Improvement of photosynthesis in rice (Oryza sativa L.) by inserting the C4 pathway
© Karki et al.; licensee Springer. 2013
Received: 9 May 2013
Accepted: 12 September 2013
Published: 28 October 2013
To boost food production for a rapidly growing global population, crop yields must significantly increase. One of the avenues being recently explored is the improvement of photosynthetic capacity by installing the C4 photosynthetic pathway into C3 crops like rice to drastically increase their yield. Crops with an enhanced photosynthetic mechanism would better utilize the solar radiation that can be translated into yield. This subsequently will help in producing more grain yield, reduce water loss and increase nitrogen use efficiency especially in hot and dry environments. This review provides a summary of the factors that need to be modified in rice so that the C4 pathway can be introduced successfully. It also discusses the differences between the C3 and C4 photosynthetic pathways in terms of anatomy, biochemistry and genetics.
To provide adequate food and nutrition to the global population that is expected to reach 9 billion by 2050 (http://www.unpopulation.org), rice yields need to increase by at least 60% (FAO 2009). Rice is the staple food of over half of the world’s population and this rice consuming population is increasing at the rate of 1.098% per annum (http://esa.un.org/wpp/Excel-Data/population.htm). Escalating population means more demand for food, water and land at a time when the natural resource base for agriculture is being degraded because large areas of farmland are being diverted from food production to industrialization and bio-fuel production. An unpredictable climate change is threatening to further reduce agriculturally viable land due to more instances of drought and flood (http://www.fao.org/docrep/017/aq191e/aq191e.pdf). As a growing population and global climate change place increasing pressure on the world’s food supply, it is essential that we continue to improve crop performance in terms of grain productivity to keep pace with population growth. The increase in crop productivity conferred by the plant types created during green revolution period supported the population boom following the two world wars. Since then, despite the use of improved varieties and advanced technologies, the yield potential of present day rice cultivars has improved just a little indicating that these varieties have hit a yield ceiling (Akita 1994). Recently an attempt is underway to increase the rice yield potential by engineering an efficient C4 type photosynthesis into rice (Kajala et al. 2011). For this, a set of genes which regulate leaf anatomy and biochemical processes have to be inserted into rice and expressed in an appropriate manner which is currently not possible solely by conventional plant breeding techniques. Therefore, genetic engineering to improve the photosynthetic pathway of rice would provide sufficient opportunity to enhance the actual grain productivity as well as the yield potential. Genetic engineering provides an efficient and precise breeding tool in which only the genes of interest can be introduced even from distantly related species.
In C3 plants like rice, CO2 is assimilated into a 3-carbon compound by the photosynthetic enzyme ribulose-1, 5-bisphosphate carboxylase oxygenase (Rubisco). As the name indicates, Rubisco also catalyzes oxidation of ribulose-1, 5-bisphosphate (RuBP) in a wasteful process known as photorespiration which can incur a loss of upto 25% of previously fixed carbon (Sage 2004). At temperature above 30°C which is typical of tropical rice growing areas of the world, rate of oxygenation increases substantially and this considerably reduces the photosynthetic efficiency of C3 plants by upto 40% (Ehleringer and Monson 1993). Thus, photosynthesis of rice in the tropics and warm temperate regions becomes inefficient. The C4 plants which have CO2 concentrating mechanism within their leaves have very much reduced levels of photorespiration and thus have evolved to thrive in hot, arid environments and offer valuable insights for crop improvement strategies. Rice with a C4 photosynthesis mechanism would have increased photosynthetic efficiency while using scarce resources such as land, water, and fertilizer specifically nitrogen more effectively (Hibberd et al. 2008). Because it will perform well under high temperature as well as require less water and nitrogen, C4 rice would confer benefits on different types of rice ecosystems including the marginal lands.
