Summary Cytochromes P450 (CYPs) play key role in generating the structural diversity of terpenoids, the largest group of plant natural products. However, functional characterization of CYPs has been challenging because of the expansive families found in plant genomes, diverse reactivity and inaccessibility of their substrates and products.Here we present the characterization of two CYPs, CYP76AH3 and CYP76AK1, that act sequentially to form a bifurcating pathway for the biosynthesis of tanshinones, the oxygenated diterpenoids from the Chinese medicinal plant Danshen.These CYPs had similar transcription profiles to that of the known gene responsible for tanshinone production in elicited Danshen hairy roots. Biochemical and RNA interference studies demonstrated that both CYPs are promiscuous. CYP76AH3 oxidizes ferruginol at two different carbon centers, and CYP76AK1 hydroxylates C-20 of two of the resulting intermediates. Together, these convert ferruginol into 11,20-dihydroxy ferruginol and 11,20-dihydroxy sugiol en route to tanshinones. Moreover, we demonstrate the utility of these CYPs by engineering yeast for heterologous production of six oxygenated diterpenoids, which in turn enabled structural characterization of three novel compounds produced by CYP-mediated oxidation.Our results highlight the incorporation of multiple CYPs in diterpenoids metabolic engineering, and a continuing trend of CYPs promiscuity generating complex networks in terpenoid biosynthesis.

SUMMARYFloral organ identity is defined by organ homoetic genes whose coordinated expression is crucial with respect to the time and place of floral organ formation. Here, we report molecular cloning and characterization of the rice STAMENLESS 1 (SL1) gene that is involved in floral development. The sl1 mutant largely resembles the rice B-class gene mutant spw1; both exhibit homeotic conversions of lodicules and stamens to palea/lemmalike organs and carpels. Additionally, sl1 produces flowers with varied numbers of inner floral organs, and amorphous tissues without floral organ identity were frequently formed in whorls 3 and 4. We also show that SL1 specifies lodicule and stamen identities through positive transcriptional regulation of SPW1/OsMADS16 expression. SL1 encodes a member of the C2H2 family of zinc finger proteins, closely related to JAG of Arabidopsis. The functional divergence between SL1 and JAG implies that SL1 was co-opted for its distinctive roles in specification of floral organ identity in rice after the lineage split from Arabidopsis.