Isoquinoline Alkaloid Biosynthesis Research Articles

MicroRNAs Participate in Morphological Acclimation of Sugar Beet Roots to Nitrogen Deficiency.

Nitrogen (N) is essential for sugar beet (Beta vulgaris L.), a highly N-demanding sugar crop. This study investigated the morphological, subcellular, and microRNA-regulated responses of sugar beet roots to low N (LN) stress (0.5 mmol/L N) to better understand the N perception, uptake, and utilization in this species. The results showed that LN led to decreased dry weight of roots, N accumulation, and N dry matter production efficiency, along with damage to cell walls and membranes and a reduction in organelle numbers (particularly mitochondria). Meanwhile, there was an increase in root length (7.2%) and branch numbers (29.2%) and a decrease in root surface area (6.14%) and root volume (6.23%) in sugar beet after 7 d of LN exposure compared to the control (5 mmol/L N). Transcriptomics analysis was confirmed by qRT-PCR for 6 randomly selected microRNAs, and we identified 22 differentially expressed microRNAs (DEMs) in beet root under LN treatment. They were primarily enriched in functions related to binding (1125), ion binding (641), intracellular (437) and intracellular parts (428), and organelles (350) and associated with starch and sucrose metabolism, tyrosine metabolism, pyrimidine metabolism, amino sugar and nucleotide sugar metabolism, and isoquinoline alkaloid biosynthesis, as indicated by the GO and KEGG analyses. Among them, the upregulated miR156a, with conserved sequences, was identified as a key DEM that potentially targets and regulates squamosa promoter-binding-like proteins (SPLs, 104889216 and 104897537) through the microRNA-mRNA network. Overexpression of miR156a (MIR) promoted root growth in transgenic Arabidopsis, increasing the length, surface area, and volume. In contrast, silencing miR156a (STTM) had the opposite effect. Notably, the fresh root weight decreased by 45.6% in STTM lines, while it increased by 27.4% in MIR lines, compared to the wild type (WT). It can be inferred that microRNAs, especially miR156, play crucial roles in sugar beet root's development and acclimation to LN conditions. They likely facilitate active responses to N deficiency through network regulation, enabling beet roots to take up nutrients from the environment and sustain their vital life processes.

Molecular Insights into the Defense of Dioscorea opposita Cultivar Tiegun Callus Against Pathogenic and Endophytic Fungal Infection Through Transcriptome Analysis.

Dioscorea opposita cultivar Tiegun is an economically important crop with high nutritional and medicinal value. Plants can activate complex and diverse defense mechanisms after infection by pathogenic fungi. Moreover, endophytic fungi can also trigger the plant immune system to resist pathogen invasion. However, the study of the effects of endophytic fungi on plant infection lags far behind that of pathogenic fungi, and the underlying mechanism is not fully understood. Here, the black spot pathogen Alternaria alternata and the endophytic fungus Penicillium halotolerans of Tiegun were identified and used to infect calli. The results showed that A. alternata could cause more severe membrane lipid peroxidation, whereas P. halotolerans could rapidly increase the activity of the plant antioxidant enzymes superoxide dismutase, peroxidase, and catalase; thus, the degree of damage to the callus caused by P. halotolerans was weaker than that caused by A. alternata. RNA sequencing analysis revealed that various plant defense pathways, such as phenylpropanoid biosynthesis, flavonoid biosynthesis, plant hormone signal transduction, and the mitogen-activated protein kinase signaling pathway, play important roles in triggering the plant immune response during fungal infection. Furthermore, the tryptophan metabolism, betalain biosynthesis, fatty acid degradation, flavonoid biosynthesis, tyrosine metabolism, and isoquinoline alkaloid biosynthesis pathways may accelerate the infection of pathogenic fungi, and the ribosome biogenesis pathway in eukaryotes may retard the damage caused by endophytic fungi. This study lays a foundation for exploring the infection mechanism of yam pathogens and endophytic fungi and provides insight for effective fungal disease control in agriculture.

A genome assembly of decaploid Houttuynia cordata provides insights into the evolution of Houttuynia and the biosynthesis of alkaloids.

