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Chinese goldthread ( Coptis chinensis Franch.), a member of the Ranunculales, represents an important early-diverging eudicot lineage with diverse medicinal applications. Here, we present a high-quality chromosome-scale genome assembly and annotation of C. chinensis . Phylogenetic and comparative genomic analyses reveal the phylogenetic placement of this species and identify a single round of ancient whole-genome duplication (WGD) shared by the Ranunculaceae. We characterize genes involved in the biosynthesis of protoberberine-type alkaloids in C. chinensis . In particular, local genomic tandem duplications contribute to member amplification of a Ranunculales clade-specific gene family of the cytochrome P450 (CYP) 719. The functional versatility of a key CYP719 gene that encodes the ( S )-canadine synthase enzyme involved in the berberine biosynthesis pathway may play critical roles in the diversification of other berberine-related alkaloids in C. chinensis . Our study provides insights into the genomic landscape of early-diverging eudicots and provides a valuable model genome for genetic and applied studies of Ranunculales. Coptis chinensis represents an early-diverging eudicot lineage with diverse medicinal applications. Here, the authors report its chromosome-scale genome assembly, infer a single ancient whole-genome duplication, and characterize the function of CYP719 in diversification of protoberberine-type alkaloids.

Coptis (Ranunculaceae) contains 15 species and is one of the pharmaceutically most important plant genera in eastern Asia. Understanding of the evolution of morphological characters and phylogenetic relationships within the genus is very limited. Here, we present the first comprehensive phylogenetic analysis of the genus based on two plastid and one nuclear markers. The phylogeny was reconstructed using Bayesian inference, as well as maximum parsimony and maximum likelihood methods. The Swofford-Olsen-Waddell-Hillis and Bayesian tests were used to assess the strength of the conflicts between traditional taxonomic units and those suggested by the phylogenetic inferences. Evolution of morphological characters was inferred using Bayesian method to identify synapomorphies for the infrageneric lineages. Our data recognize two strongly supported clades within Coptis. The first clade contains subgenus Coptis and section Japonocoptis of subgenus Metacoptis, supported by morphological characters, such as traits of the central leaflet base, petal color, and petal shape. The second clade consists of section Japonocoptis of subgenus Metacoptis. Coptis morii is not united with C. quinquefolia, in contrast with the view that C. morii is a synonym of C. quinquefolia. Two varieties of C. chinensis do not cluster together. Coptis groenlandica and C. lutescens are reduced to C. trifolia and C. japonica, respectively. Central leaflet base, sepal shape, and petal blade carry a strong phylogenetic signal in Coptis, while leaf type, sepal and petal color, and petal shape exhibit relatively higher levels of evolutionary flexibility.

Background The Plant-specific AP2/ERF gene family encodes proteins involved in various biological and physiological processes. Although the genome of Coptis chinensis Franch, a plant producing benzylisoquinoline alkaloids (BIAs), has been sequenced at the chromosome level, studies on the AP2/ERF gene family in C. chinensis are lacking. Thus, a genome-wide identification of AP2/ERF gene family in C. chinensis was conducted to explore its role in BIAs biosynthesis. Results A total of 96 CcAP2/ERF genes were identified and categorized into five subfamilies, including 43 ERFs, 32 DREBs, 17 AP2s, 3 RAVs, and 1 Soloist, based on their structural domains. These CcAP2/ERF genes were unevenly distributed across nine chromosomes. Analysis of gene duplication events identified 17 CcAP2/ERF gene pairs in the genome, with 7 involved in tandem duplication events and 10 involved in segmental duplicate events, indicating that both types of duplications contributed to the expansion of the AP2/ERF gene family. The Ka / Ks ratio analysis suggested that the CcAP2/ERF gene family underwent strong purifying selection. Two phytohormones, methyl jasmonate and abscisic acid, were identified as potential key inducers of BIAs biosynthesis due to the cis -acting element prediction. Analysis of the spatial transcriptomic data revealed that 28 differentially expressed AP2/ERF genes had the highest or relatively higher expression levels in the rhizome, 17 of which positively correlated with the tissue-specific accumulation of BIAs. Further real-time PCR verification and protein-protein interaction analysis indicated that DREB1B might be one of the central regulators in a highly complex BIAs biosynthesis network. Conclusion These findings provide significant insight into the function of AP2/ERF genes in C. chinensis , particularly in the regulatory network of BIAs biosynthesis in C. chinensis . This study also identifies candidate genes for metabolic engineering to increase BIAs content in C. chinensis .

