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Objectives: Mirabilis jalapa L. and Bougainvillea spectabilis are two Mirabilis species known for their ornamental and pharmaceutical values. The organelle genomes are highly conserved with a rapid evolution rate making them suitable for evolutionary studies. Therefore, mitochondrial and chloroplast genomes of B. spectabilis and M. jalapa were sequenced to understand their evolutionary relationship with other angiosperms. Data description: Here, we report the complete mitochondrial genomes of B. spectabilis and M. jalapa (343,746 bp and 267,334 bp, respectively) and chloroplast genomes of B. spectabilis (154,520 bp) and M. jalapa (154,532 bp) obtained from Illumina NovaSeq. The mitochondrial genomes of B. spectabilis and M. jalapa consisted of 70 and 72 genes, respectively. Likewise, the chloroplast genomes of B. spectabilis and M. jalapa contained 131 and 132 genes, respectively. The generated genomic data will be useful for molecular characterization and evolutionary studies. [ABSTRACT FROM AUTHOR]

Objectives: Mirabilis jalapa L. and Bougainvillea spectabilis are two Mirabilis species known for their ornamental and pharmaceutical values. The organelle genomes are highly conserved with a rapid evolution rate making them suitable for evolutionary studies. Therefore, mitochondrial and chloroplast genomes of B. spectabilis and M. jalapa were sequenced to understand their evolutionary relationship with other angiosperms. Data description: Here, we report the complete mitochondrial genomes of B. spectabilis and M. jalapa (343,746 bp and 267,334 bp, respectively) and chloroplast genomes of B. spectabilis (154,520 bp) and M. jalapa (154,532 bp) obtained from Illumina NovaSeq. The mitochondrial genomes of B. spectabilis and M. jalapa consisted of 70 and 72 genes, respectively. Likewise, the chloroplast genomes of B. spectabilis and M. jalapa contained 131 and 132 genes, respectively. The generated genomic data will be useful for molecular characterization and evolutionary studies. [ABSTRACT FROM AUTHOR]

Scopolia lurida , a medicinal plant native to the Tibetan Plateau, is among the most effective producers of pharmaceutical tropane alkaloids (TAs). The hyoscyamine 6β-hydroxylase genes of Hyoscyamus niger ( HnH6H ) and S. lurida ( SlH6H ) were cloned and respectively overexpressed in hairy root cultures of S. lurida , to compare their effects on promoting the production of TAs, especially the high-value scopolamine. Root cultures with SlH6H / HnH6H overexpression were confirmed by PCR and real-time quantitative PCR, suggesting that the enzymatic steps defined by H6H were strongly elevated at the transcriptional level. Tropane alkaloids, including hyoscyamine, anisodamine and scopolamine, were analyzed by HPLC. Scopolamine and anisodamine contents were remarkably elevated in the root cultures overexpressing SlH6H / HnH6H , whereas that of hyoscyamine was more or less reduced, when compared with those of the control. These results also indicated that SlH6H and HnH6H promoted anisodamine production at similar levels in S. lurida root cultures. More importantly, HnH6H -overexpressing root cultures had more scopolamine in them that did SlH6H -overexpressing root cultures. This study not only provides a feasible way of overexpressing H6H to produce high-value scopolamine in engineered root cultures of S. lurida but also found that HnH6H was better than SlH6H for engineering scopolamine production. [ABSTRACT FROM AUTHOR]