Houttuynia cordata Thunb., commonly known as yuxingcao in China, is known for its characteristic fishy smell and is widely recognized as an important herb and vegetable in many parts of Asia. However, the lack of genomic information on H. cordata limits the understanding of its population structure, genetic diversity, and biosynthesis of medicinal compounds. Here we used single-molecule sequencing, Illumina paired-end sequencing, and chromosome conformation capture technology to construct the first chromosome-scale decaploid H. cordata reference genome. The genome assembly was 2.63Gb in size, with 1348 contigs and a contig N50 of 21.94Mb further clustered and ordered into 88 pseudochromosomes based on Hi-C analysis. The results of genome evolution analysis showed that H. cordata underwent a whole-genome duplication (WGD) event ~17 million years ago, and an additional WGD event occurred 3.3 million years ago, which may be the main factor leading to the high abundance of multiple copies of orthologous genes. Here, transcriptome sequencing across five different tissues revealed significant expansion and distinct expression patterns of key gene families, such as l-amino acid/l-tryptophan decarboxylase and strictosidine synthase, which are essential for the biosynthesis of isoquinoline and indole alkaloids, along with the identification of genes such as TTM3, which is critical for root development. This study constructed the first decaploid medicinal plant genome and revealed the genome evolution and polyploidization events of H. cordata .

Metabolome and transcriptome integration explored the mechanism of browning in Glycyrrhiza uralensis Fisch cells.

Glycyrrhiza uralensis Fisch, a traditional Chinese medicinal herb known for its diverse pharmacological effects including heat-clearing, detoxification, phlegm dissolving, and cough relief, has experienced an exponential increase in demand due to its expanding clinical use and development prospects. Currently, large-scale cell culture stands out as one of the most promising biotechnological approaches for producing bioactive compounds from medicinal plants. However, the problem of cell browning represents a significant bottleneck in industrial applications of cell culture. This study focuses on the Glycyrrhiza uralensis Fisch cells from the Ordos plateau, aiming to elucidate the enzymatic browning process during plant cell culture. Key substrates and genes involved in enzymatic browning were identified by metabolome and transcriptome analysis of normal and browning cells. Metabolome analysis reveals significant changes in the levels of chalcone, isoflavone, imidazole-pyrimidine, purine nucleosides, organic oxides, carboxylic acids and their derivatives, benzene and its derivatives, flavonoids, 2-arylated benzofuran flavonoids, diazanaphthalenes and fatty acyls within browning cells. In particular, chalcones, isoflavones, and flavones compounds account for a higher proportion of these changes. Furthermore, these compounds collectively show enrichment in four metabolic pathways: Isoflavone biosynthesis pathway; Cutin suberine and wax biosynthesis pathway; Aminoacyl-tRNA biosynthesis pathway; Isoquinoline alkaloid biosynthesis pathway; Transcriptome analysis revealed that the MYB transcription factor is a key regulator of flavonoid synthesis during the browning process in cells. In addition, 223 differentially expressed genes were identified, including phenylpropane, shikimic acid, glycolysis, and pentose phosphate pathways. Among these genes, 23 are directly involved in flavonoid biosynthesis; qPCR validation showed that eight genes (GlPK, GlPAL, Gl24CL, Gl1PDT, Gl3CHI, GlC4H, Gl2F3'H, and Gl2CCR) were up-regulated in browning cells compared to normal cells. These findings corroborate the sequencing results and underscore the critical role of these genes in cellular browning. Consequently, modulation of their expression offers promising strategies for effective control of cellular browning issues.

Whole-Genome Sequencing of Two Potentially Allelopathic Strains of Bacillus from the Roots of C. equisetifolia and Identification of Genes Related to Synthesis of Secondary Metabolites.

