ARPI, β-AS, and UGE regulate glycyrrhizin biosynthesis in Glycyrrhiza uralensis hairy roots.

key messageARPI, β-AS, and UGE were cloned from G. uralensis and their regulatory effects on glycyrrhizin biosynthesis were investigated. β-AS and UGE but not ARPI positively regulate the biosynthesis of glycyrrhizin. Glycyrrhiza uralensis Fisch. has been used to treat respiratory, gastric, and liver diseases since ancient China. The most important and widely studied active component in G. uralensis is glycyrrhizin (GC). Our pervious RNA-Seq study shows that GC biosynthesis is regulated by multiple biosynthetic pathways. In this study, three target genes, ARPI, β-AS, and UGE from different pathways were selected and their regulatory effects on GC biosynthesis were investigated using G. uralensis hairy roots. Our data show that hairy roots knocking out ARPI or UGE died soon after induction, indicating that the genes are essential for the growth of G. uralensis hairy roots. Hairy roots with β-AS knocked out grew healthily. However, they failed to produce GC, suggesting that β-AS is required for triterpenoid skeleton formation. Conversely, overexpression of UGE or β-AS significantly increased the GC content, whereas overexpression of ARPI had no obvious effects on GC accumulation in G. uralensis hairy roots. Our findings demonstrate that β-AS and UGE positively regulate the biosynthesis of GC.