C4 type photosynthesis is one of the three types of biochemical mechanisms adopted by plants to fix atmospheric CO2, others being C3 and Crassulacean acid metabolism (CAM) pathways. C4 photosynthesis has evolved more than 66 times independently (Sage et al. 2012) at least in 19 families during angiosperm evolution from C3 ancestors (Muhaidat et al. 2007) and it entails alternations of cellular structures, biochemistry and hence the development of leaves. This highly specialized form of photosynthesis essentially has developed a CO2 concentrating mechanism around the Rubisco enzyme thus eliminating the oxygenase function of Rubisco thereby reducing the wastage of energy due to photorespiration (Douce and Heldt 2000). Rubisco from C4 species is more efficient than from C3 species in terms of carboxylation (Kubien et al. 2008). The other associated benefits of the C4 system include higher water use efficiency because steeper concentration gradient for CO2 diffusion can be maintained through partly closed stomata, higher radiation use efficiency as C4 photosynthesis efficiency does not get saturated at high light intensity (Rizal et al. 2012) and higher nitrogen use efficiencies because it will require less Rubisco and hence less nitrogen.
C4 plants are potentially more productive at higher temperatures typically experienced by rice. To take advantage of this more efficient photosynthetic system at a time when the population and food prices are soaring, there are efforts towards inserting the C4 mechanism such as that found in maize into rice (Rizal et al. 2012). This novel approach to modify the photosynthesis system of rice is a challenging and long term endeavor because the C4 pathway is very complex and many factors controlling the mechanism are still unknown. Therefore, it requires ingenuity and expertise of scientists involved in diverse disciplines such as genetic engineering, biochemistry, bioinformatics, molecular biology, photosynthesis, systems biology, physiology, plant breeding, metabolomics, etc. For the same, the C4 rice consortium was conceptualized and established which began the practical work of C4 rice engineering since 2009 (http://photosynthome.irri.org/C4rice/). This review provides an update on the requirements to develop C4 rice and progress made in the field of genetic engineering. Based on the study of the evolution of C4 from C3 species and the associated changes, the following modifications are essential to establish a functional C4 photosynthetic pathway in rice.
Increase the number and size of chloroplasts in bundle sheath cells of rice
Golden2-like (GLK) gene family members encode nuclear transcription factors that have been implicated to regulate chloroplast development in Arabidopsis, Zea mays, and the moss Physcomitrella patens (Rossini et al. 2001). In each of these species, GLK genes exist as a homologous pair named as GLK1 and GLK2 (Waters et al. 2009). In moss and Arabidopsis the GLK genes are redundant and functionally equivalent whereas in maize and sorghum GLK genes act in a cell-type-specific manner to direct the development of dimorphic chloroplasts (Waters et al. 2008; Wang et al. 2013a). In maize, Golden2 (G2) and its homologue ZmGLK1 transcripts accumulate primarily in BS and M cells, respectively, suggesting a specific role for each gene regulating the dimorphic chloroplast differentiation (Wang et al. 2013a).
Reduce the vein spacing thereby increasing the vein density in the leaf
The activity of the Calvin cycle should be significantly reduced in MC and greatly enhanced in the BSC of rice
The photorespiration in mesophyll cells has to be greatly reduced
In C3 plants carbon fixation and Calvin cycle take place in MCs. During carbon fixation, ribulose- 1, 5- bisphosphate (RuBP) - a five carbon compound, catalyzed by an enzyme ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco, EC.184.108.40.206) reacts with CO2 to form two molecules of 3-carbon compound called the 3-phosphoglycerate (3-PGA). Within the Calvin cycle, the two PGA molecules form an energy rich molecule of sugar (triose phosphate) and regenerates RuBP for the next cycle. At current atmospheric CO2 concentrations (ca. 400 ppm) Rubisco also catalyses a reaction between RuBP and O2 resulting in one molecule each of 2-phosphoglycolate and 3-PGA (Peterhansel and Maurino 2011). The 2-phosphoglycolate has to be converted back to 3-PGA through the process called photorespiration, which involves a series of biochemical reactions. During this process a loss of previously fixed carbon and nitrogen occurs and extra energy must also be used (Sharpe and Offermann 2013).
