Functional diversification in the Nudix hydrolase gene family drives sesquiterpene biosynthesis in Rosa x wichurana.

Roses use a non-canonical pathway involving a Nudix hydrolase, RhNUDX1, to synthesize their monoterpenes, especially geraniol. Here we report the characterization of another expressedNUDX1gene from the rose cultivarRosaxwichurana,RwNUDX1-2. In order to study the function of the RwNUDX1-2 protein, we analyzed the volatile profiles of an F(1)progeny generated by crossingR. chinensiscv. 'Old Blush' withR. xwichurana. A correlation test of the volatilomes with gene expression data revealed thatRwNUDX1-2is involved in the biosynthesis of a group of sesquiterpenoids, especiallyE,E-farnesol, in addition to other sesquiterpenes.In vitroenzyme assays and heterologousin plantafunctional characterization of theRwNUDX1-2gene corroborated this result. A quantitative trait locus (QTL) analysis was performed using the data ofE,E-farnesol contents in the progeny and a genetic map was constructed based on gene markers. TheRwNUDX1-2gene co-localized with the QTL forE,E-farnesol content, thereby confirming its function in sesquiterpenoid biosynthesis inR. xwichurana. Finally, in order to understand the structural bases for the substrate specificity of rose NUDX proteins, the RhNUDX1 protein was crystallized, and its structure was refined to 1.7 angstrom. By molecular modeling of different rose NUDX1 protein complexes with their respective substrates, a structural basis for substrate discrimination by rose NUDX1 proteins is proposed.