Protoberberine alkaloids are a characteristic group of natural products in Coptis plants known for their notable pharmacological activities. However, the structural similarity and the substrate promiscuity of their biosynthetic enzymes have left the precise synthetic pathways remain unclarified, posing challenges to regulate product formation. In this study, we identified CcOMT8, a key enzyme responsible for C2‐methoxylation in the biosynthesis of epiberberine in C. chinensis, through methyl jasmonate elicitation analysis and comparative genomics‐based microsynteny analysis. Functional characterisation demonstrated that CcOMT8 specifically catalyses 2‐O‐methylation of (S)‐scoulerine, as verified by heterologous expression in both microbial and plant systems. Its lack of activity toward (S)‐cheilanthifoline further confirmed the specific route for epiberberine biosynthesis. Structural investigations of CcOMT8 and its complexes revealed key aspects of substrate recognition and a catalytic mechanism mediated by the His253‐Asp254‐Glu312 triad. Comparative structural analysis with 9‐O‐methyltransferases indicated that hydrophilic residues and reduced steric hindrance in the substrate binding pocket govern the regioselectivity of CcOMT8. Using focused rational iterative site‐specific mutagenesis (FRISM), we developed an optimised mutant, S109L/C250A/L300A, with 4.88‐fold enhanced catalytic efficiency. This study elucidates the biosynthetic pathway of epiberberine in Coptis, clarifies the molecular basis of enzyme‐directed metabolic flux, and provides efficient biocatalysts for the synthetic biosynthesis of protoberberine alkaloids.

Root rot disease poses a devastating threat to Coptis chinensis Franch , a medicinal plant prized for its bioactive alkaloids. To dissect its defense mechanisms, we conducted integrated transcriptomic and metabolomic analyses on resistant (R), early-stage infected (S-ES), and late-stage infected (S-LS) plants Our findings reveal a disease severity-dependent escalation in flavonoid metabolism. Key metabolites, such as kaempferol and quercetin derivatives, were significantly increased compared to R, paralleled by progressive upregulation of biosynthetic genes ( PAL , CHS , CHI , FLS ). Strikingly, salicylic acid (SA)-associated metabolites and pathway genes ( NPR1 , NPR3 / NPR4 ) showed no differential expression across groups, contrasting with typical SA-mediated defenses in other species. This study uncovers flavonoid biosynthesis as the primary defense strategy in C. chinensis during root rot progression, while SA signaling may not be the main defense mechanism. These results provide actionable targets for enhancing disease resistance in medicinal plants through metabolic engineering.

The medicinal plant Coptis chinensis is highly susceptible to root rot caused predominantly by Fusarium spp., threatening yield and medicinal quality. Biological control agents (BCAs) from the genus Bacillus are promising, yet their host mechanisms remain incompletely defined. We evaluated a four-strain Bacillus consortium as BCAs in field-grown C. chinensis and profiled host roots using transcriptomics and metabolomics. Pre-inoculation with the BCA mixture resulted in a modest, albeit statistically non-significant, reduction in disease index following Fusarium solani challenge. Multi-omics analyses showed that BCA treatment reprogrammed genes and metabolites associated with amino acid metabolism, accompanied by increased glutamine (Gln) accumulation. Supplementing culture medium with exogenous Gln inhibited F. solani growth in a concentration-dependent manner in vitro. Notably, F. solani or BCAs alone elicited modest regulation of canonical defense genes, whereas BCA pre-inoculation followed by F. solani robustly up-regulated defense modules across MAPK signaling, plant–pathogen interaction, and hormone signaling (JA, ET, SA), consistent with molecular immune priming. These results suggest that the BCA consortium induces resistance- and priming-related molecular markers in C. chinensis by modulating amino acid metabolism and host immunity. These findings provide a mechanistic basis for the potential application of BCAs in the sustainable management of C. chinensis root rot.

GRAS transcription factors are essential for plant growth and stress adaptation, yet they remain uncharacterized in the medicinal herb Coptis chinensis. To address this gap, we performed a genome-wide identification of the GRAS family and investigated its transcriptional responses to temperature and light stress, integrating comparative transcriptomics with promoter analysis to explore potential co-expression with bZIP factors. A total of 48 CcGRAS genes were identified and found to be unevenly distributed across nine chromosomes. Expression profiling revealed that CcGRAS genes are markedly more responsive to varying light intensities (476, 8340 lx) than to temperature stresses (15, 35 °C), relative to controls (2060 lx for light, 25 °C for temperature). Co-expression analysis uncovered an underground tissue-specific module in which CcbZIP16 is upregulated with four CcGRAS genes (CcGRAS11, CcGRAS12, CcGRAS43, CcGRAS48) that are coordinately upregulated specifically under low-light conditions. The promoters of these co-expressed genes are significantly enriched in canonical light-responsive cis-elements, providing correlative evidence for their coordinated transcriptional control. Together, these findings identify a tissue-specific GRAS-bZIP co-expressed gene set under light stress and suggest a candidate regulatory framework for dissecting light adaptation mechanisms. This work also provides a foundation for targeted genetic improvements in stress tolerance and alkaloid biosynthesis in this important medicinal plant.

