EsERF transcription factor regulates triterpenoid saponin biosynthesis in Eleutherococcus senticosus through three mechanisms

As the principal bioactive constituents of Eleutherococcus senticosus, triterpenoid saponins exhibit diverse pharmacological activities. The E. senticosus ethylene responsive factor (EsERF) critically regulates triterpenoid saponin biosynthesis; however, the specific EsERF TFs modulating this process in E. senticosus and their underlying mechanisms remain uncharacterized. This study identified four EsERF TFs associated with saponin synthesis. Among them, EsERF66, EsERF127, and EsERF130 promoted saponin synthesis by upregulating the expression of the key biosynthetic enzymes EsFPS, EsSS, and EsSE, whereas EsERF138 suppressed saponin synthesis by downregulating these enzymes. All four TFs were localized to the nucleus, and only EsERF66 had self-transcriptional activation activity, the promoting efficacy of which was affected by light quality and DNA methylation. In addition, the four EsERFs dynamically modulated the binding stability to their target genes by forming complexes with interacting proteins. Specifically, EsERF66–EsCPI1, EsERF127–EsIL1, and EsERF130–EsERFBP3 complexes enhanced the transcriptional activity of EsERFs, whereas EsERF127–EsIL1 and EsERF138–EsCS1 complexes attenuated the regulatory effects of EsERFs on target genes. In summary, EsERFs can synergistically regulate triterpenoid saponin biosynthesis in E. senticosus through three mechanisms: direct binding to target genes, DNA methylation modification, and formation of EsERF–protein interaction complexes.