Panax quinquefolius is a globally valued medicinal plant rich in bioactive flavonoids, yet the molecular mechanisms underlying their biosynthesis remain poorly understood. In this study, we integrated transcriptomic and metabolomic analyses to investigate tissue-specific flavonoid accumulation and regulatory networks in roots, leaves, and flowers. Metabolomic profiling identified 141 flavonoid metabolites, with flavones, flavonols, and C-glycosylflavones predominantly enriched in aerial tissues (leaves and flowers), while specific glycosides like tricin 7-O-acetylglucoside showed root-specific accumulation. Transcriptome sequencing revealed 15,551–18,946 DEGs across tissues, and the reliability of the transcriptomic data was validated by qRT-PCR. KEGG and GO annotation analyses suggested that these DEGs may play a crucial role in the biosynthesis and metabolism of secondary metabolites. From the DEGs, UGTs and MYB TFs were identified and subjected to correlation analysis. Functional validation through in vitro enzymatic assays confirmed that PqUGT71A1 catalyzes apigenin and naringenin glycosylation at the 7-OH position. Additionally, subcellular localization and yeast one-hybrid assays demonstrated that PqMYB7 and PqMYB13 interact with the PqUGT71A1 promoter and activate its expression.. This study unveils the spatial dynamics of flavonoid metabolism in P. quinquefolius and establishes a MYB-UGT regulatory axis, providing critical insights for metabolic engineering and bioactive compound optimization in medicinal plants.

Auxin-responsive factors (ARFs) are an important class of transcription factors and are an important component of auxin signaling. This study conducted a genome-wide analysis of the ARF gene family in ginseng and presented its findings. Fifty-three ARF genes specific to ginseng (PgARF) were discovered after studying the ginseng genome. The coding sequence (CDS) has a length of 1092–4098 base pairs and codes for a protein sequence of 363–1565 amino acids. Among them, PgARF32 has the least number of exons (2), and PgARF16 has the most exons (18). These genes were then distributed into six subgroups based on the results obtained from phylogenetic analysis. In each subgroup, the majority of the PgARF genes displayed comparable intron/exon structures. PgARF genes are unevenly distributed on 20 chromosomes. Most PgARFs have B3 DNA binding, Auxin_resp, and PB1 domains. The PgARF promoter region contains various functional domains such as plant hormones, light signals, and developmental functions. Segmental duplications contribute to the expansion of the ARF gene family in ginseng, and the genes have undergone purifying selection during evolution. Transcriptomic results showed that some PgARFs had different expression patterns in different parts of ginseng; most PgARFs were affected by exogenous hormones, and a few PgARFs responded to environmental stress. It is suggested that PgARF is involved in the development of ginseng by regulating hormone-mediated genes. PgARF14, PgARF42, and PgARF53 are all situated in the nucleus, and both PgARR14 and PgARF53 noticeably enhance the growth length of roots in Arabidopsis. Our findings offer a theoretical and practical foundation for exploring PgARFs’ role in the growth of ginseng roots.

Panax quinquefolius L. is an important medicinal plant, and flavonoids are among its main secondary metabolites. The R2R3-MYB transcription factor plays an irreplaceable role in plant growth, development, and secondary metabolism. In our study, we identified 159 R2R3-MYBs and analyzed their physical and chemical properties in P. quinquefolius. The protein length of 159 PqMYBs varied from 107 to 1050 amino acids. The molecular weight ranged from 12.21 to 116.44 kDa. The isoelectric point was between 4.57 and 10.34. We constructed a phylogenetic tree of P. quinquefolius and Arabidopsis thaliana R2R3-MYB family members, and PqMYB members were divided into 33 subgroups. Transcriptome data analysis showed that the expression patterns of PqMYBs in root, leaf, and flower were significantly different. Following the MeJA treatment of seedlings, five candidate PqMYB genes demonstrated a response. A correlation analysis of PqMYBs and candidate flavonoid pathway genes showed that PqMYB2, PqMYB46, and PqMYB72 had correlation coefficients that were higher than 0.8 with PqCHS, PqANS4, and PqCCoAMT10, respectively. Furthermore, a transient expression assay confirmed that the three PqMYBs were localized in the nucleus. We speculated that these three PqMYBs were related to flavonoid biosynthesis in P. quinquefolius. These results provided a theoretical basis and a new perspective for further understanding the R2R3-MYB gene family and the biosynthesis mechanism of secondary metabolites in P. quinquefolius.

