Comprehensive genome-wide identification of PgCYP450 genes and functional characterization of PgCYP71BE217 in saponin biosynthesis of Platycodon grandiflorus

Platycodon grandiflorus (Jacq.) A. DC., a well-known traditional Chinese medicinal herb, is particularly abundant in saponins. The cytochrome P450 monooxygenase (CYP450) superfamily constitutes the key group of enzymes responsible for regulating saponin biosynthesis. To date, a comprehensive analysis of CYP450 genes associated with saponin biosynthesis in P. grandiflorus has not been conducted. In the present study, 175 PgCYP450 genes were systematically identified in P. grandiflorus. Phylogenetic analysis classified these genes into nine distinct clans. Transcriptome (RNA-seq) data further demonstrated that the majority of genes within the CYP71 family were upregulated in response to methyl jasmonate (MeJA) treatment. qRT-PCR revealed that PgCYP71BE217 had the highest expression level after MeJA induction, which was over 60 times higher than the control, and displayed elevated expression primarily in the roots. Subcellular localization and yeast self-activation experiments indicated that PgCYP71BE217 was localized to both the plasma membrane and cytoplasm, and lacked self-activation activity. Overexpression of PgCYP71BE217 using a hairy root transformation system enhanced the transcript levels of PgHMGS (3-hydroxy-3-methylglutaryl-CoA synthase), PgbAS1 (β-amyrin synthase 1), and PgSS (squalene synthase), while simultaneously elevating the accumulation of saponins such as platycodin D, platycoside E, and platycodin D3. The identification and functional analysis of the CYP450 gene family in this study enhances understanding of saponin synthesis and provides valuable genetic resources for P. grandiflorus breeding.