Self‐activating and chromosomally colocalized LsERF166 positively regulates costunolide biosynthesis in lettuce

Costunolide, an important natural product with a diverse array of bioactivities, serves as a key precursor in the biosynthesis of sesquiterpene lactones in Asteraceae species. While the biosynthetic pathway of costunolide has been elucidated, the regulatory network remains poorly understood. Lettuce (Lactuca sativa), which is abundant in sesquiterpene lactones, has been selected as a model system to investigate the transcriptional regulation of costunolide biosynthesis. A multi-tissue transcriptome dataset of lettuce has been constructed, revealing that the costunolide biosynthetic genes are coexpressed across various tissues. By integrating coexpression analysis with chromosomal localization studies, an ethylene-responsive factor (ERF) transcription factor (TF), LsERF166, has been identified as being physically adjacent to the genes encoding costunolide synthases (COSs). LsERF166, a typical ethylene-inducible ERF IX TF, positively regulates the biosynthesis of costunolide by directly activating the expression of costunolide biosynthetic genes. Besides, LsERF166 also exhibits self-activation, thereby forming a positive feedback loop. The colocalization of ERF166 and COS is observed in several species in the Asteraceae family, underscoring its functional significance. Our findings reveal a self-activating and chromosomally colocalized ERF TF involved in costunolide biosynthesis, providing further insights into the transcriptional regulation of plant secondary metabolism.