The coastal Casuarina equisetifolia is the most common tree species in Hainan's coastal protection forests. Sequencing the genomes of its allelopathic endophytes can allow the protective effects of these bacteria to be effectively implemented in protected forests. The goal of this study was to sequence the whole genomes of the endophytes Bacillus amyloliquefaciens and Bacillus aryabhattai isolated from C. equisetifolia root tissues. The results showed that the genome sizes of B. amyloliquefaciens and B. aryabhattai were 3.854 Mb and 5.508 Mb, respectively. The two strains shared 2514 common gene families while having 1055 and 2406 distinct gene families, respectively. The two strains had 283 and 298 allelochemical synthesis-associated genes, respectively, 255 of which were shared by both strains and 28 and 43 of which were unique to each strain, respectively. The genes were putatively involved in 11 functional pathways, including secondary metabolite biosynthesis, terpene carbon skeleton biosynthesis, biosynthesis of ubiquinone and other terpene quinones, tropane/piperidine and piperidine alkaloids biosynthesis, and phenylpropanoid biosynthesis. NQO1 and entC are known to be involved in the biosynthesis of ubiquinone and other terpenoid quinones, and rfbC/rmlC, rfbA/rmlA/rffH, and rfbB/rmlB/rffG are involved in the biosynthesis of polyketide glycan units. Among the B. aryabhattai-specific allelochemical synthesis-related genes, STE24 is involved in terpene carbon skeleton production, atzF and gdhA in arginine biosynthesis, and TYR in isoquinoline alkaloid biosynthesis. B. amyloliquefaciens and B. aryabhattai share the genes aspB, yhdR, trpA, trpB, and GGPS, which are known to be involved in the synthesis of carotenoids, indole, momilactones, and other allelochemicals. Additionally, these bacteria are involved in allelochemical synthesis via routes such as polyketide sugar unit biosynthesis and isoquinoline alkaloid biosynthesis. This study sheds light on the genetic basis of allelopathy in Bacillus strains associated with C. equisetifolia, highlighting the possible use of these bacteria in sustainable agricultural strategies for weed management and crop protection.

Deep integration of metabolome and transcriptome characterizes alkaloid metabolism in Houttuynia cordata

Alkaloids are the main medicinal components in Houttuynia cordata. In this study, two accessions 6# and 7# of H. cordata underwent thorough metabolomic analyses to identify and quantify alkaloid phytometabolites. It turned out that the alkaloid types were largely similar between 6# and 7#, and the identified 81 alkaloids could be divided into nine structural classes. However, the content of alkaloids in the two accessions was quite different. According to transcriptome data, a total of 114 differentially expressed genes related to alkaloid metabolism were screened. The alkaloid synthesis pathway of the two varieties was mainly different in the isoquinoline alkaloid biosynthesis and indole alkaloid biosynthesis; four genes A22110063c_transcript_59323, A22110063c_transcript_60118, A22110063c_transcript_51672 and A22110063c_transcript_48784 were highly expressed in 7#, which could be key candidate genes of alkaloid metabolism and warrant further analysis. These results provide a reference for the medicinal application of H. cordata and breeding alkaloid rich varieties.

Screening and Genomic Analysis of Alkaloid-Producing Endophytic Fungus Fusarium solani Strain MC503 from Macleaya cordata.

The extensive harvesting of Macleaya cordata, as a biomedicinal plant and a wild source of quaternary benzo[c]phenanthridine alkaloids, has led to a rapid decline in its population. An alternative approach to the production of these bioactive compounds, which are known for their diverse pharmacological effects, is needed. Production of these compounds using alkaloid-producing endophytic fungi is a promising potential approach. In this research, we isolated an alkaloid-producing endophytic fungus, strain MC503, from the roots of Macleaya cordata. Genomic analysis was conducted to elucidate its metabolic pathways and identify the potential genes responsible for alkaloid biosynthesis. High-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) analyses revealed the presence and quantified the content of sanguinarine (536.87 μg/L) and chelerythrine (393.31 μg/L) in the fungal fermentation extract. Based on our analysis of the morphological and micromorphological characteristics and the ITS region of the nuclear ribosomal DNA of the alkaloid-producing endophyte, it was identified as Fusarium solani strain MC503. To the best of our knowledge, there is no existing report on Fusarium solani from Macleaya cordata or other medicinal plants that produce sanguinarine and chelerythrine simultaneously. These findings provide valuable insights into the capability of Fusarium solani to carry out isoquinoline alkaloid biosynthesis and lay the foundation for further exploration of its potential applications in pharmaceuticals.

Salicylic acid modulates secondary metabolism and enhanced colchicine accumulation in long yellow daylily (Hemerocallis citrina).

