimilation rate in air in the growth conditions and maximal extractable NR activity in the same leaves. Samples were taken from individual water-replete plants (▪) or plants deprived of water for 7 d (♦). Each value represents the mean of three replicate experiments. Chl, Chlorophyll.

Figure 6

The NR (A), PEPCase (B), or SPS (C) transcript levels in leaves of water-replete controls (WR) or water-deprived plants (WD) at 3 d (1), 5 d (2), and 7 d (3) after withholding irrigation or 24 h after irrigation to soil capacity of maize plants that had been deprived of water for 7 d (RW). Denatured total RNA (10 μg) samples were fractionated on a formaldehyde-agarose gel (1.5% agarose) and transferred onto a membrane. The RNA blots were probed with the partial 2.2-kb cDNA clone pZmnrl encoding NR of maize (Gowrie and Campbell, 1989), a partial 1.0-kb cDNA clone corresponding to PEPCase of maize (Sheen and Bogorad, 1987), or a full-length 3.4-kb cDNA clone encoding SPS of maize (Worrell et al., 1991).

Figure 7

The contribution of major amino acids to the total amino acid pool in droughted maize leaves (A) and in water-replete controls (B). Measurements were made 7 d after the onset of water deprivation.