Multi-omics elucidation of the phytohormone biosynthetic pathways in Epichloë sinensis

Abstract

Epichloë sinensis is an endophyte species that forms a mutualistic symbiosis with Festuca sinensis, which improves host fitness by promoting growth and enhancing stress tolerance. Phytohormones play essential roles in regulating plant–microbial interactions. Previous research has found that E. sinensis is able to produce growth-promoting hormones, including auxin (IAA), cytokinins (CTK), abscisic acid (ABA), and gibberellins (GA). However, the underlying biosynthetic pathways remain uncharacterized. In the present study, E. sinensis strain 57A was used to systematically investigate phytohormone biosynthetic pathways based on genomic and transcriptomic analyses. Genome annotation identified 53 putative phytohormone biosynthesis genes, including key enzymes such as 6 aromatic amino acid aminotransferases, 1 indole-3-pyruvate decarboxylase, 14 aldehyde dehydrogenases, 5 nitrile hydratases, 1 ent-copalyl diphosphate synthase, and 6 cytochrome P450 monooxygenase. Comparative pathway analysis indicated that IAA in E. sinensis is primarily synthesized through the indole-3-pyruvic acid (IPyA) pathway, involving sequential reactions catalysed by aminotransferase, indole-3-pyruvate decarboxylase, and dehydrogenase. Additionally, E. sinensis may synthesize CTK through both the de novo biosynthesis pathway and the tRNA degradation pathway. The former transfers the isopentenyl chain from dimethylallyl pyrophosphate or 4-hydroxy-3-methyl-but-2-enyl pyrophosphate to the N6-amino group of adenosine monophosphate, while the latter generates CTK through the breakdown of prenylated tRNAs. Notably, the canonical GA and ABA biosynthesis pathways in E. sinensis appear incomplete, with absent homologs of alpha-ionylidenethane synthase aba3, ent-kaurene synthase, and ent-kaurenoic acid oxidase. This suggests that E. sinensis may utilize non-canonical or currently unknown pathways to produce GA and ABA, which warrants further investigation. Transcriptome analysis of E. sinensis in vitro showed that the majority of genes involved in IAA and CTK biosynthesis were expressed, which provides molecular evidence supporting our hypothesis regarding the phytohormone biosynthesis pathways in E. sinensis. In conclusion, this study provides the first comprehensive elucidation of phytohormone biosynthesis in E. sinensis and reconstructs the putative IAA and CTK biosynthetic pathways. These findings contribute to a deeper understanding of the E. sinensis endophyte and lay a theoretical foundation for further investigation into the symbiotic mechanisms.