Cloning and characterization of the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway genes of a natural-rubber producing plant, Hevea brasiliensis.
Natural rubber is synthesized as rubber particles in the latex, the fluid cytoplasm of laticifers, of Hevea brasiliensis. Although it has been found that natural rubber is biosynthesized through the mevalonate pathway, the involvement of an alternative 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway is uncertain. We obtained all series of the MEP pathway candidate genes by analyzing expressed sequence tag (EST) information and degenerate PCR in H. brasiliensis. Complementation experiments with Escherichia coli mutants were performed to confirm the functions of the MEP pathway gene products of H. brasiliensis together with those of Arabidopsis thaliana, and it was found that 1-deoxy-D-xylulose-5-phosphate reductoisomerase, 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase, and 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase of H. brasiliensis were functionally active in the E. coli mutants. Gene expression analysis revealed that the expression level of the HbDXS2 gene in latex was relatively high as compared to those of other MEP pathway genes. However, a feeding experiment with [1-(13)C] 1-deoxy-D-xylulose triacetate, an intermediate derivative of the MEP pathway, indicated that the MEP pathway is not involved in rubber biosynthesis, but is involved in carotenoids biosynthesis in H. brasiliensis.
Sando
Tomoki
T
Department of Biotechnology, Graduate School of Engineering, Osaka University, Japan.
Takeno
Shinya
S
Watanabe
Norie
N
Okumoto
Hiroshi
H
Kuzuyama
Tomohisa
T
Yamashita
Atsushi
A
Hattori
Masahira
M
Ogasawara
Naotake
N
Fukusaki
Eiichiro
E
Kobayashi
Akio
A
D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D003001:Cloning, Molecular; D030541:Databases, Genetic; D004896:Erythritol / Q000031:analogs & derivatives* / Q000378:metabolism; D019665:Euphorbiaceae / Q000235:genetics* / Q000378:metabolism*; D020224:Expressed Sequence Tags; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics*; D028482:Hevea / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D012408:Rubber / Q000378:metabolism*; D036226:Seedlings / Q000235:genetics / Q000378:metabolism; D013194:Staining and Labeling; D013403:Sugar Phosphates / Q000378:metabolism*; D014996:Xylulose / Q000031:analogs & derivatives / Q000378:metabolism
C114232:2-C-methylerythritol 4-phosphate; D002247:Carbon Isotopes; D013403:Sugar Phosphates; D014996:Xylulose; C029724:1-deoxy-2-pentulose; D012408:Rubber; D004896:Erythritol