Salicylic acid (SA) is an essential phytoregulator that is widely used to promote the synthesis of high-value nutraceuticals in plants. However, its application in daylily, an ornamental plant highly valued in traditional Chinese medicine, has not been reported. Herein, we investigated the exogenous SA-induced physiological, transcriptional and biochemical changes in long yellow daylily (LYD). We found that 2mg/L foliar SA treatment significantly improved LYD plant growth and yield. Transcriptome sequencing and differentially expressed genes (DEGs) analysis revealed that the phenylpropanoid biosynthesis, isoquinoline alkaloid biosynthesis, sulfur metabolism, plant hormone signal transduction and tyrosine metabolism were significantly induced in SA-treated leaves. Many transcription factors and antioxidant system-related DEGs were induced under the SA treatment. Biochemical analyses showed that the leaf contents of soluble sugar, soluble protein (Cpr), ascorbic acid (AsA) and colchicine were significantly increased by 15.15% (from 30.16 ± 1.301 to 34.73 ± 0.861mg/g), 19.54% (from 60.3 ± 2.227 to 72.08 ± 1.617mg/g), 30.45% (from 190.1 ± 4.56 to 247.98 ± 11.652 μg/g) and 73.05% (from 3.08 ± 0.157 to 5.33 ± 0.462 μg/g), respectively, under the SA treatment. Furthermore, we identified 15 potential candidate genes for enhancing the growth, production and phytochemical content of LYD. Our results provide support for the bioaccumulation of colchicine in yellow daylily and valuable resources for biotechnological-assisted production of this important nutraceutical in Hemerocallis spp.

Integrated multi-omics investigation revealed the importance of phenylpropanoid metabolism in the defense response of Lilium regale Wilson to fusarium wilt.

Lilies (genus Lilium) play a significant role in the global cut-flower industry, but they are highly susceptible to fusarium wilt caused by Fusarium oxysporum. However, Lilium regale, a wild lily species, exhibits remarkable resistance to F. oxysporum. To investigate the quantitative resistance of L. regale to fusarium wilt, a comprehensive multi-omics analysis was conducted. Upon inoculation with F. oxysporum, L. regale roots showed a significant accumulation of phenylpropane metabolites, including lignin precursors, flavonoids, and hydroxycinnamic acids. These findings were consistent with the upregulated expression of phenylpropanoid biosynthesis-related genes encoding various enzymes, as revealed by transcriptomics and proteomics analyses. Furthermore, metabolomics and proteomics data demonstrated differential activation of monoterpenoid and isoquinoline alkaloid biosynthesis. Colorimetry and high-performance liquid chromatography analyses revealed significantly higher levels of total flavonoids, lignin, ferulic acid, phlorizin, and quercetin contents in L. regale scales compared with susceptible lily 'Siberia' scales during F. oxysporum infection. These phenylpropanes exhibited inhibitory effects on F. oxysporum growth and suppressed the expression of pathogenicity-related genes. Transcriptional regulatory network analysis suggested that ethylene-responsive transcription factors (ERFs) may positively regulate phenylpropanoid biosynthesis. Therefore, LrERF4 was cloned and transiently overexpressed in the fusarium wilt-susceptible Oriental hybrid lily 'Siberia'. The overexpression of LrERF4 resulted in increased levels of total flavonoids, lignin, ferulic acid, phlorizin, and quercetin, while the silencing of LrERF4 in L. regale through RNAi had the opposite effect. In conclusion, phenylpropanoid metabolism plays a crucial role in the defense response of L. regale against fusarium wilt, with LrERF4 acting as a positive regulator of phenylpropane biosynthesis.

Integrated metabolomic and transcriptomic analyses provide insights into regulation mechanisms during bulbous stem development in the Chinese medicinal herb plant, Stephania kwangsiensis

