Overexpression of the chloroplastic 2-oxoglutarate/malate transporter disturbs carbon and nitrogen homeostasis in rice

Abstract

The chloroplastic 2-oxaloacetate/malate transporter (OMT1 or DiT1) takes part in the malate valve that protects chloroplasts from excessive redox poise through export of malate and import of oxaloacetate (OAA). Together with the glutamate/malate transporter (DCT1 or DiT2), it connects carbon with nitrogen assimilation, by providing 2-oxoglutarate for the GS/GOGAT reaction and exporting glutamate to the cytoplasm. OMT1 further plays a prominent role in C4 photosynthesis: OAA resulting from PEP-carboxylation is imported into the chloroplast, reduced to malate by plastidic NADP-MDH, and then exported for transport to bundle sheath cells. Both transport steps are catalyzed by OMT1, at the rate of net carbon assimilation. Therefore, to engineer C4 photosynthesis into C3 crops, OMT1 must be expressed in high amounts on top of core C4 metabolic enzymes. We report here high-level expression of ZmOMT1 from maize in rice (Oryza sativa ssp. indica IR64). Increased activity of the transporter in transgenic rice was confirmed by reconstitution of transporter activity into proteoliposomes. Unexpectedly, over-expression of ZmOMT1 in rice negatively affected growth, CO2 assimilation rate, total free amino acid contents, TCA cycle metabolites, as well as sucrose and starch contents. Accumulation of high amounts of aspartate and the impaired growth phenotype of OMT1 rice lines could be suppressed by simultaneous over-expression of ZmDiT2. Implications for engineering C4-rice are discussed.

Diversity in Acidity between Core and Pulp of Asian Pear Fruit Is Mainly Regulated by the Collaborative Activity of PH8.1 and DIC2 Genes during Fruit Development

“…This study showed that the ALMT9like2 expression level was significantly higher at 60 DAF, while at 110 DAF, another gene tDT2 expression level was significantly lower in the core than in the pulp; moreover, the tDT2 relative mRNA level showed a positive correlation with the malate contents (Figure 4B). Likewise, in plants, a di-carboxylate transporter gene (DIT) is known to be an important malate valve regulator to maintain the redox balance [17,18]. This study showed that the relative expression level of DIT2 was significantly lower in the core than in the pulp at 85 and 110 DAF; Pearson correlation analysis also showed a significant and positive correlation between the DIT2 relative mRNA level and the malate content (Figure 4B).…”

“…The transportation of additional malate from the cytosol into the cell vacuole is an important step to maintain the cytosolic pH and prevent cellular damage, which is possibly controlled by two key genes named aluminum-activated malate transporter 9 (ALMT9) and tonoplast di-carboxylate transporter (tDT) in grapes, apples, and tomatoes [14][15][16]. Additionally, di-carboxylate transporter (DIT) is homologous to chloroplastic 2-oxoglutarate/malate transporter (OMT1) and is an important malate valve regulator to maintain redox balance and nitrogen assimilation in plant cells [17,18].…”

Suppression of metabolite shuttles for export of chloroplast and mitochondrial ATP and NADPH increases the cytosolic NADH:NAD+ ratio in tobacco leaves in the dark

“…ZmOMT1, was overexpressed in rice as part of efforts to introduce C4 photosynthesis in C3 crops (Zamani-Nour et al, 2021). The transgenic lines with the highest expression levels showed reduced height and leaf lesions in mature leaves, accompanied by small reductions in CO2 assimilation rates and increased photorespiration and dark respiration rates (Zamani-Nour et al, 2021).…”

“…ZmOMT1, was overexpressed in rice as part of efforts to introduce C4 photosynthesis in C3 crops (Zamani-Nour et al, 2021). The transgenic lines with the highest expression levels showed reduced height and leaf lesions in mature leaves, accompanied by small reductions in CO2 assimilation rates and increased photorespiration and dark respiration rates (Zamani-Nour et al, 2021). The phenotype was rescued by simultaneously introducing ZmDiT1 into the ZmOMT1 transgenic lines, which suggested that defects were caused by an imbalance between the transport of OAA, malate, and 2-oxoglutarate by pOMT1 and glutamate and aspartate by DiT1.…”

Transport Proteins Enabling Plant Photorespiratory Metabolism

“…Experiments overexpressing Dit1/OMT in rice resulted in strong reduction of photosynthesis and growth, and it could only be rescued by concomitant expression of DiT2.2/DCT2. The results suggest that the DiT1/OMT transporter activity needs to be regulated in close coordination with photosynthesis [ 97 ]. The DiT1/OMT gene was also shown to be under strong evolutionary pressure during evolution of C 4 in NADP-ME type grasses [ 98 ].…”