A cytosolic bifunctional geranyl/farnesyl diphosphate synthase provides MVA-derived GPP for geraniol biosynthesis in rose flowers

Geraniol derived from essential oils of various plant species is widely used in the cosmetic and perfume industries. It is also an essential trait of the pleasant smell of rose flowers. In contrast to other monoterpenes which are produced in plastids via the methyl erythritol phosphate pathway, geraniol biosynthesis in roses relies on cytosolic NUDX1 hydrolase which dephosphorylates geranyl diphosphate (GPP). However, the metabolic origin of cytosolic GPP remains unknown. By feeding Rosa chinensis “Old Blush” flowers with pathway-specific precursors and inhibitors, combined with metabolic profiling and functional characterization of enzymes in vitro and in planta, we show that geraniol is synthesized through the cytosolic mevalonate (MVA) pathway by a bifunctional geranyl/farnesyl diphosphate synthase, RcG/FPPS1, producing both GPP and farnesyl diphosphate (FPP). The downregulation and overexpression of RcG/FPPS1 in rose petals affected not only geraniol and germacrene D emissions but also dihydro-β-ionol, the latter due to metabolic cross talk of RcG/FPPS1-dependent isoprenoid intermediates trafficking from the cytosol to plastids. Phylogenetic analysis together with functional characterization of G/FPPS orthologs revealed that the G/FPPS activity is conserved among Rosaceae species. Site-directed mutagenesis and molecular dynamic simulations enabled to identify two conserved amino acids that evolved from ancestral FPPSs and contribute to GPP/FPP product specificity. Overall, this study elucidates the origin of the cytosolic GPP for NUDX1-dependent geraniol production, provides insights into the emergence of the RcG/FPPS1 GPPS activity from the ancestral FPPSs, and shows that RcG/FPPS1 plays a key role in the biosynthesis of volatile terpenoid compounds in rose flowers.