Molecular characterization of the Oncidium orchid HDR gene encoding 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase, the last step of the methylerythritol phosphate pathway.
Two pathways are used by higher plants for the biosynthesis of isoprenoid precursors: the mevalonate pathway in the cytosol and a 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway in the plastids, with 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase (HDR) catalyzing the last step in the MEP pathway. In order to understand the contribution of MEP pathway in isoprenoid biosynthesis of Oncidium orchid, a full-length cDNA corresponding to HDR from the flower tissues of Oncidium Gower Ramsey was cloned. The deduced OncHDR amino acid sequence contains a plastid signal peptide at the N-terminus and four conserved cysteine residues. RT-PCR analysis of HDR in Oncidium flowering plants revealed ubiquitous expression in organs and tissues, with preferential expression in the floral organs. Phylogenetic analysis revealed evolutionary conservation of the encoding HDR protein sequence. The genomic sequence of the HDR in Oncidium is similar to that in Arabidopsis, grape, and rice in structure. Successful complementation by OncHDR of an E. coli hdr(-) mutant confirmed its function. Transgenic tobacco carrying the OncHDR promoter-GUS gene fusion showed expression in most tissues, as well as in reproductive organs, as revealed by histochemical staining. Light induced strong GUS expression driven by the OncHDR promoter in transgenic tobacco seedlings. Taken together, our data suggest a role for OncHDR as a light-activated gene.
Huang
Jian-Zhi
JZ
Institute of Biotechnology, National Pingtung University of Science and Technology, Pingtung 91201, Taiwan.
Cheng
Ting-Chi
TC
Wen
Pei-Jung
PJ
Hsieh
Ming-Hsiun
MH
Chen
Fure-Chyi
FC
D000595:Amino Acid Sequence; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D004896:Erythritol / Q000031:analogs & derivatives* / Q000096:biosynthesis; D018506:Gene Expression Regulation, Plant; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D029595:Orchidaceae / Q000201:enzymology / Q000235:genetics*; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D013403:Sugar Phosphates / Q000096:biosynthesis*; D014026:Nicotiana / Q000235:genetics
C114232:2-C-methylerythritol 4-phosphate; D018076:DNA, Complementary; D018744:DNA, Plant; D010940:Plant Proteins; D013403:Sugar Phosphates; D010088:Oxidoreductases; D004896:Erythritol