Background MADS-box transcription factors have been shown to be involved in multiple developmental processes, including the regulation of floral organ formation and pollen maturation. However, the role of the MADS-box gene family in floral development of the alpine plant species Coptis teeta Wall, which is widely used in Traditional Chinese Medicine (TCM), is unknown. Results Sixty-six MADS-box genes were identified in the C. teeta genome. These genes were shown to be unevenly distributed throughout the genome of C. teeta . The majority of which (49) were classified as type I MADS-box genes and were further subdivided into four groups (Mα, Mβ, Mγ and Mδ). The remainder were identified as belonging to the type II MADS-box gene category. It was observed that four pairs of segmental and tandem duplication had occurred in the C. teeta MADS-box gene family, and that the ratios of Ka/Ks were less than 1, suggesting that these genes may have experienced purifying selection during evolution. Gene expression profiling analysis revealed that 38 MADS-box genes displayed differential expression patterns between the M and F floral phenotypes. Sixteen of these MADS-box genes were further verified by RT-qPCR. The 3D structure of each subfamily gene was predicted, further indicating that MADS-box genes of the same type possess structural similarities to the known template. Conclusions These data provide new insights into the molecular mechanism of dichogamy and herkogamy formation in C. teeta and establish a solid foundation for future studies of the MADS-box genes family in this medicinal plant species.

Coptis chinensis is a perennial herb of the Ranunculaceae family. The isoquinoline alkaloid is the main active component of C. chinensis, mainly exists in its rhizomes and has high clinical application potential. The in vitro synthesis of isoquinoline alkaloids is difficult because their structures are complex; hence, plants are still the main source of them. In this study, two-year and four-year rhizomes of C. chinensis were selected to investigate the effect of growth years on the accumulation of isoquinoline alkaloids. Two-year and four-year C. chinensis were selected for metabolomics detection and transcriptomic analysis. A total of 413 alkaloids were detected by metabolomics analysis, of which 92 were isoquinoline alkaloids. (S)-reticuline was a significantly different accumulated metabolite of the isoquinoline alkaloids biosynthetic pathway in C. chinensis between the two groups. The results of transcriptome analysis showed that a total of 464 differential genes were identified, 36 of which were associated with the isoquinoline alkaloid biosynthesis pathway of C. chinensis. Among them, 18 genes were correlated with the content of important isoquinoline alkaloids. Overall, this study provided a comprehensive metabolomic and transcriptomic analysis of the rapid growth stage of C. chinensis rhizome from the perspective of growth years. It brought new insights into the biosynthetic pathway of isoquinoline alkaloids and provided information for utilizing biotechnology to improve their contents in C. chinensis.

Key messageThe study carry out comprehensive transcriptome analysis of C. deltoidea and exploration of BIAs biosynthesis and accumulation based on UHPLC-MS/MS and combined sequencing platforms.Coptis deltoidea is an important medicinal plant with a long history of medicinal use, which is rich in benzylisoquinoline alkaloids (BIAs). In this study, Ultra performance liquid chromatography-electrospray ionization tandem mass spectrometry (UHPLC-ESI-MS/MS) and combined sequencing platforms were performed for exploration of BIAs biosynthesis, accumulation and comprehensive transcriptome analysis of C. deltoidea. By metabolism profiling, the accumulation of ten BIAs was analyzed using UHPLC-MS/MS and different contents were observed in different organs. From transcriptome sequencing result, we applied single-molecule real-time (SMRT) sequencing to C. deltoidea and generated a total of 75,438 full-length transcripts. We proposed the candidate biosynthetic pathway of tyrosine, precursor of BIAs, and identified 64 full length-transcripts encoding enzymes putatively involved in BIAs biosynthesis. RNA-Seq data indicated that the majority of genes exhibited relatively high expression level in roots. Transport of BIAs was also important for their accumulation. Here, 9 ABC transporters and 2 MATE transporters highly homologous to known alkaloid transporters related with BIAs transport in roots and rhizomes were identified. These findings based on the combined sequencing platforms provide valuable genetic information for C. deltoidea and the results of transcriptome combined with metabolome analysis can help us better understand BIAs biosynthesis and transport in this medicinal plant. The information will be critical for further characterization of C. deltoidea transcriptome and molecular-assisted breeding for this medicinal plant with scarce resources.