BackgroundStephania kwangsiensis Lo (Menispermaceae) is a well-known Chinese herbal medicine, and its bulbous stems are used medicinally. The storage stem of S. kwangsiensis originated from the hypocotyls. To date, there are no reports on the growth and development of S. kwangsiensis storage stems.ResultsThe bulbous stem of S. kwangsiensis, the starch diameter was larger at the stable expanding stage (S3T) than at the unexpanded stage (S1T) or the rapidly expanding stage (S2T) at the three different time points. We used ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and Illumina sequencing to identify key genes involved in bulbous stem development. A large number of differentially accumulated metabolites (DAMs) and differentially expressed genes (DEGs) were identified. Based on the differential expression profiles of the metabolites, alkaloids, lipids, and phenolic acids were the top three differentially expressed classes. Compared with S2T, significant changes in plant signal transduction and isoquinoline alkaloid biosynthesis pathways occurred at both the transcriptional and metabolic levels in S1T. In S2T compared with S3T, several metabolites involved in tyrosine metabolism were decreased. Temporal analysis of S1T to S3T indicated the downregulation of phenylpropanoid biosynthesis, including lignin biosynthesis. The annotation of key pathways showed an up-down trend for genes and metabolites involved in isoquinoline alkaloid biosynthesis, whereas phenylpropanoid biosynthesis was not completely consistent.ConclusionsDownregulation of the phenylpropanoid biosynthesis pathway may be the result of carbon flow into alkaloid synthesis and storage of lipids and starch during the development of S. kwangsiensis bulbous stems. A decrease in the number of metabolites involved in tyrosine metabolism may also lead to a decrease in the upstream substrates of phenylpropane biosynthesis. Downregulation of lignin synthesis during phenylpropanoid biosynthesis may loosen restrictions on bulbous stem expansion. This study provides the first comprehensive analysis of the metabolome and transcriptome profiles of S. kwangsiensis bulbous stems. These data provide guidance for the cultivation, breeding, and harvesting of S. kwangsiensis.

Integrative leaf anatomy structure, physiology, and metabolome analyses revealed the response to drought stress in sainfoin at the seedling stage.

Sainfoin (Onobrychis viciaefolia) is a vital legume forage, and drought is the primary element impeding sainfoin growth. The anatomical structure, physiological indexes, and metabolites of the leaves of sainfoin seedlings with a drought-resistant line of P1 (DRL) and a drought-sensitive material of 2049 (DSM) were analyzed under drought (-1.0 MPa) with polyethylene glycol-6000 (PEG-6000). The leaf anatomy was studied by the paraffin section method. The related physiological indexes were measured by the hydroxylamine oxidation method, titanium sulfate colorimetric method, thiobarbituric acid method, acidic ninhydrin colorimetric method, and Coomassie brilliant blue method. The metabolomics analysis was composed of liquid chromatography tandem high-resolution mass spectrometry (LC-MS/MS). The results revealed that the thickness of the epidermis, palisade tissue, and sponge tissue of DRL were significantly greater than those of DSM. The leaves of DRL exhibited lower levels of superoxide anion (O2 •-) production rate, hydrogen peroxide (H2O2) content, and malondialdehyde (MDA) content compared with DSM, while proline (Pro) content and soluble protein (SP) content were significantly higher than those of DSM. A total of 391 differential metabolites were identified in two samples. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment showed that the primary differential metabolites were concentrated into the tyrosine metabolism; isoquinoline alkaloid biosynthesis; ubiquinone and other terpenoid quinone biosynthesis; neomycin, kanamycin, and gentamicin biosynthesis; and anthocyanin biosynthesis metabolic pathways. Compared with DSM, DRL had more complete anatomical structure, lower active oxygen content, and higher antioxidant level. The results improved our insights into the drought-resistant mechanisms in sainfoin.

Untargeted metabolomic analysis of the metabolites in roots of Pugionium cornutum seedlings under drought stress.

Pugionium cornutum is an annual or biennial xerophyte distributed in arid regions, with drought resistance properties. While previous studies have predominantly focused on the physiological changes of P. cornutum , the understanding of its metabolite variations remains limited. In this study, untargeted metabolomic technology was performed to analyse the change of metabolites in the roots of P. cornutum seedlings under drought stress. Our findings revealed that compared to the R1, the root water potential and the number of lateral roots increased, while the length of the tap root and fresh weight increased first and then decreased. In the R1-R2, a total of 45 differential metabolites (DMs) were identified, whereas in the R1-R3 82 DMs were observed. Subsequently, KEGG analysis revealed a significant enrichment of microbial metabolism in diverse environments and aminobenzoate degradation in the R1-R2, and phenylpropanoid biosynthesis, ubiquinone, and other terpenoid-quinone biosynthesis and isoquinoline alkaloid biosynthesis were significantly enriched in the R1-R3. The upregulation DMs, including L-arginosuccinate, L-tyrosine, p-coumarate, caffeate, ferulate, vanillin, coniferin, 5-aminopentanoate, 2-methylmaleate and 2-furoate in P. cornutum seedlings may play a crucial role in enhancing root growth and improving drought resistance. These findings provide a basis for future investigations into the underlying mechanisms of drought resistance in P. cornutum .

