Integration of full-length transcriptomics and targeted metabolomics to identify benzylisoquinoline alkaloid biosynthetic genes in Corydalis yanhusuo

Corydalis yanhusuo W.T.Wang is a classic herb that is frequently used in traditional Chinese medicine and is efficacious in promoting blood circulation,enhancing energy,and relieving pain.Benzylisoquinoline alkaloids(BIAs)are the main bioactive ingredients in Corydalis yanhusuo.However,few studies have investigated the BIA biosynthetic pathway in C.yanhusuo,and the biosynthetic pathway of species-specific chemicals such as tetrahydropalmatine remains unclear.We performed full-length transcriptomic and metabolomic analyses to identify candidate genes that might be involved in BIA biosynthesis and identified a total of 101 full-length transcripts and 19 metabolites involved in the BIA biosynthetic pathway.Moreover,the contents of 19 representative BIAs in C.yanhusuo were quantified by classical targeted metabolomic approaches.Their accumulation in the tuber was consistent with the expression patterns of identified BIA biosynthetic genes in tubers and leaves,which reinforces the validity and reliability of the analyses.Full-length genes with similar expression or enrichment patterns were identified,and a complete BIA biosynthesis pathway in C.yanhusuo was constructed according to these findings.Phylogenetic analysis revealed a total of ten enzymes that may possess columbamine-O-methyltransferase activity,which is the final step for tetrahydropalmatine synthesis.Our results span the whole BIA biosynthetic pathway in C.yanhusuo.Our full-length transcriptomic data will enable further molecular cloning of enzymes and activity validation studies.

The chromosome-scale assembly of the Notopterygium incisum genome provides insight into the structural diversity of coumarins

Coumarins,derived from the phenylpropanoid pathway,represent one of the primary metabolites found in angiosperms.The alignment of the tetrahydropyran(THP)and tetrahydrofuran(THF)rings with the lactone structure results in the formation of at least four types of complex coumarins.However,the mechanisms underlying the structural diversity of coumarin remain poorly understood.Here,we report the chromosome-level genome assembly of Notopterygium incisum,spanning 1.64 Gb,with a contig N50 value of 22.7 Mb and 60,021 annotated protein-coding genes.Additionally,we identified the key enzymes responsible for shaping the structural diversity of coumarins,including two p-coumaroyl CoA 2′-hydroxylases crucial for simple coumarins basic skeleton architecture,two UbiA prenyltransferases responsible for angular or linear coumarins biosynthesis,and five CYP736 cyclases involved in THP and THF ring formation.Notably,two bifunctional enzymes capable of catalyzing both demethylsuberosin and osthenol were identified for the first time.Evolutionary analysis implies that tandem and ectopic duplications of the CYP736 subfamily,specifically arising in the Apiaceae,contributed to the structural diversity of coumarins in N.incisum.Conclusively,this study proposes a parallel evolution scenario for the complex coumarin biosynthetic pathway among different angiosperms and provides essential synthetic biology elements for the heterologous industrial production of coumarins.