Differences in carboxylation efficiency (CE) and CO 2 compensation point (CP) between rice (C3) and sorghum (C4) at 21 and 2% oxygen level
CE (mol mol-1)
Increase in CE when O2level was reduced from 21 to 2%
CO2CP (mol mol-1)
Engineering of C4 pathway into rice
It was thought that single cell C4 system could be faster to install in C3 plants. There are attempts to engineer single cell C4 photosynthesis system in rice too (Miyao et al. 2011). To introduce single cell C4–like pathway in which MC is made to capture and release CO2 in the manner it takes place in Hydrilla verticillata (L.f) Royle., four enzymes (PEPC, PPDK, NADP-MDH, and NADP-ME) involved in the pathway were overproduced in the transgenic rice leaves (Ku et al. 1999; Fukayama et al. 2001; Tsuchida et al. 2001; Taniguchi et al. 2008). A few of the major problems encountered that need to be addressed to make a single-cell C4-like pathway in rice are: mechanism to facilitate transport activity of PEP across the chloroplast envelope, the import of OAA into the chloroplasts and the direction of the NADP-ME reaction, involvement of NADP-MDH, the presence of endogenous PEPC inside the MC chloroplast of rice and further elevation of NADP-MDH activity were reported to be necessary (Miyao et al. 2011). Terrestrial single cell C4 species such as Bienertia cycloptera, B. sinuspersici and Suaeda aralocaspica, belonging to Chenopodiaceae family also need spatial compartmentalization of the carbon assimilation and decarboxylation (Chuong et al. 2006). These species have dimorphic chloroplasts in those compartments. The earlier attempts produced futile cycle which was due to no change in anatomy, lack of appropriate transporters and the maize genes transformed into rice were not appropriately expressed in cell specific manner and were not regulated like in maize but were regulated like the endogenous rice C3 isoforms (Miyao et al. 2011).
To engineer the photosynthetic pathway from C3 to C4 within two decades, which took million of years in nature, C4 rice consortium began the simultaneous gene discovery and engineering of already known genes into rice aiming to form C4 rice with Kranz type anatomy. C4 genes such as CA, PEPC, PPDK, NADP-ME, and NADP-MDH are cloned from maize and transformed into rice. Also the transporters that were over expressed in the C4 metabolic pathways such as 2-oxoglutarate/malate transporter (OMT1), dicarboxylate transporter1 (DiT1), dicarboxylate transporter2 (DiT2), PEP/phosphate transporter (PPT1), mesophyll envelope protein (MEP) and triose-phosphate phosphate translocator (TPT) that were recently identified through proteomics of maize BS and MS cells (Friso et al. 2010) are being transformed into rice (Figure 3). The C4 rice consortium members are also involved in discovering novel genes related to Kranz anatomy (Wang et al. 2013b). Once tested, the promising candidate genes controlling the Kranz anatomy will also be introduced in the rice plants that have been engineered with the C4 biochemical pathway genes.
The C4 photosynthetic pathway has evolved more than 66 times in different species, suggesting that C3 plants may be in some way preconditioned to C4 photosynthesis. Developing crop plants with enhanced photosynthesis will improve crop yield and make efficient use of resources in a sustainable manner. The C4 rice consortium is striving to install a maize-like photosynthetic mechanism in rice to break its yield barrier and to breed a new generation of “climate-ready” rice which will yield more even under the situations of increasing temperature and decreasing water availability. This initiative demands a holistic approach of understanding the regulatory networks and the complex mechanism underlying the C4 photosynthetic pathway and their systematic introduction into the rice genome.
We are grateful to Joan Salonga and Glenn Dimayuga for gas exchange measurements, Abigail Elmido-Mabilangan for microscopy and other staff of C4 Rice Center, International Rice Research Institute (IRRI) for their technical support. This study under the C4 Rice Center at IRRI was supported by Bill and Melinda Gates Foundation and UKAID.
