Characterization and expression analysis of genes encoding phosphoenolpyruvate carboxylase and phosphoenolpyruvate carboxylase kinase of Lotus japonicus, a model legume.
Phosphoenolpyruvate carboxylases (PEPCs), one form of which in each legume species plays a central role in the carbon metabolism in symbiotic root nodules, are activated through phosphorylation of a conserved residue by a specific protein kinase (PEPC-PK). We characterized the cDNAs for two PEPC isoforms of Lotus japonicus, an amide-translocating legume that forms determinate nodules. One gene encodes a nodule-enhanced form, which is more closely related to the PEPCs in amide-type indeterminate nodules than those in ureide-type determinate nodules. The other gene is expressed in shoots and roots at a low level. Both forms have the putative phosphorylation site, Ser11. We also isolated a cDNA and the corresponding genomic DNA for PEPC-PK of L. japonicus. The recombinant PEPC-PK protein expressed in Escherichia coli phosphorylated recombinant maize C4-form PEPC efficiently in vitro. The level of mRNA for PEPC-PK was high in root nodules, and those in shoots and roots were also significant. In situ hybridization revealed that the expression patterns of the transcripts for PEPC and PEPC-PK were similar in mature root nodules, but were different in emerging nodules. When L. japonicus seedlings were subjected to prolonged darkness and subsequent illumination, the activity of PEPC-PK and the mRNA levels of both PEPC and PEPC-PK in nodules decreased and then recovered, suggesting that they are regulated according to the amounts of photosynthates transported from shoots.
Laboratory of Plant Physiology, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
Tomomi
T
Nakagawa
Tomoko
T
Izumi
Mari
M
Banba
Yosuke
Y
Umehara
Hiroshi
H
Kouchi
Katsura
K
Izui
Shingo
S
Hata
D003001:Cloning, Molecular; D017931:DNA Primers; D007887:Fabaceae / Q000201:enzymology / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D000070116:Lotus / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D010730:Phosphoenolpyruvate Carboxylase / Q000235:genetics*; D010802:Phylogeny; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D014158:Transcription, Genetic
D017931:DNA Primers; D012333:RNA, Messenger; D011994:Recombinant Proteins; C069767:phosphoenolpyruvate carboxylase kinase; D017346:Protein Serine-Threonine Kinases; D010730:Phosphoenolpyruvate Carboxylase