Mirabilis himalaica is an endangered medicinal plant species in the Tibetan Plateau. The two genes respectively encoding chalcone synthase ( MhCHS) and chalcone isomerase ( MhCHI) were isolated and characterized from M. himalaica. The sequence analysis revealed that the two genes were similar with their corresponding homologous genes in other plants. The tissue profiles showed that both MhCHS and MhCHI had higher expression levels in roots than in stems and leaves. Transgenic hairy root cultures respectively with overexpressing MhCHS and MhCHI were established. The genomic PCR detection confirmed the authority of transgenic hairy root lines, in which either MhCHS or MhCHI expression levels were much higher than that in non-transgenic hairy root line. Finally, the HPLC detection results demonstrated that the rotenoid contents in MhCHS/ MhCHI-transformed hairy root lines were enhanced. This study provided two candidate genes that could be used to genetic engineering rotenoid biosynthesis in M. himalaica and an alternative method to produce rotenoid using transgenic hairy root cultures. [ABSTRACT FROM AUTHOR]

Tyrosine decarboxylase initializes salidroside biosynthesis. Metabolic characterization of tyrosine decarboxylase gene from Rhodiola crenulata (RcTYDC) revealed that it played an important role in salidroside biosynthesis. Recombinant 53 kDa RcTYDC converted tyrosine into tyramine. RcTYDC gene expression was induced coordinately with the expression of RcUDPGT (the last gene involved in salidroside biosynthesis) in SA/MeJA treatment; the expression of RcTYDC and RcUDPGT was dramatically upregulated by SA, respectively 49 folds and 36 folds compared with control. MeJA also significantly increased the expression of RcTYDC and RcUDPGT in hairy root cultures. The tissue profile of RcTYDC and RcUDPGT was highly similar: highest expression levels found in stems, higher expression levels in leaves than in flowers and roots. The gene expressing levels were consistent with the salidroside accumulation levels. This strongly suggested that RcTYDC played an important role in salidroside biosynthesis in R. crenulata. Finally, RcTYDC was used to engineering salidroside biosynthetic pathway in R. crenulata hairy roots via metabolic engineering strategy of overexpression. All the transgenic lines showed much higher expression levels of RcTYDC than non-transgenic one. The transgenic lines produced tyramine, tyrosol and salidroside at higher levels, which were respectively 3.21–6.84, 1.50–2.19 and 1.27–3.47 folds compared with the corresponding compound in non-transgenic lines. In conclusion, RcTYDC overexpression promoted tyramine biosynthesis that facilitated more metabolic flux flowing toward the downstream pathway and as a result, the intermediate tyrosol was accumulated more that led to the increased production of the end-product salidroside. [ABSTRACT FROM AUTHOR]

Anthocyanins are water-soluble flavonoid pigments that play a crucial role in plant growth and metabolism. They serve as attractants for animals by providing plants with red, blue, and purple pigments, facilitating pollination and seed dispersal. The fruits of solanaceous plants, tomato (Solanum lycopersicum) and eggplant (Solanum melongena), primarily accumulate anthocyanins in the fruit peels, while the ripe fruits of Atropa belladonna (Ab) have a dark purple flesh due to anthocyanin accumulation. In this study, an R2R3-MYB transcription factor (TF), AbMYB1, was identified through association analysis of gene expression and anthocyanin accumulation in different tissues of A. belladonna. Its role in regulating anthocyanin biosynthesis was investigated through gene overexpression and RNA interference (RNAi). Overexpression of AbMYB1 significantly enhanced the expression of anthocyanin biosynthesis genes, such as AbF3H, AbF3′5′H, AbDFR, AbANS, and Ab3GT, leading to increased anthocyanin production. Conversely, RNAi-mediated suppression of AbMYB1 resulted in decreased expression of most anthocyanin biosynthesis genes, as well as reduced anthocyanin contents in A. belladonna. Overall, AbMYB1 was identified as a fruit-expressed R2R3-MYB TF that positively regulated anthocyanin biosynthesis in A. belladonna. This study provides valuable insights into the regulation of anthocyanin biosynthesis in Solanaceae plants, laying the foundation for understanding anthocyanin accumulation especially in the whole fruits of solanaceous plants. [ABSTRACT FROM AUTHOR]