- Akita S: Eco-physiological aspect of raising the yield plateau of irrigated rice in the tropics. Proceedings of a workshop on rice yield potential in favorable environments, IRRI, 29 November - 4 December 1993. In Breaking the yield barrier. Edited by: Cassman KG. International Rice Research Institute, Philippines; 1994.Google Scholar
- Chuong SDX, Franceschi VR, Edwards GE: The cytoskeleton maintains organelle partitioning required for single-cell C4 photosynthesis in Chenopodiaceae species. Plant Cell 2006, 18: 2207–2223. 10.1105/tpc.105.036186PubMed CentralView ArticlePubMedGoogle Scholar
- Dengler NG, Nelson T: Leaf structure and development in C4 plants. In C4 plant biology. Edited by: Sage RF, Monson RK. Academic Press, San Diego, California; 1999:133–172.View ArticleGoogle Scholar
- Douce R, Heldt HW: Photorespiration. In Photosynthesis: Physiology and metabolism. Edited by: Leegood RC, Sharkey TD, von Caemmerer S. Kluwer Academic, Dordrecht, The Netherlands; 2000:115–136.View ArticleGoogle Scholar
- Ehleringer JR, Monson RK: Evolutionary and ecological aspects of photosynthetic pathway variation. Annu Rev Ecol Syst 1993, 24: 411–439. 10.1146/annurev.es.24.110193.002211View ArticleGoogle Scholar
- FAO: FAO's Director-General on how to feed the World in 2050. Popul Dev Rev 2009, 35: 837–839. doi: 10.1111/j.1728–4457.2009.00312.x doi: 10.1111/j.1728-4457.2009.00312.xView ArticleGoogle Scholar
- Friso G, Majeran W, Huang MS, Sun Q, van Wijk KJ: Reconstruction of metabolic pathways, protein expression, and homeostasis machineries across maize bundle sheath and mesophyll chloroplasts: large-scale quantitive proteomics using the first maize genome assembly. Plant Physiol 2010, 152: 1219–1250. 10.1104/pp.109.152694PubMed CentralView ArticlePubMedGoogle Scholar
- Fukayama H, Tsuchida H, Agarie S, Nomura M, Onodera H, Ono K, Lee BH, Hirose S, Toki S, Ku MSB, Makino A, Matsuoka M, Miyao M: Significant accumulation of C4-specific Pyruvate, orthophosphate dikinase in a C3 plant, rice. Plant Physiol 2001, 127: 1136–1146. 10.1104/pp.010641PubMed CentralView ArticlePubMedGoogle Scholar
- Hibberd JM, Sheehy JE, Langdale JA: Using C4 photosynthesis to increase the yield of rice- rationale and feasibility. Curr Opin Plant Biol 2008, 11: 228–231. 10.1016/j.pbi.2007.11.002View ArticlePubMedGoogle Scholar
- Kajala K, Covshoff S, Karki S, Woodfield H, Tolley BJ, Dionora MJ, Mogul RT, Mabilangan AE, Danila FR, Hibberd JM, Quick WP: Strategies for engineering a two-celled C4 photosynthetic pathway into rice. J Exp Bot 2011, 62: 3001–3010. 10.1093/jxb/err022View ArticlePubMedGoogle Scholar
- Kebeish R, Niessen M, Thiruveedhi K, Bari R, Hirsch HJ, Rosenkranz R, Stabler N, Schonfeld B, Kreuzaler F, Peterhansel C: Chloroplastic photorespiratory bypass increases photosynthesis and biomass production in Arabidopsis thaliana. Nat Biotechnol 2007, 25: 593–599. doi: 10.1038/nbt1299 doi: 10.1038/nbt1299 10.1038/nbt1299View ArticlePubMedGoogle Scholar
- Kim GT, Tsukaya H, Uchimiya H: The CURLY LEAF gene controls both division and elongation of cells during the expansion of the leaf blade in Arabidopsis thaliana. Planta 1998, 206: 175–183. 10.1007/s004250050389View ArticlePubMedGoogle Scholar
- Ku MSB, Agarie S, Nomura M, Fukayama H, Tsuchida H, Ono K, Hirose S, Toki S, Miyao M, Matsuoka M: High-level expression of maize phospho enol pyruvate carboxylase in transgenic rice plants. Nat Biotechnol 1999, 17: 76–80. 10.1038/5256View ArticlePubMedGoogle Scholar
