Molecular cloning and characterization of an intronless farnesyl diphosphate synthase (FDP) gene from Indian rubber clone (Hevea brasiliensis Muell. Arg. RRII105): A gene involved in isoprenoid biosynthesis

Abstract

Farnesyl diphosphate synthase (FDP) is a key intermediate in the biosynthesis of isoprenoid compounds and it is also the allylic diphosphate initiator of natural rubber (cis-1,4-polyisoprene) biosynthesis in plants. This report describes the isolation and characterization of full-length cDNA as well as genomic DNA encoding HbFDP from an elite Indian rubber clone (RRII 105) and its expression analysis in Arabidopsis plant. A 1.2kb cDNA encoding farnesyl diphosphate synthase was isolated and the nucleotide sequence analysis revealed an open reading frame of 1029 nucleotides which encoded a protein of 342 amino acids with a predicated molecular mass of 39.4kDa. It is interesting to note that the genomic FDP gene fragment did not contain any introns (intronless) and has a continuous open reading frame encoding a peptide of 342 amino acids. The deduced amino acid sequence of the HbFDP named HbFDP exhibited a high homology with other plant FDPs. Phylogenetic tree analysis showed that HbFDP belonged to the plant FDP group. The putative protein sequence analysis of predicated HbFDP cDNA showed the presence of 2 transmembrane motifs (DDIMD; DDYLD) in the catalytic site. It also contains the DDXXD motifs that are characteristic of prenyltransferases. Genomic southern blot analysis indicated that HbFDP is a member of small gene family in Hevea. Semi-quantitative RT-PCR results clearly indicated that HbFDP gene was differentially expressed in various tissues examined. However, the abundance of mRNA transcript level was found to be high in laticifer cells which indicate the possible involvement of HbFDP gene in natural rubber biosynthesis in Hevea. Further to validate the role of HbFDP gene in plants, HbFDP cDNA was over-expressed in transgenic Arabidopsis plants. Semi-quantitative RT-PCR analysis showed that accumulation of HbFDP mRNA transcripts was 3–5 folds higher in transgenic Arabidopsis plants than control. This data suggest that HbFDP gene might play an important role in the regulation of plant growth and isoprenoid biosynthesis.