As an end product of purine metabolism, uric acid (UA) is a major endogenous antioxidant in humans. However, impaired UA synthesis and excretion can lead to hyperuricemia (HUA), which may in turn induce endothelial dysfunction (ED) and contribute to the pathogenesis of cardiovascular diseases (CVDs; e.g., atherosclerosis and hypertension). In this review, we discuss recent advances and novel insights into the effects exerted by HUA conditions in ED and related underlying mechanisms focusing on impaired UA metabolism, reduction in the synthesis and bioavailability of nitric oxide, endothelial cell injury, the endothelial‐to‐mesenchymal transition, insulin resistance, procoagulant activity, and acquisition of an inflammatory phenotype. We additionally discuss intervention strategies for HUA‐induced ED and the paradoxical roles of UA in endothelial function. We summarize major conclusions and perspectives: the deleterious effects of HUA contribute to the initiation and progression of CVD‐related ED. However, the treatment strategies (in addition to urate‐lowering therapy) for increasing endothelial function are limited because the majority of literature on pharmacological and pathophysiological mechanisms underlying HUA‐induced ED solely describes in vitro models. Therefore, a better understanding of the mechanisms involved in HUA‐induced ED is critical to the development of novel therapies for preventing and treating CVD‐HUA comorbidities. [ABSTRACT FROM AUTHOR]

Polyamines, including putrescine, spermidine, and spermine, play critical roles in cell physiology by different forms. As a rate-limiting enzyme that converts ornithine to putrescine, ornithine decarboxylase (ODC, EC 1.1.1.37) has been studied in detail in animals and microorganisms, but its specific functions are poorly understood in plants. In this study, the metabolic and developmental roles of the ODC gene were studied through RNAi-mediated suppression of the ODC gene (AbODC) in A. belladonna. Suppression of AbODC reduced the production of precursors of medicinal tropane alkaloids, including putrescine and N -methylputrescine, as well as hyoscyamine and scopolamine. In AbODC -RNAi roots, the production of putrescine and spermidine in free form was reduced, but in the AbODC -RNAi leaves, the content of free polyamines was not altered. In the roots/leaves of AbODC -RNAi plants, the production of conjugated and bound polyamines was reduced. In addition, suppression of the ODC gene resulted in reduction of polyamines and pollen sterility in AbODC- RNAi flowers. In floral organs, GUS-staining results indicated that AbODC was domainantly expressed in pollen. In summary, ornithine decarboxylase not only plays a key role in regulating the biosynthesis of diverse forms of polyamines and medicinal tropane alkaloids, but also participates in pollen development. [Display omitted] • Suppression of AbODC reduced the yield of tropane alkaloids and its precursors. • Inhibition of AbODC expression suppressed the biosynthesis of various polyamines. • In flower, AbODC was dominantly expressed in pollen. • Suppression of the ODC gene resulted in pollen sterility in AbODC- RNAi plants. [ABSTRACT FROM AUTHOR]

Background: As a valuable medicinal plant, Rhodiola has a very long history of folk medicine used as an important adaptogen, tonic, and hemostatic. However, our knowledge of the chloroplast genome level of Rhodiola is limited. This drawback has limited studies on the identification, evolution, genetic diversity and other relevant studies on Rhodiola. Results: Six Rhodiola complete chloroplast genomes were determined and compared to another Rhodiola cp genome at the genome scale. The results revealed a cp genome with a typical quadripartite and circular structure that ranged in size from 150,771 to 151,891 base pairs. High similarity of genome organization, gene number, gene order, and GC content were found among the chloroplast genomes of Rhodiola. 186 (R. wallichiana) to 200 (R. gelida) SSRs and 144 pairs of repeats were detected in the 6 Rhodiola cp genomes. Thirteen mutational hotspots for genome divergence were determined and could be used as candidate markers for phylogenetic analyses and Rhodiola species identification. The phylogenetic relationships inferred by members of Rhodiola cluster into two clades: dioecious and hermaphrodite. Our findings are helpful for understanding Rhodiola's taxonomic, phylogenetic, and evolutionary relationships. Conclusions: Comparative analysis of chloroplast genomes of Rhodiola facilitates medicinal resource conservation, phylogenetic reconstruction and biogeographical research of Rhodiola. [ABSTRACT FROM AUTHOR]