Perilla frutescens (L.) Britt. is a traditional medicinal and culinary plant with a long history of cultivation and significant potential for broader utilization. The basic helix-loop-helix (bHLH) gene family is essential for regulating plant growth, development, stress responses, and secondary metabolism. However, the bHLH gene family in P. frutescens has not yet been characterized. In this study, a total of 205 bHLH genes were identified in P. frutescens through genome mining and analysis. Phylogenetic analysis classified these PfbHLH genes into 23 distinct subfamilies. Promoter analysis revealed an enrichment of cis-acting elements linked to plant hormone signaling and stress responses, suggesting their potential regulatory roles in development, growth, and stress adaptation. Expression profiling based on publicly available RNA-seq data demonstrated tissue-specific expression patterns of PfbHLH genes in roots, stems, and leaves. Four PfbHLH genes (PfbHLH66, PfbHLH45, PfbHLH13, and PfbHLH5) showed significant responses to methyl jasmonate (MeJA) induction. Yeast one-hybrid assays confirmed that these PfbHLH proteins could bind to the cis-acting G-box (CACGTG) element. This study offers new perspectives on the evolution, regulatory mechanisms, and functional roles of the bHLH gene family in P. frutescens. The findings deepen our understanding of the functional diversity within this gene family and establish a foundation for genetic enhancement and the biosynthesis of medicinal compounds in the species.

Lobetyolin is a key bioactive compound in Codonopsis pilosula, but the existing literature has not well explored the regulatory mechanisms governing its biosynthesis. This study performed a genome-wide identification of 175 AP2/ERF transcription factors (TFs), categorizing them into the AP2, ERF, and RAV subfamilies. Promoter analysis revealed numerous hormone- and stress-responsive cis-elements, with a notable prevalence of methyl jasmonate (MeJA)-related motifs. In MeJA-treated hairy root cultures, we observed a significant increase in lobetyolin accumulation. Integrated transcriptomic and metabolomic analyses demonstrated the activation of jasmonate signaling and secondary metabolism pathways. Two MeJA-induced TFs, CpERF140 and CpERF138, were identified as key regulatory candidates. Subsequent experiments confirmed their nuclear localization and specific binding affinity for GCC-box motifs. This study constitutes the inaugural genome-wide characterization of the AP2/ERF family in C. pilosula, employing hairy root culture as an effective platform for functional investigation. Our findings offer novel perspectives on the transcriptional regulation of lobetyolin biosynthesis, providing a foundation for the future metabolic engineering of this medicinal plant.

Perilla frutescens (L.) Britt., a plant rich in phenolic acids, contains rosmarinic acid (RA) as its main phenolic component. RA exhibits various pharmacological activities, including anti-allergic, anti-inflammatory, and antioxidant effects. In this study, we established hairy root cultures of P. frutescens using Agrobacterium rhizogenes strains C58C1, A4, and R1000, and assessed phenolic acid content. Among the strains, C58C1 yielded the highest number of rooted explants and the highest induction rate, with the RA content reaching 19.08 mg/g DW in the fastest-growing lines. We also investigated the effects of salicylic acid (SA) and methyl jasmonate (MeJA) on phenolic acid accumulation in hairy roots. SA treatment promoted RA accumulation, reaching 20.92 mg/g DW on day nine, while MeJA increased RA to 17.36 mg/g DW on day three, followed by a decline to 4.27 mg/g DW by day nine. Furthermore, transgenic analysis of PfbHLH13 and PfbHLH66 transcription factors showed that overexpression of these genes significantly increased RA content. Quantitative real-time PCR revealed that PfbHLH13 enhances phenolic acid accumulation by regulating the phenylpropanoid pathway, while PfbHLH66 influences both the phenylpropanoid and tyrosine-derived pathways. These findings highlight the potential of P. frutescens hairy roots for large-scale RA production.