title: Phosphoenolpyruvate carboxylase intrinsically located in the chloroplast of rice plays a crucial role in ammonium assimilation.
abstract: Phosphoenolpyruvate carboxylase (PEPC) is a key enzyme of primary metabolism in bacteria, algae, and vascular plants, and is believed to be cytosolic. Here we show that rice (Oryza sativa L.) has a plant-type PEPC, Osppc4, that is targeted to the chloroplast. Osppc4 was expressed in all organs tested and showed high expression in the leaves. Its expression in the leaves was confined to mesophyll cells, and Osppc4 accounted for approximately one-third of total PEPC protein in the leaf blade. Recombinant Osppc4 was active in the PEPC reaction, showing V(max) comparable to cytosolic isozymes. Knockdown of Osppc4 expression by the RNAi technique resulted in stunting at the vegetative stage, which was much more marked when rice plants were grown with ammonium than with nitrate as the nitrogen source. Comparison of leaf metabolomes of ammonium-grown plants suggested that the knockdown suppressed ammonium assimilation and subsequent amino acid synthesis by reducing levels of organic acids, which are carbon skeleton donors for these processes. We also identified the chloroplastic PEPC gene in other Oryza species, all of which are adapted to waterlogged soil where the major nitrogen source is ammonium. This suggests that, in addition to glycolysis, the genus Oryza has a unique route to provide organic acids for ammonium assimilation that involves a chloroplastic PEPC, and that this route is crucial for growth with ammonium. This work provides evidence for diversity of primary ammonium assimilation in the leaves of vascular plants.
authors.affiliation: Photobiology and Photosynthesis Research Unit, National Institute of Agrobiological Sciences, Kannondai, Tsukuba 305-8602, Japan.
authors.forename: Chisato
authors.initials: C
authors.lastname: Masumoto
authors.forename: Shin-Ichi
authors.initials: S
authors.lastname: Miyazawa
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ohkawa
authors.forename: Takuya
authors.initials: T
authors.lastname: Fukuda
authors.forename: Yojiro
authors.initials: Y
authors.lastname: Taniguchi
authors.forename: Seiji
authors.initials: S
authors.lastname: Murayama
authors.forename: Miyako
authors.initials: M
authors.lastname: Kusano
authors.forename: Kazuki
authors.initials: K
authors.lastname: Saito
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Fukayama
authors.forename: Mitsue
authors.initials: M
authors.lastname: Miyao
mesh_terms: D002736:Chloroplasts / Q000201:enzymology* / Q000235:genetics; D055785:Gene Knockdown Techniques; D017343:Genes, Plant; D007700:Kinetics; D012275:Oryza / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D010730:Phosphoenolpyruvate Carboxylase / Q000235:genetics / Q000378:metabolism*; D053147:Plant Exudates / Q000378:metabolism; D018515:Plant Leaves / Q000166:cytology / Q000378:metabolism; D018520:Plant Shoots / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D052584:Xylem / Q000378:metabolism
chemical_list: D053147:Plant Exudates; D000644:Quaternary Ammonium Compounds; D011994:Recombinant Proteins; D010730:Phosphoenolpyruvate Carboxylase