- Kubien DS, Whitney SM, Moore PV, Jesson LK: The biochemistry of Rubisco in Flaveria. J Exp Bot 2008, 59: 1767–1777.View ArticlePubMedGoogle Scholar
- Leegood RC: Roles of the bundle sheath cells in leaves of C3 plants. J Exp Bot 2008, 59: 1663–1673.View ArticlePubMedGoogle Scholar
- Li Y, Gao Y, Xu X, Shen Q, Guo S: Light saturated photosynthetic rate in high-nitrogen rice (Oryza sativa L.) leaves is related to chloroplast CO2 concentration. J Exp Bot 2009, 60: 2531–2360.Google Scholar
- Matsuoka M, Kyozuka J, Shimamoto K, Kano-Murakami Y: The promoters of two carboxylases in a C4 plant (maize) direct cell-specific, light regulated expression in a C3 plant (rice). Plant J 1994, 6: 311–319. 10.1046/j.1365-313X.1994.06030311.xView ArticlePubMedGoogle Scholar
- Miyao M, Masumoto C, Miyazawa SI, Fukayama H: Lessons from engineering a single-celled C4 photosynthetic pathway into rice. J Exp Bot 2011, 62: 3021–3029. 10.1093/jxb/err023View ArticlePubMedGoogle Scholar
- Monson RK, Rawsthorne S: CO2 assimilation in C3-C4 intermediate plants. In Photosynthesis: physiology and metabolism. Edited by: Leegood RC, Sharkey TD, von Caemmerer S. Kluwer Academic Publishers, Dordrecht, The Netherlands; 2000:533–550.View ArticleGoogle Scholar
- Muhaidat R, Sage RF, Dengler NG: Diversity of Kranz anatomy and biochemistry in C4 eudicots. Am J Bot 2007, 94: 362–381. 10.3732/ajb.94.3.362View ArticlePubMedGoogle Scholar
- Nelson T, Langdale JA: Patterns of leaf development in C4 plants. Plant Cell 1989, 1: 3–13.PubMed CentralView ArticlePubMedGoogle Scholar
- Nikolopoulos D, Liakopoulos G, Drossopoulos I, Karabourniotis G: The relationship between anatomy and photosynthetic performance of heterobaric leaves. Plant Physiol 2002, 129: 235–243. 10.1104/pp.010943PubMed CentralView ArticlePubMedGoogle Scholar
- Nomura M, Higuchi T, Ishida Y, Ohta S, Komari T, Imaizumi N, Miyao-Tokutomi M, Matsuoka M, Tajima S: Differential expression pattern of C4 bundle sheath expression genes in rice, a C3 plant. Plant Cell Physiol 2005, 46: 754–761. 10.1093/pcp/pci078View ArticlePubMedGoogle Scholar
- Peterhansel C, Maurino VG: Photorespiration redesigned. Plant Physiol 2011, 155: 49–55. 10.1104/pp.110.165019PubMed CentralView ArticlePubMedGoogle Scholar
- Rizal G, Karki S, Thakur V, Chatterjee J, Coe RA, Wanchana S, Quick WP: Towards a C4 rice. Asian J Cell Biol 2012, 7: 13–31. 10.3923/ajcb.2012.13.31View ArticleGoogle Scholar
- Robles P, Fleury D, Candela H, Cnops G, Alonso-Peral MM, Anami S, Falcone A, Caldana C, Willmitzer L, Ponce MR, Van Lijsebettens M, Micol JL: The RON1/FRY1/SAL1 gene is required for leaf morphogenesis and venation patterning in Arabidopsis. Plant Physiol 2010, 152: 1357–1372. 10.1104/pp.109.149369PubMed CentralView ArticlePubMedGoogle Scholar
- Rossini L, Cribb L, Martin DJ, Langdale JA: The maize Golden2 gene defines a novel class of transcriptional regulators in plants. Plant Cell 2001, 13: 1231–1244.PubMed CentralView ArticlePubMedGoogle Scholar
- Sage RF: The evolution of C4 photosynthesis. New Phytol 2004, 1618: 341–370.View ArticleGoogle Scholar
- Sage RF, Sage TL, Kocacinar F: Photorespiration and the evolution of C4 photosynthesis. Annu Rev Plant Biol 2012, 63: 19–47. 10.1146/annurev-arplant-042811-105511View ArticlePubMedGoogle Scholar