Atropa belladonna is an important industrial crop for producing anticholinergic tropane alkaloids (TAs). Using glyphosate as selection pressure, transgenic homozygous plants of A. belladonna are generated, in which a novel calmodulin gene (AbCaM1) and a reported EPSPS gene (G2-EPSPS) are co-overexpressed. AbCaM1 is highly expressed in secondary roots of A. belladonna and has calcium-binding activity. Three transgenic homozygous lines were generated and their glyphosate tolerance and TAs' production were evaluated in the field. Transgenic homozygous lines produced TAs at much higher levels than wild-type plants. In the leaves of T2GC02, T2GC05, and T2GC06, the hyoscyamine content was 8.95-, 10.61-, and 9.96 mg/g DW, the scopolamine content was 1.34-, 1.50- and 0.86 mg/g DW, respectively. Wild-type plants of A. belladonna produced hyoscyamine and scopolamine respectively at the levels of 2.45 mg/g DW and 0.30 mg/g DW in leaves. Gene expression analysis indicated that AbCaM1 significantly up-regulated seven key TA biosynthesis genes. Transgenic homozygous lines could tolerate a commercial recommended dose of glyphosate in the field. In summary, new varieties of A. belladonna not only produce pharmaceutical TAs at high levels but tolerate glyphosate, facilitating industrial production of TAs and weed management at a much lower cost. • AbCaM1 is a novel calmodulin regulating alkaloid biosynthesis in Atropa belladonna. • The method using glyphosate as direct selection pressure is established to obtain engineered plants of A. belladonna. • The plants of A. belladonna overexpressing AbCaM1 and G2-EPSPS have high-yield tropane alkaloids and glyphosate resistance. [ABSTRACT FROM AUTHOR]

Nine previously undescribed iridoids, ptehosides A-I (1 – 9), together with 12 known ones (10 – 21), were isolated from Pterocephalus hookeri (C.B. Clarke) Höeck. Their structures were elucidated using various spectroscopic methods including HR-ESI-MS, NMR, UV, IR and CD, etc. The cytotoxic activities of all isolates were evaluated using MTT method in three human cancer cell lines (Caco2, Huh-7, and SW982). As result, compound 9 exhibited substantial inhibitory activity on Caco2, Huh-7, and SW982 cells with IC 50 values of 1.17 ± 0.05, 1.15 ± 0.05 and 1.14 ± 0.04 μM, respectively. A preliminary mechanism study showed that 9 arrested the cell cycle of SW982 cells in the G0/G1 phase and induced apoptosis by upregulating Bax expression and downregulating Bcl-2 expression. Nine undescribed iridoids, together with twelve known ones were isolated from the whole plant of Pterocephalus hookeri. Among them, seven compounds showed significant cytotoxic activities against SW982 cells in vitro. Mechanically, ptehoside I (9) could arrest the cell-cycle in G0/G1 phase and induce apoptosis. [Display omitted] • Nine undescribed iridoids were isolated from the whole plant of Pterocephalus hookeri. • The stereochemistry was identified by acid hydrolysis, experimental and calculated ECD. • Ptehoside I (9) induced G0/G1 cell cycle arrest against SW982 cells in vitro. • 9 induced apoptosis and regulated the expression levels of Bax and Bcl-2 proteins in SW982 cells. [ABSTRACT FROM AUTHOR]

