Molecular characterisation and regulation of expression of a novel isopentenyl diphosphate isomerase 1 gene from autumn olive fruit (Elaeagnus umbellata)

摘要

Isopentenyl diphosphate isomerases (IPI) catalyse the interconversion of two five-carbon isoprenoid units, isopentenyl diphosphate and dimethylallyl diphosphate, during the biosynthesis of a large variety of isoprenoids. Here, we report the cloning and functional validation of a novel gene (designated EutIPI1) encoding an IPI from the fruit of Elaeagnus umbellata Thunb., which produces high amounts of lycopene. The open reading frame of EutIPI1 was 705 bp in length and was interrupted by five introns. It encoded a putative polypeptide of 234 amino acids, without a variable N-terminal extension, which may be targeted to peroxisomes due to the presence of a peroxisome targeting sequence at its C-terminus. The activity of the recombinant protein was confirmed by a colour complementation assay in recombinant Escherichia coli cells. The EutIPI1 gene was highly expressed in leaves, moderately expressed in roots and in red fruit, but only weakly expressed in stems and flowers. During the natural ripening process, transcription of the EutIPI1 gene was significantly up-regulated only in dark-yellow fruit. The 484 bp promoter region of the EutIPI1 gene was isolated using PCR-based DNA-walking and contained a cis-acting salicylic acid (SA)-responsive element and a gibberellin (GA(3))-responsive element. Our results showed that exogenous SA or GA(3), as well as four other plant growth regulators (PGRs), including ethylene, auxin, abscisic acid, and jasmonic acid regulated expression of the EutIPI1 gene in PGR-treated yellow fruit. Lycopene accumulation was promoted by GA(3) and ethylene, correlating with up-regulation of EutIPI1 gene expression. Characterisation of the EutIPI1 gene and regulation of its expression will contribute to our understanding of its function during the ripening of autumn olive fruit.