Abstract
Phosphoenolpyruvate carboxylase (PEPC) was overproduced in the leaves of rice plants by introducing the intact maize C4-specific PEPC gene. Maize PEPC in transgenic rice leaves underwent activity regulation through protein phosphorylation in a manner similar to endogenous rice PEPC but contrary to that occurring in maize leaves, being downregulated in the light and upregulated in the dark. Compared with untransformed rice, the level of the substrate for PEPC (phosphoenolpyruvate) was slightly lower and the product (oxaloacetate) was slightly higher in transgenic rice, suggesting that maize PEPC was functioning even though it remained dephosphorylated and less active in the light. 14CO2 labeling experiments indicated that maize PEPC did not contribute significantly to the photosynthetic CO2 fixation of transgenic rice plants. Rather, it slightly lowered the CO2 assimilation rate. This effect was ascribable to the stimulation of respiration in the light, which was more marked at lower O2 concentrations. It was concluded that overproduction of PEPC does not directly affect photosynthesis significantly but it suppresses photosynthesis indirectly by stimulating respiration in the light. We also found that while the steady-state stomatal aperture remained unaffected over a wide range of humidity, the stomatal opening under non-steady-state conditions was destabilized in transgenic rice.
Similar content being viewed by others
References
Agarie S, Miura A, Sumikura R, Tsukamoto S, Nose A, Arima S, Matsuoka M and Miyao-Tokutomi M (2002) Overexpression of C4 PEPC caused O2-insensitive photosynthesis in transgenic rice plants. Plant Sci 162: 257–265
Assmann SM (1993) Signal transduction in guard cells. Ann Rev Cell Biol 9: 345–375
Atkin OK, Evans JR, Ball MC, Siebke K, Pons TL and Lambers H (1998) Light inhibition of leaf respiration: The role of irradiance and temperature. In: Moller IM, Gardeström P, Glimelius K and Glaser E (eds) Plant Mitochondria: From Gene to Function, pp 25–32, Backhuys Publishers, Leiden, The Netherlands
Bakrim N, Nhiri M, Pierre J-N and Vidal J (1998) Metabolite control of Sorghum C4 phosphoenolpyruvate carboxylase catalytic activity and phosphorylation state. Photosynth Res 58: 153–162
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254
Chollet R, Vidal J and O'Leary MH (1996) Phosphoenolpyruvate carboxylase: A ubiquitous, highly regulated enzyme in plants. Ann Rev Plant Physiol Plant Mol Biol 47: 273–298
Dong L-Y, Masuda T, Kawamura T, Hata S and Izui K (1998) Cloning, expression, and characterization of a root-form phosphoenolpyruvate carboxylase from Zea mays: Comparison with the C4-form enzyme. Plant Cell Physiol 39: 865–873
Duff SMG and Chollet R (1995) In vivo regulation of wheat-leaf phosphoenolpyruvate carboxylase by reversible phosphorylation. Plant Physiol 107: 775–782
Fukayama H, Tsuchida H, Agarie S, Nomura M, Onodera H, Ono K, Lee BH, Hirose S, Toki S, Ku MSB, Makino A, Matsuoka M and Miyao M (2001) Significant accumulation of C4-specific pyruvate, orthophosphate dikinase in a C3 plant, rice. Plant Physiol 127: 1136–1146
Gehlen J, Panstruga R, Smets H, Merkelbach S, Kleines M, Porsch P, Fladung M, Becker I, Rademacher T, Häusler RE and Hirsch HJ (1996) Effects of altered phosphoenolpyruvate carboxylase activities on transgenic C3 plant Solanum tuberosum. PlantMol Biol 32: 831–848
Hatch MD (1971) The C4-pathway of photosynthesis. Evidence for an intermediate pool of carbon dioxide and the identity of the donor C4-dicarboxylic acid. Biochem J 125: 425–432
Hatch MD (1976) The C4 pathway of photosynthesis: Mechanism and function. In: Burris RH and Black CC (eds) CO2 Metabolism and Plant Productivity, pp 59–81. University Park Press, Baltimore, Maryland
Hatch MD (1979) Mechanism of C4 photosynthesis in Chloris gayana: Pool sizes and kinetics of 14CO2 incorporation into 4-carbon and 3-carbon intermediates. Arch Biochem Biophys 194: 117–127
Hatch MD and Kagawa T (1976) Photosynthetic activities of isolated bundle sheath cells in relation to differing mechanisms of C4 pathway photosynthesis. Arch Biochem Biophys 175: 39–53
Hatch MD and Slack CR (1966) Photosynthesis by sugar-cane leaves. A new carboxylation reaction and the pathway of sugar formation. Biochem J 101: 103–111
Häusler RE, Kleines M, Uhrig H, Hirsch H-J and Smets H (1999) Overexpression of phosphoenolpyruvate carboxylase from Corynebacterium glutamicum lowers the CO2 compensation point (Γ*) and enhances dark and light respiration in transgenic potato. J Exp Bot 50: 1231–1242
Häusler RE, Rademacher T, Li J, Lipka V, Fischer KL, Schubert S, Kreuzaler F and Hirsch H-J (2001) Single and double overexpression of C4-cycle genes had differential effects on the pattern of endogenous enzymes, attenuation of photorespiration and on contents of UV protectants in transgenic potato and tobacco plants. J Exp Bot 52: 1785–1803
Häusler RE, Hirsch H-J, Kreuzaler F and Peterhänsel C (2002) Overexpression of C4-cycle enzymes in transgenic C3 plants: a biotechnological approach to improve C3-photosynthesis. J Exp Bot 53: 591–607
Heineke D, Riens B, Grosse H, Hoferichter P, Peter U, Flügge UI and Heldt HW (1991) Redox transfer across the inner chloroplast envelope membrane. Plant Physiol 95: 1131–1137
Jiao D, Li X, Huang X, Chi W, Kuang T and Ku MSB (2001) The characteristics of CO2 assimilation of photosynthesis and chlorophyll fluorescence in transgenic PEPC rice. Chin Sci Bull 46: 1080–1084
Jiao D, Huang X, Li X, Chi W, Kuang T, Zhang Q, Ku MSB and Cho D (2002) Photosynthetic characteristics and tolerance to photo-oxidation of transgenic rice expressing C4 photosynthesis enzymes. Photosynth Res 72: 85–93
Kogami H, Shono M, Koike T, Yanagisawa S, Izui K, Sentoku N, Tanifuji S, Uchimiya H and Toki S (1994) Molecular and physiological evaluation of transgenic tobacco plants expressing amaize phosphoenolpyruvate carboxylase gene under the control of the cauliflower mosaic virus 35S promoter. Transgen Res 3: 287–296
Ku MSB, Agarie S, Nomura M, Fukayama H, Tsuchida H, Ono K, Hirose S, Toki S, Miyao M and Matsuoka M (1999) Highlevel expression of maize phosphoenolpyruvate carboxylase in transgenic rice plants. Nat Biotechnol 17: 76–80.
