Sucrose phosphate synthase (SPS) is a key enzyme in sucrose synthesis, which controls sucrose content in plants. This study was designed to examine the efficacy of the overexpression of SoSPS1 gene on sucrose accumulation and carbon partitioning in transgenic sugarcane. The overexpression of SoSPS1 gene increased SPS activity and sucrose content in transgenic sugarcane leaves. More importantly, the overexpression enhanced soluble acid invertase (SAI) activity concomitant with the increase of glucose and fructose levels in the leaves, whereas sucrose synthase activity exhibited almost no change. In the stalk, a similar correlation was observed, but a higher correlation was noted between SPS activity and sugar content. These results suggest that SPS overexpression has both direct and indirect effects on sugar concentration and SAI activity in sugarcane. In addition, SPS overexpression resulted in a significant increase in plant height and stalk number in some transgenic lines compared to those in non-transgenic control. Taken together, these results strongly suggest that enhancing SPS activity is a useful strategy for improving sugarcane yield.

We previously showed that the selective accumulation of phosphoenolpyruvate carboxylase (PEPC) in photosynthetically maturing maize (Zea mays L.) leaf cells induced by nitrate supply to nitrogen-starved plants was primarily a consequence of the level of its mRNA (B Sugiharto, K Miyata, H Nakamoto, H Sasakawa, T Sugiyama [1990] Plant Physiol 92: 963-969). To determine the specificity of inorganic nitrogen sources for the regulation of PEPC gene expression, nitrate (16 millimolar) or ammonium (6 millimolar) was supplied to plants grown previously in low nitrate (0.8 millimolar), and changes in the level of PEPC and its mRNA were measured in the basal region of the youngest, fully developed leaves of plants during recovery from nitrogen stress. The exogenous supply of nitrogen selectively increased the levels of protein and mRNA for PEPC. This increase was more pronounced in plants supplemented with ammonium than with nitrate. The accumulation of PEPC during nitrogen recovery increased in parallel with the increase in the activity of glutamine synthetase and/or ferredoxin-dependent glutamate synthase. Among the major amino acids, glutamine was the most influenced during recovery, and its level increased in parallel with the steady-state level of PEPC mRNA for 7 hours after nitrogen supply. The administration of glutamine (12 millimolar) to nitrogen-starved plants increased the steady-state level of PEPC mRNA 7 hours after administration, whereas 12 millimolar glutamate decreased the level of PEPC mRNA. The results indicate that glutamine and/or its metabolite(s) can be a positive control on the nitrogen-dependent regulation of PEPC gene expression in maize leaf cells.

We have utilized the cellular differentiation gradient of the developed, youngest leaf to examine the regulation by nitrogen of levels of phosphoenolpyruvate carboxylase (PEPCase), pyruvate orthophosphate dikinase (PPDK), and ribulose 1,5-bisphosphate carboxylase in maize (Zea mays L.). The protein whose level regulated most preferentially by N availability was PEPCase, followed by PPDK, and the changes in level occurred most conspicuously at the photosynthetically maturing cells. Pulse and pulse-chase experiments to analyze photosynthetic fixation of [(14)C]CO(2) indicate that maize leaf primarily exploited a C(4)-mode of photosynthetic fixation of carbon dioxide even under a selective reduction in levels of these proteins. The effects of N on the synthesis of these proteins and the accumulation of corresponding mRNAs during recovery from a deficiency were examined by pulse and pulse-chase labeling with [(35)S]Met and by hybridization, respectively. The rate of turnover of PPDK was substantially higher than that of the other proteins. Results also showed that the reduced accumulation of PEPCase, as well as PPDK, under N deficiency could largely be accounted for a reduced level of synthesis of protein with a concomitant reduction in level of their mRNAs. This indicates that the N-dependent selective accumulation of these enzymes is primarily a consequence of level of its mRNAs.

Previous studies with intact maize (Zea mays L.) plants indicated that phosphoenolpyruvate carboxylase (PEPC) levels are controlled by nitrogen (N) availability and that this regulation is presumably at the transcriptional level (B. Sugiharto, K. Miyata, H. Nakamoto, H. Sasakawa, T. Sugiyama [1990] Plant Physiol 92: 963-969; B. Sugiharto, T. Sugiyama [1992] Plant Physiol 98: 1403-1408). In the present study, detached maize leaves were used to investigate further the mechanism of N-dependent regulation of gene expression in C(4) plants. PEPC and carbonic anhydrase (CA) mRNA levels decreased in leaves detached from maize plants. Addition of high nitrate did not prevent this decrease. However, the addition of zeatin to solutions bathing the cut ends of the detached leaves inhibited the decrease of PEPC and CA mRNA levels. Simultaneous addition of high nitrate and zeatin to leaves detached from N-deficient maize plants caused a large and rapid increase in PEPC and CA mRNA levels. Zeatin could be replaced by benzyladenine, but not by indoleacetic acid or abscisic acid. Both CA isozymes were effected and responded in an identical manner. We conclude that detached maize leaves provide an excellent experimental system to study the mechanism(s) of N-mediated regulation of PEPC and CA gene expression. However, zeatin is an essential component of this system.

We have used detached leaves to study the N-dependent control of expression of phosphoenolpyruvate carboxylase (PEPC) and carbonic anhydrase (CA) genes in maize (Zea mays L. cv Golden Cross Bantam T51). Following supplementation with an N-source and zeatin, PEPC and CA mRNA levels increased in leaves detached from N-deficient maize plants. Addition of methionine sulfoximine (MSX), a specific inhibitor of glutamine synthetase, inhibited the nitrate-dependent increase of PEPC and CA mRNA but did not affect the glutamine-dependent increase of PEPC and CA mRNA levels. Glutamine levels in detached maize leaves treated with various N sources in the presence or absence of MSX correlated with the levels of PEPC and CA mRNA. We conclude that glutamine is the most likely effector for controlling the N-dependent expression of PEPC and CA in maize plants.

Plant growth-promoting rhizobacteria (PGPR) are promising candidates that enhance plant growth under stressful conditions. In this study, 10 bacterial isolates were screened for their IAA production, among them JTB1 and MT22 isolates were selected which produced high IAA levels under 10% PEG and 2% NaCl stress. The isolates showed a high capacity for phosphate solubilization and ACC deaminase activity. Phytogenic analysis showed that the isolate belonged to Bacillus megaterium species JTB1 and MT22. Application of JTB1, MT22, and their consortia as PGPR significantly promoted root development and sugarcane growth under moderate and severe drought stress. Sugarcane growth promotion resulted from the retardation of reactive oxygen species (ROS) synthesis, malondialdehyde (MDA), electrolyte leakage, and cell damage by increasing antioxidant scavenging systems, such as catalase (CAT) and ascorbate peroxidase (APX), owing to PGPR inoculation under drought stress. Inoculation with PGPR resulted in increased auxin transporter expression, which modulated the increase in photosynthetic gene expression of RBC-L, PEPC, SPS in sugarcane under drought stress. The application of JTB1, MT22, or their consortia seemed to have similar effects on all observed parameters. Collectively, these results indicated that inoculation with PGPR enhanced root development and increased the antioxidant system and photosynthetic activity, which promoted sugarcane growth under drought stress.