Real-time quantitative PCR (RT-qPCR) is an important technique for studying gene expression analysis, but accurate and reliable results depend on the use of a stable reference gene. This study proposes to test the expression stability of candidate reference genes in the callus of Saussurea laniceps, a unique Tibetan medicinal plant. Based on the S. laniceps callus transcriptome, eleven candidate reference genes, including TUA2, TUB3, TUB8, TIF3B1, TIF3H1, ELF5A, PP2AA2, UEV1D, UBL5, UBC36, and SKIP1), were validated for RT-qPCR normalization in the callus under abiotic stress (salt, cold, and UV) and hormone treatments (abscisic acid, MeJA, and salicylic acid). The stability of the candidate genes was evaluated in all the samples of S. laniceps. Comprehensive analysis of all samples showed that the best reference genes were UBC36 and UBL5. ELF5A and TIF3B1 were ranked as the most stable genes in the sample sets under abiotic stress. For hormone stimulation, UBC36 and TIF3H1 genes had the best stability. This study provides useful guidelines and a starting point for reference gene selection for expression analysis using RT-qPCR techniques in S. laniceps. [ABSTRACT FROM AUTHOR]

Medicinal tropane alkaloids (TAs), including hyoscyamine, anisodamine and scopolamine, are essential anticholinergic drugs specifically produced in several solanaceous plants. Atropa belladonna is one of the most important medicinal plants that produces TAs. Therefore, it is necessary to cultivate new A. belladonna germplasm with the high content of TAs. Here, we found that the levels of TAs were elevated under low nitrogen (LN) condition, and identified a LN-responsive bHLH transcription factor (TF) of A. belladonna (named LNIR) regulating the biosynthesis of TAs. The expression level of LNIR was highest in secondary roots where TAs are synthesized specifically, and was significantly induced by LN. Further research revealed that LNIR directly activated the transcription of hyoscyamine 6 β -hydroxylase gene (H6H) by binding to its promoter, which converts hyoscyamine into anisodamine and subsequently epoxidizes anisodamine to form scopolamine. Overexpression of LNIR upregulated the expression levels of TA biosynthesis genes and consequently led to the increased production of TAs. In summary, we functionally identified a LN-responsive bHLH gene that facilitated the development of A. belladonna with high-yield TAs under the decreased usage of nitrogen fertilizer. • Biosynthesis of TAs was improved under low nitrogen (LN) condition. • LN treatment induced the expression of LNIR, a bHLH transcription factor in Atropa belladonna. • LNIR positively regulated the biosynthesis of TAs by promoting the expression of TAs biosynthesis genes. [ABSTRACT FROM AUTHOR]

Putrescine, produced via the arginine decarboxylase (ADC)/ornithine decarboxylase (ODC)-mediated pathway, is an initial precursor for polyamines metabolism and the root-specific biosynthesis of medicinal tropane alkaloids (TAs). These alkaloids are widely used as muscarinic acetylcholine antagonists in clinics. Although the functions of ODC in biosynthesis of polyamines and TAs have been well investigated, the role of ADC is still poorly understood. In this study, enzyme inhibitor treatment showed that ADC was involved in the biosynthesis of putrescine-derived metabolites and root growth in Atropa belladonna. Further analysis found that there were six ADC unigenes in the A. belladonna transcriptome, with two of them, AbADC1 and AbADC2 , exhibiting high expression in the roots. To investigate their roles in TAs/polyamines metabolism and root growth, RNA interference (RNAi) was used to suppress either AbADC1 or AbADC2 expression in A. belladonna hairy roots. Suppression of the AbADC1 expression resulted in a significant reduction in the putrescine content and hairy root biomass. However, it had no noticeable effect on the levels of N -methylputrescine and the TAs hyoscyamine, anisodamine, and scopolamine. On the other hand, suppression of AbADC2 expression markedly reduced the levels of putrescine, N -methylputrescine, and TAs, but had no significant effect on hairy root biomass. According to β -glucuronidase (GUS) staining assays, AbADC1 was mainly expressed in the root elongation and division region while AbADC2 was mainly expressed in the cylinder of the root maturation region. These differences in expression led to functional divergence, with AbADC1 primarily regulating root growth and AbADC2 contributing to TA biosynthesis. • RNAi reduced putrescine biosynthesis by suppressing AbADC1 or AbADC2 in A. belladonna hairy roots. • Knocking down AbADC1 reduced hairy root biomass, while knocking down AbADC2 did not. • Knocking down AbADC2 suppressed the biosynthesis of N -methylputrescine and TAs. • AbADC1 exhibited high expression in the root elongation and division region. • AbADC2 was highly expressed in the cylinder of the root maturation region. [ABSTRACT FROM AUTHOR]