Ku MSB, Cho D, Ranade U, Hsu T-P, Li X, Jiao D-M, Ehleringer J, Miyao M and Matsuoka M(2000) Photosynthetic performance of transgenic rice plants overexpressing maize C4 photosynthesis enzymes. In: Sheehy JE, Mitchell PL and Hardy B (eds) Redesigning Rice Photosynthesis to Increase Yield, pp 193–204. Elsevier Science Publishers, Amsterdam
Lepiniec L, Vidal J, Chollet R, Gadal P and Crétin C (1994) Phosphoenolpyruvate carboxylase: Structure, regulation and evolution. Plant Sci 99: 111–124
Leport L, Kandlbinder A, Baur B and Kaiser WM (1996) Diurnal modulation of phosphoenolpyruvate carboxylation in pea leaves and roots as related to tissue malate concentrations and to the nitrogen source. Planta 198: 495–501
Li B, Zhang XQ and Chollet R (1996) Phosphoenolpyruvate carboxylase kinase in tobacco leaves is activated by light in a similar but not identical way as in maize. Plant Physiol 111: 497–505
Matsuoka M, Fukayama H, Tsuchida H, Nomura M, Agarie S, Ku MSB and Miyao M (2000) How to express some C4 photosynthesis genes at high levels in rice. In: Sheehy JE, Mitchell PL and Hardy B (eds) Redesigning Rice Photosynthesis to Increase Yield, pp 167–175. Elsevier Science Publishers, Amsterdam
Matsuoka M, Furbank RT, Fukayama H and Miyao M (2001) Molecular engineering of C4 photosynthesis. Annu Rev Plant Physiol Plant Mol Biol 52: 297–314
Miyao M (1994) Involvement of active oxygen species in degradation of the D1 protein under strong illumination in isolated subcomplexes of Photosystem II. Biochemistry 33: 9722–9730
Nimmo HG (1998) Circadian regulation of a plant protein kinase. Chronobiol Int 15: 109–118
Svensson P, Bläsing O and Westhoff P (1997) Evolution of the enzymatic characteristics of C4 phosphoenolpyruvate carboxylase. A comparison of the orthologous PPCA phosphoenolpyruvate carboxylases of Flaveria trinervia (C4) and Flaveria pringlei (PC3). Eur J Biochem 246: 452–460
Toh H, Kawamura T and Izui K (1994) Molecular evolution of phosphoenolpyruvate carboxylase. Plant Cell Environ 17: 31–43
Ueno Y, Imanari E, Emura J, Yoshizawa-Kumagaye K, Nakajima K, Inami K, Shiba T, Sakakibara H, Sugiyama T and Izui K (2000) Immunological analysis of the phosphorylation state of maize C4-form phosphoenolpyruvate carboxylase with specific antibodies raised against a synthetic phosphorylated peptide. Plant J 21: 17–26
Vidal J and Chollet R (1997) Regulatory phosphorylation of C4 PEP carboxylase. Trends Plant Sci 2: 230–237
von Caemmerer S and Farquhar GD (1981) Some relationships between the biochemistry of photosynthesis and the gas exchange of leaves. Planta 153: 376–387
Wang YH and Chollet R (1993) Partial purification and characterization of phosphoenolpyruvate carboxylase protein-serine kinase from illuminated maize leaves. Arch Biochem Biophys 304: 496–502
Winter H, Robinson DG and Heldt HW (1993) Subcellular volumes and metabolite concentrations in barley leaves. Planta 191: 180–190
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Fukayama, H., Hatch, M.D., Tamai, T. et al. Activity regulation and physiological impacts of maize C4-specific phosphoenolpyruvate carboxylase overproduced in transgenic rice plants. Photosynthesis Research 77, 227–239 (2003). https://doi.org/10.1023/A:1025861431886
Issue Date:
DOI: https://doi.org/10.1023/A:1025861431886