Rice contains a biosynthetic gene cluster associated with production of the casbane‐type diterpenoid phytoalexin <i>ent</i>‐10‐oxodepressin
Liang Jin, Qinqin Shen, Liping Wang, Jiang Liu, Jingye Fu, Le Zhao, Meimei Xu, Reuben J. Peters, Qiang Wang
Abstract
Diterpenoids play important roles in rice microbial disease resistance as phytoalexins, as well as acting in allelopathy and abiotic stress responses. Recently, the casbane-type phytoalexin ent-10-oxodepressin was identified in rice, but its biosynthesis has not yet been elucidated. Here ent-10-oxodepressin biosynthesis was investigated via co-expression analysis and biochemical characterisation, with use of the CRISPR/Cas9 technology for genetic analysis. The results identified a biosynthetic gene cluster (BGC) on rice chromosome 7 (c7BGC), containing the relevant ent-casbene synthase (OsECBS), and four cytochrome P450 (CYP) genes from the CYP71Z subfamily. Three of these CYPs were shown to act on ent-casbene, with CYP71Z2 able to produce a keto group at carbon-5 (C5), while the closely related paralogues CYP71Z21 and CYP71Z22 both readily produce a keto group at C10. Together these C5 and C10 oxidases can elaborate ent-casbene to ent-10-oxodepressin (5,10-diketo-ent-casbene). OsECBS knockout lines no longer produce casbane-type diterpenoids and exhibit impaired resistance to the rice fungal blast pathogen Magnaporthe oryzae. Elucidation of ent-10-oxodepressin biosynthesis and the associated c7BGC provides not only a potential target for molecular breeding, but also, gives the intriguing parallels to the independently assembled BGCs for casbene-derived diterpenoids in the Euphorbiaceae, further insight into plant BGC evolution, as discussed here.