Transcriptome analysis reveals the potential mechanism of altering viability, yield, and isoquinoline alkaloids in Coptis chinensis through Cunninghamia lanceolata understory cultivation

The dried rhizomes of Coptis chinensis hold significance in Chinese medicine. Monocropping C. chinensis under the shade of a manmade scaffold, the primary planting mode, poses a threat to the ecological balance. In recent years, the practice of C. chinensis–Cunninghamia lanceolata understory cultivation has gained prevalence in southwest China. However, there is no evidence to suggest that understory cultivation enhances the viability, yield, or isoquinoline alkaloid content of C. chinensis. This study examined the physiological properties, yield indicators, and isoquinoline alkaloid content to investigate variations in C. chinensis in response to understory cultivation. Transcriptome analysis was conducted to elucidate potential mechanisms driving these alterations. The results indicate that understory cultivation significantly enhances the viability, yield, and levels of epiberberine, palmatine, and berberine in C. chinensis while reducing coptisine content. Transcriptomic analyses identified 2062 upregulated and 1853 downregulated genes in the understory cultivation system. Pathways such as “phenylpropanoid biosynthesis,” “zeatin biosynthesis,” “photosynthesis,” “tyrosine metabolism,” “isoquinoline alkaloid biosynthesis,” and “starch and sucrose metabolism” exhibited significant enrichment of differentially expressed genes (DEGs). DEGs involved in these pathways were thoroughly analyzed. INV, BGL-2, BGL-4, SPS-2, AMY-3, Psb B, Psb R, Psb S, Psa D, Psa E, Psa H, Psa O, Pet C, Pet H, deta, and b exhibited significant positive correlations with plant fresh weight, aboveground fresh weight, and underground fresh weight. 6-OMT-2 and COMT1-3 displayed significant positive correlations with coptisine content, but negative correlations with epiberberine, palmatine, and berberine content. ZOG-1, ZOG-3, TAT, PPO, POD-13 and UGT 73C5-1 showed noteworthy positive correlations with berberine content. Conversely, MIFH, POD-4, POD-5, and POD-8 displayed significant positive correlations with epiberberine, palmatine, and berberine content. POD-5, and POD-7 were significantly negatively correlated with coptisine content. Furthermore, gene expression levels determined by qRT–PCR aligned with the transcriptomic sequencing results, confirming the reliability of the transcriptomic findings. The results of this study contribute evidence elucidating potential mechanisms through which C. chinensis responds to understory cultivation.Graphical

Genome sequencing and functional analysis of a multipurpose medicinal herb Tinospora cordifolia (Giloy).

Tinospora cordifolia (Willd.) Hook.f. & Thomson, also known as Giloy, is among the most important medicinal plants that have numerous therapeutic applications in human health due to the production of a diverse array of secondary metabolites. To gain genomic insights into the medicinal properties of T. cordifolia, the genome sequencing was carried out using 10× Genomics linked read and Nanopore long-read technologies. The draft genome assembly of T. cordifolia was comprised of 1.01 Gbp, which is the genome sequenced from the plant family Menispermaceae. We also performed the genome size estimation for T. cordifolia, which was found to be 1.13 Gbp. The deep sequencing of transcriptome from the leaf tissue was also performed. The genome and transcriptome assemblies were used to construct the gene set, resulting in 17,245 coding gene sequences. Further, the phylogenetic position of T. cordifolia was also positioned as basal eudicot by constructing a genome-wide phylogenetic tree using multiple species. Further, a comprehensive comparative evolutionary analysis of gene families contraction/expansion and multiple signatures of adaptive evolution was performed. The genes involved in benzyl iso-quinoline alkaloid, terpenoid, lignin and flavonoid biosynthesis pathways were found with signatures of adaptive evolution. These evolutionary adaptations in genes provide genomic insights into the presence of diverse medicinal properties of this plant. The genes involved in the common symbiosis signalling pathway associated with endosymbiosis (Arbuscular Mycorrhiza) were found to be adaptively evolved. The genes involved in adventitious root formation, peroxisome biogenesis, biosynthesis of phytohormones, and tolerance against abiotic and biotic stresses were also found to be adaptively evolved in T. cordifolia.

