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

Abstract

The chloroplastic 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 -ketoglutarate 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.