miRNA-TF-gene regulatory network in metabolite differentiation of stem and root parts in Ephedra sinica Stapf

Ephedra sinica Stapf is a important plant and it is widely used in pharmaceutical drugs and dietary supplements. Interestingly, its stem and root exhibit different effects and even opposite pharmacological action. Researches have found that these significant differences between the stems and roots are caused by the specific metabolites(SMs) such as the ephedrine alkaloids, ephedradines and mahuannins. However, the exploration for the formation of the differential metabolites still focus on identification of key biosynthetic genes. The process of SMs biosynthesis and upstream regulatory factors of synthetic genes have not yet been deciphered. In this work, through integrated metabolomics and transcriptomics, we found that ephedradines, mahuannins and ephedrannins were mainly synthesized in the roots and therefore accumulated there. Monoterpenoids were synthesized and accumulated in the stems. Additionally, ephedrine alkaloids accumulated in the stems, but the biosynthesis started from the roots. Furthermore, we identified 49 transcription factor families and 143 miRNAs from the mRNA library and the non-redundant miRNA library, respectively. We studied regulatory roles of key TFs and miRNAs, and constructed a key miRNA-TF-gene regulatory network in SMs biosynthesis. These key miRNAs and TFs were considered to co-regulate the expression of the biosynthetic genes. These results would further explain where these SMs are distributed and how they are synthesized. This work provide new insights into the intrinsic mechanism of metabolites differentiation, which lead to the significant pharmacological differences between the stems and roots.