Alkaloids and phenolic constituents from Glaucium corniculatum

The aerial parts of Glaucium corniculatum were phytochemically investigated. Consequently, 14 compounds, including six alkaloids (1–6) namely dehydroberberine (1), berberine (2), protpine (3), allocryptopine (4), (+)-corydine (5) and glaucine (6); six flavonoids (7–12) including quercetin (7), isorhamnetin (8), rutin (9), isorhamnetin 3-O-rutinoside (narcissoside, narcissin) (10), quercetin-3-O-glucoside-7-O-rutinoside (11), and isorhamnetin 3-O-glucoside-7-O-rutinoside (12); and hydroxycinnamic acid malate esters, (+)-phaselic acid ((+)-2-O-caffeoyl-L-malate) (13), and (+)-2-O-feruloyl-L-malate (14) were identified. Notably, the protoberberine-type isoquinoline alkaloid dehydroberberine (1) was naturally isolated for the first time, and the two flavonoids triosides (11 and 12) were identified for the first time from genus Glaucium and family Papaveraceae. Their structures were assigned through 1D, 2D NMR, HR-ESI-MS experiments, and by comparing the obtained data with those reported in the literature. The identified metabolites in G. corniculatum have important chemotaxonomic implications, suggesting that the biosynthesis of isoquinoline alkaloids is influenced by the plant's geographical location and could serve as valuable chemotaxonomic markers for identifying the plant sources. Furthermore, flavonoids and phenolic acids can be utilized to differentiate between subfamilies within the family Papaveraceae or to distinguish it from closely related families.

Transcriptome analysis reveals cadmium exposure enhanced the isoquinoline alkaloid biosynthesis and disease resistance in Coptis chinensis

Coptis chinensis Franch is a perennial herb from the Ranunculaceae family with a long history of medicinal use. As the medicinal part, the rhizome of coptis often accumulates excessive cadmium (Cd) even at low concentrations in the soil, which not only compromises its medicinal safety but also raises concerns about adverse effects on human health. Therefore, effective strategies are needed to mitigate this accumulation and ensure its safe use in traditional medicine. This study utilized transcriptome profiling and physiological analysis to explore molecular mechanisms associated with ecological significance and the active accumulation of Cd in C. chinensis. The response to Cd in C. chinensis was assessed through RNA sequencing, Cd determination and isoquinoline alkaloid measurement using its roots, stems, and leaves. The transcriptome revealed, a total of 2667, 2998, or 2815 up-regulated deferentially expressed genes in roots, stems or leaves in response to Cd exposure. Furthermore, we identified phenylpropanoid and isoquinoline alkaloid biosynthesis as the key pathways response to Cd exposure, which suggests that C. chinensis may improve its tolerance to Cd through regulating the phenylpropanoid biosynthesis pathway. Under Cd exposure, plant-pathogen interaction in leaves was identified as the key pathway, which indicates that upregulation of genes involved in plant-pathogen interaction could enhance disease resistance in C. chinensis. WGCNA analysis identified WRKY8 (Cluster-55763.31419) and WRKY47 (Cluster-55763.221590) as potential regulators of secondary metabolic synthesis and plant-pathogen interaction pathway in C. chinensis triggered by Cd. The measurement of berberine, coptisine, palmatine, and epiberberine also demonstrated that Cd simulated the four isoquinoline alkaloids in roots. Therefore, our study not only presented a transcriptome expression profiles that revealed significant upregulation of genes involved in metal transport and detoxification pathways but also suggested a possible mechanism to cope with Cd accumulation. This knowledge provides a new insight into gene manipulation for controlling Cd accumulation, enhancing resistance and promoting synthesis of secondary metabolites with potential medicinal properties in other medicinal plant species.

Mycorrhiza-triggered changes in leaf food quality and secondary metabolite profile in tea at low temperatures

