Cis-3-hexenal modulates 3-hydroxy-3-methylglutaryl-CoA reductase at multiple levels to enhance triterpenoid production in the root cultures of Astragalus mongholicus Bunge

3-Hydroxy-3-methylglutaryl-CoA reductase (HMGR) catalyzes the rate-limiting step of the mevalonate (MVA) pathway for cytosolic biosynthesis of isoprenoids such as triterpenoids and sterols. The current study aimed to investigate whether and how HMGR was modulated by cis -3-hexenal in this process in A . mongholicus root cultures. In addition to cis -3-hexenal, lovastatin, an inhibitor of HMGR, was also applied. Compared with the control, cis -3-hexenal similarly reduced HMGR activity to the same level as lovastatin but increased the contents of astragaloside Ⅰ, Ⅲ, cyclocephaloside Ⅱ, and cycloastragenol to higher levels than those of lovastatin. Interestingly, the combination of cis -3-hexenal and lovastatin enhanced the contents of astragaloside Ⅰ, Ⅱ, Ⅲ, and cyclocephaloside Ⅱ to much higher levels within 24 hours. This result revealed that the reduced HMGR activity was not due to inhibition by cis -3-hexenal, which was confirmed by molecular docking, in vitro incubation, and measuring HMGR level. Transcriptome analysis revealed that cis -3-hexenal significantly increased the abundance of four transcripts encoding HMGR and sterol C -22 desaturase. Further real-time PCR and correlation analysis indicated that SNF5, encoded by isoform_35315, was a key transcription regulatory protein involved in multilevel control of HMGR by cis -3-hexenal and its combination with lovastatin in an isoform-specific and time-dependent pattern. In summary, our results elucidated the modulating mechanism and potential of cis -3-hexenal and lovastatin in enhancing astragalosides and cycloastragenol production in A. mongholicus root cultures. • Cis -3-hexenal reduced HMGR activity and level. • Cis -3-hexenal upregulated the transcription of HMGR in an isoform-specific manner. • Transcription regulatory protein SNF5 was involved in multilevel control of HMGR.