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

Shirin Zamani-Nour,Berkley J Walker,W Paul Quick,Hsiang-Chun Lin,Efren Bagunu,Tammy L Sage,Tabea Mettler-Altmann,Andreas P M Weber,Shanta Karki,Roxana Khoshravesh,Christine Raines

doi:10.1093/jxb/eraa343

Abstract

The chloroplastic 2-oxaloacetate (OAA)/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 OAA. Together with the glutamate/malate transporter (DCT1 or DiT2), it connects carbon with nitrogen assimilation, by providing 2-oxoglutarate for the GS/GOGAT (glutamine synthetase/glutamate synthase) reaction and exporting glutamate to the cytoplasm. OMT1 further plays a prominent role in C4 photosynthesis: OAA resulting from phosphoenolpyruvate carboxylation is imported into the chloroplast, reduced to malate by plastidic NADP-malate dehydrogenase, and then exported for transport to bundle sheath cells. Both transport steps are catalyzed by OMT1, at the rate of net carbon assimilation. 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, overexpression of ZmOMT1 in rice negatively affected growth, CO2 assimilation rate, total free amino acid content, tricarboxylic acid 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 overexpression of ZmDiT2. Implications for engineering C4 rice are discussed.

Highlights

Population growth, climate change, and lack of arable land are causing greater dependence on crop yield improvement
OMT1 further plays a prominent role in C4 photosynthesis: OAA resulting from phosphoenolpyruvate carboxylation is imported into the chloroplast, reduced to malate by plastidic NADP-malate dehydrogenase, and exported for transport to bundle sheath cells
Carbon:nitrogen (C:N) ratio measurement The ratio of carbon to nitrogen as well as δ 13C were analyzed based on leaf dry weight of 30-day-old and 50-day-old transgenic plants using the ISOTOPE cube elemental analyzer connected to an Isoprime 100 isotope ratio mass spectrometer (Elementar, Germany)

Summary

Introduction

Population growth, climate change, and lack of arable land are causing greater dependence on crop yield improvement. Its highest consumption is in Asia (Muthayya et al, 2014) where 60% of the world population exists (Bai et al, 2018), with the highest and lowest rates of poverty and income, respectively (FAO, 2017).boosting rice yield and performance is an important goal for improving the quality of life for a large share of the global population. C3 photosynthesis performs both initial carbon fixation and Calvin–Benson cycle reactions in the mesophyll. In C4 photosynthesis, initial carbon fixation and the Calvin–Benson cycle are carried out separately in the mesophyll and one or more layers of sheath cells (bundle and/or mestome sheath) surrounding the vascular tissue, respectively.This spatial separation concentrates CO2 around the enzyme Rubisco, thereby reducing Rubisco oxygenase activity and the subsequent loss of energy and previously fixed CO2 during photorespiration (Sage et al, 2012). C4 photosynthesis represents an adaptation for coping with stressful conditions, such as drought, high temperature, and light intensity (Edwards et al, 2010)

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