Tea is a popular cash crop around the world that often endures low temperatures in the spring, thereby negatively affecting tea yield and quality. The objective of this study was to examine the effects of an arbuscular mycorrhizal fungus, Clariodeoglomus etunicatum, on plant growth, root morphology, leaf food quality, associated gene expression, and secondary metabolite profiles in tea (Camellia sinensis cv. Fuding Dabaicha) seedlings grown at low temperatures. A seven-day low temperature (4 °C) treatment significantly reduced root colonization rate by C. etunicatum by 21.08%. Uninoculated plants' growth was considerably impeded by low temperatures, whereas C. etunicatum inoculation significantly increased leaf number and leaf, stem and root biomass at low temperatures. Low temperatures had little effect on root morphology, whereas C. etunicatum inoculation significantly increased root surface area at low temperatures. In addition, low temperatures reduced leaf quality variables, whereas C. etunicatum significantly increased the levels of glucose, fructose, sucrose, tea polyphenols, catechin, free amino acids, and caffeine at low temperatures, along with up-regulated expression of CsGDH, CsGOGAT, and CsHMGR, as well as down-regulated expression of CsGs. Non-targeted metabolomics identified 767 differential metabolites, of which 65 differential metabolites were observed at low temperatures by C. etunicatum inoculation, including 42 (e.g., catechin, (−)-epicatechingallate, protoanemonin, epicatechin, and 4-dodecylbenzenesulfonic acid) up-regulated and 23 (e.g., 4-hydroxybenzoic acid, raffinose, L-theanine, melezitose, and D-serine) down-regulated. Mycorrhiza-triggered differential metabolites enriched 268 metabolic pathways at low temperatures, including isoquinoline alkaloid biosynthesis, lysine biosynthesis, tyrosine metabolism, and others. These findings reveal that C. etunicatum can function as a fungal stimulant to improve the plant growth and leaf quality in tea while also changing the secondary metabolite profile at low temperatures.

Transcriptome and Metabolome Analysis of Isoquinoline Alkaloid Biosynthesis of Coptis chinensis in Different Years.

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.

Identification of physiological and metabolic networks involved in postharvest browning of cigar tobacco leaves

The surface color of cigar tobacco leaves (CTLs) is largely determined by the browning response and is one of the most important quality traits affecting consumer preferences. The physiological changes and metabolic network of browning in CTLs after harvest have not been reported. We investigated the molecular mechanism of browning in CTLs by comparing the physiological characteristics and metabolites of CTLs browning at five postharvest periods. Phenotypic traits and browning-related enzyme activity data indicated that the browning reaction was most intense at approximately 12 d when polyphenol oxidase and peroxidase activities reached their maximum. Postharvest CTLs polyphenols, polyacids, and carbon and nitrogen compounds were rapidly degraded during the yellowing phase, and the biosynthesis rate of polyphenols and carotenoids was greater than the degradation rate during the browning phase. Metabolomic analysis discovered 2027 metabolites that were annotated mainly to lipids and lipid-like molecules, organic acids and their derivatives, and organic heterocyclic components. Hierarchical cluster analysis discovered glutamate, serine, threonine, ornithine, and arginine as the major amino acids involved in the browning reaction. KEGG pathway enrichment analysis indicated that the metabolic pathways with important effects on enzymatic browning were phenylpropanoid biosynthesis, flavonoid biosynthesis, isoquinoline alkaloid biosynthesis, phenylalanine metabolism, and linoleic acid metabolism. The metabolic pathway network was mined, and the main polyphenols involved in the browning reaction of CTLs were found to be cyanidin, rutin, caffeoylquinic acid, kaempferol, naringin, and neohesperidin. This study provides a reference for the browning physiology and metabolism network of postharvest CTLs.Graphical

Integrating Full-Length and Second-Generation Transcriptomics to Reveal Differentially Expressed Genes Associated with the Development of Corydalis yanhusuo Tuber.

Corydalis yanhusuo is a medicinal herb in China that has been widely used to treat various kinds of pain. The tuber is the main organ of C. yanhusuo used for medicinal purposes, but changes in related genes during the development of the tuber have rarely been reported. To identify the differentially expressed genes during tuber development, C. yanhusuo full-length transcriptomic sequencing was performed using single-molecule real-time technology, and tubers at three development stages were selected for comparative transcriptome analysis. A total of 90,496 full-length non-chimeric transcripts were obtained, and 19,341 transcripts were annotated in at least one public database. A total of 9221 differentially expressed genes were identified during the swelling process of C. yanhusuo tuber. A Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis revealed that differentially expressed genes associated with a "starch and sucrose metabolism pathway", "phenylpropanoid biosynthesis pathway", "isoquinoline alkaloid biosynthesis pathway", "zeatin biosynthesis pathway", and "brassinosteroid biosynthesis pathway" were predominantly enriched. In addition, the genes involved in cell wall metabolism were potentially associated with tuber swelling. These processes regulated and were involved in C. yanhusuo tuber development. The results provide a foundation for further research on tuber formation in medicinal plants.