De novo transcriptome assembly for the basal angiosperm Illicium anisatum provides insights into the biosynthesis of shikimate and neurotoxin anisatin

Illicium anisatum, an ancient angiosperm belonging to the ANITA grade, contains abundant plant metabolites with antimicrobial activity, including shikimate and sesquiterpene lactones (STLs) such as anisatin. The aim of this study was to generate a full-length transcriptome resource for identifying genes related to the shikimate and STL biosynthetic pathways in I. anisatum and for studying the evolution of Illicium species. We performed RNA isoform sequencing of I. anisatum leaf, stem, flower, and floral bud samples and annotated the assembled transcripts based on a homology search. A total of 148,593 transcripts with an average length of 2.2 kb and high assembly completeness were generated. Functional pathway analysis revealed the evolutionary conservation of the shikimate and aromatic amino acid (AAA) biosynthetic pathways in I. anisatum and the upregulation of genes that encode chorismite synthase, which is highly associated with the production of shikimate and serve as a precursor for the biosynthesis of AAAs, in floral organs. I. anisatum germacrene A oxidase (IaGAO) and germacrene D synthase (IaGDS), which participate in STL biosynthesis, showed evolutionary divergence from their homologs in the plants of the family Asteraceae, and their genes were mainly expressed in flower organs. Interestingly, alternative splicing isoforms of IaGDS transcripts were found, probably resulting in differential expression in flower and floral bud. We also constructed a map of the I. anisatum chloroplast genome. Isoform sequencing provided a high level of transcriptome assembly completeness and gene annotation and enabled effective prediction of protein domains. The reported long-read sequencing-based de novo assembly of the I. anisatum transcriptome should aid in exploring genes related to the shikimate and STL biosynthesis pathways and associated molecular mechanisms in Illicium species.