发表期刊：The Plant Journal

发表时间：2026

Abstract:

β-carboline alkaloids such as harmine and harmaline are prominent specialized metabolites in Peganum multisectum and exert potent inhibitory effects on seedling growth of treated plants, highlighting their potential for development as novel herbicides. However, the enzymatic logic underlying their biosynthesis remains unresolved. Here, we identified two O-methyltransferases, PmOMT1 and PmOMT2, that catalyze the 7-O-methylation of harmol and harmolol, respectively, to form harmine and harmaline. Heterologous expression, biochemical assays, and substrate-feeding experiments in Nicotiana benthamiana confirm their catalytic activities. Intriguingly, phylogenetic analysis and ancestral sequence reconstruction revealed that these enzymes evolved from broadly distributed caffeic acid O-methyltransferases (COMTs). COMT enzymes, which function in S-lignin and melatonin biosynthesis, also have weak β-carboline 7-O-methylation activity and provide a latent foundation for subsequent specialization. Structural modeling and site-directed mutagenesis further revealed key residues and pocket remodeling events that underlie the shift in substrate recognition in PmOMT1 and PmOMT2 relative to canonical COMTs. More importantly, phytotoxicity assays showed that harmine and harmaline exhibit stronger inhibitory effects on Amaranthus tricolor seedling root growth than harmol and harmolol, pointing to possible ecological roles for β-carboline methylation in Peganum. Together, our findings uncover the O-methylation step in β-carboline biosynthesis and illuminate how pre-existing catalytic potential in COMTs can be co-opted to generate lineage-specific alkaloid chemistry in Peganum.

https://doi.org/10.1111/tpj.70998