- Sharpe RM, Offermann S: One decade after the discovery of single-cell C4 species in terrestrial plants: what did we learn about the minimal requirement of C4 photosynthesis? Photosynth Res 2013. doi: 10.1007/s1120–013–9810–9 doi: 10.1007/s1120-013-9810-9Google Scholar
- Slewinski TL, Anderson AA, Zhang C, Turgeon R: Scarecrow plays a role in establishing Kranz anatomy in maize leaves. Plant Cell Physiol 2012, 53: 2030–2037. doi: 10.1093/pcp/pcs147 doi: 10.1093/pcp/pcs147 10.1093/pcp/pcs147View ArticlePubMedGoogle Scholar
- Taniguchi Y, Ohkawa H, Masumoto C, Fukuda T, Tamai T, Lee K, Sudoh S, Tsuchida H, Sasaki H, Fukayama H, Miyao M: Overproduction of C4 photosynthetic enzymes in transgenic rice plants: an approach to introduce the C4-like photosynthetic pathway into rice. J Exp Bot 2008, 59: 1799–1809.View ArticlePubMedGoogle Scholar
- Tolley BJ, Sage TL, Langdale JA, Hibberd JM: Individual maize chromosomes in the C3 plant oat can increase bundle sheath cell size and vein density. Plant Physiol 2012, 159: 1418–1427. 10.1104/pp.112.200584PubMed CentralView ArticlePubMedGoogle Scholar
- Tsuchida H, Tamai T, Fukayama H, Agarie S, Nomura M, Onodera H, Ono K, Nishizawa Y, Lee BH, Hirose S, Toki S, Ku MS, Matsuoka M, Miyao M: High level expression of C4-specific NADP-malic enzyme in leaves and impairment of photoautotrophic growth in a C3 plant, rice. Plant Cell Physiol 2001, 42: 138–145. 10.1093/pcp/pce013View ArticlePubMedGoogle Scholar
- Tsukaya H, Naito S, Redei GP, Komeda Y: A new class of mutations in Arabidopsis thaliana, acaulis1, affecting the development of both inflorescences and leaves. Development 1993, 118: 751–764.Google Scholar
- Ueno O: Structural and biochemical characterization of the C3-C4 intermediate Brassica gravinae and relatives, with particular reference to cellular distribution of Rubisco. J Exp Bot 2011, 62: 5347–5355. 10.1093/jxb/err187PubMed CentralView ArticlePubMedGoogle Scholar
- Wang P, Fouracre J, Kelly S, Karki S, Gowik U, Aubry S, Shaw MK, Westhoff P, Slamet-Loedin IH, Quick WP, Hibberd JM, Langdale JA: Evolution of GOLDEN2-LIKE gene function in C3 and C4 plants. Planta 2013a, 237: 481–495. 10.1007/s00425-012-1754-3View ArticleGoogle Scholar
- Wang P, Kelly S, Fouracre JP, Langdale JA: Genome-wide transcript analysis of early maize leaf development reveals gene cohorts associated with the differentiation of C4 Kranz anatomy. Plant J 2013b, 75: 656–670. 10.1111/tpj.12229View ArticleGoogle Scholar
- Waters MT, Moylan EC, Langdale JA: GLK transcription factors regulate chloroplast development in a cell-autonomous manner. Plant J 2008, 56: 432–444. 10.1111/j.1365-313X.2008.03616.xView ArticlePubMedGoogle Scholar
- Waters MT, Wang P, Korkaric M, Capper RG, Saunders NJ, Langdale JA: GLK transcription factors coordinate expression of the photosynthetic apparatus in Arabidopsis. Plant Cell 2009, 21: 1109–1128. 10.1105/tpc.108.065250PubMed CentralView ArticlePubMedGoogle Scholar
- Weber APM, von Caemmerer S: Plastid transport and metabolism of C3 and C4 plants: comparative analysis and possible biotechnological exploitation. Curr Opin Plant Biol 2010, 13: 257–265.View ArticlePubMedGoogle Scholar
- Yoshimura Y, Kubota F, Ueno O: Structural and biochemical bases of photorespiration in C4 plants: quantification of organelles and glycine decarboxylase. Planta 2004, 220: 307–317. 10.1007/s00425-004-1335-1View ArticlePubMedGoogle Scholar
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