Structural and interactions analysis of a transcription factor PnMYB2 in Panax notoginseng

Abstract

The main active ingredients of the traditional Chinese medicinal plant, Panax notoginseng, are the Panax notoginseng saponins (PNS). They can be synthesized via the mevalonate pathway; PnSS and PnSE1 are the key rate-limiting enzymes in this pathway. In this study, an interaction between PnMYB2 and the key enzymes was identified and characterized from the P. notoginseng cDNA library using the Y1H technique. Subsequently, X-α-gal color reaction confirmed the interaction between PnMYB2 and the upstream sequences of PnSS and PnSE1 promoters. Full-length cDNA sequence of PnMYB2 was isolated and characterized. PnMYB2 has an open reading frame of 864 bp, encoding 287 amino acids. 3D structural analysis of PnMYB2 indicated that its structure was similar to that of the template. Phylogenetic analysis revealed that PnMYB2 and PgMYB2 are highly homologous and belong to the R2R3 MYB transcription factor (TF). Subcellular localization analysis showed that PnMYB2 was localized in the nucleus. The recombinant protein PnMYB2 was successfully obtained through prokaryotic expression and was confirmed to be an inclusion body protein. Furthermore, electrophoretic mobility shift assay (EMSA) experiments demonstrated that PnMYB2 specifically binds to MYB core and AC-rich elements. This study provides a theoretical basis for transcriptional regulation of saponin biosynthesis in P. notoginseng.