In the present study, an immunological arabinan, LCP70-2A, was isolated from Ligusticum chuanxiong for the first time. The absolute molecular weight of LCP70-2A was determined to be 6.46 × 104 g/mol using the HPSEC-MALLS-RID method. The absolute configuration of arabinose in LCP70-2A was determined to be L-configuration. Physicochemical characterization revealed that LCP70-2A was a homogeneous polysaccharide and had a backbone of (1 → 5)-linked α -L-Ara f with terminal α -L-arabinose residues at position O -2 and O -3. Molecular conformation analysis showed that LCP70-2A was a branching polysaccharide with a compact coil chain conformation in 0.1 M NaCl solution. In addition, in vitro cell assays showed that LCP70-2A can activate macrophages by enhancing the phagocytosis and potentiating the secretion of immunoregulatory factors including NO, TNF- α , IL-6, and IL-1 β. Furthermore, LCP70-2A was proved to promote the production of ROS and NO using the zebrafish model, suggesting that LCP70-2A can be further developed as a candidate supplement for immunological enhancement. Unlabelled Image • A novel polysaccharide (LCP70-2A) was isolated from the rhizome of L. chuanxiong. • LCP70-2A was determined to be an arabinan. • LCP70-2A increased the levels of NO, IL-6, IL-1 β , and TNF- α in RAW264.7 cells. • LCP70-2A was proven to enhance the immunoregulatory activity of zebrafish. [ABSTRACT FROM AUTHOR]

Dracocephalum tanguticum and Dracocephalum moldavica are important herbs from Lamiaceae and have great medicinal value. We used the Illumina sequencing technology to sequence the complete chloroplast genome of D. tanguticum and D. moldavica and then conducted de novo assembly. The two chloroplast genomes have a typical quadripartite structure, with the gene's lengths of 82,221 bp and 81,450 bp, large single-copy region's (LSC) lengths of 82,221 bp and 81,450 bp, and small single-copy region's (SSC) lengths of 17,363 bp and 17,066 bp, inverted repeat region's (IR) lengths of 51,370 bp and 51,352 bp, respectively. The GC content of the two chloroplast genomes was 37.80% and 37.83%, respectively. The chloroplast genomes of the two plants encode 133 and 132 genes, respectively, among which there are 88 and 87 protein-coding genes, respectively, as well as 37 tRNA genes and 8 rRNA genes. Among them, the rps2 gene is unique to D. tanguticum, which is not found in D. moldavica. Through SSR analysis, we also found 6 mutation hotspot regions, which can be used as molecular markers for taxonomic studies. Phylogenetic analysis showed that Dracocephalum was more closely related to Mentha. [ABSTRACT FROM AUTHOR]

Paeonia ludlowii is indigenous to Tibet and has an important ecological and economic value in China. In Tibet, P. ludlowii has been used in folk medicine with relative success. Plant microbial endophytes play an important role in plant growth, health and ecological function. The diversity of endophytic bacteria associated with P. ludlowii remains poorly understood. In this study, the structure of the endophytic bacterial communities associated with different tissues, including fruits, flowers, leaves, stems, and roots, and rhizosphere soils was analyzed with Illumina MiSeq sequencing of bacterial 16S rDNA. A total of 426,240 sequences and 4847 operational taxonomic units (OTUs) were obtained. The OTUs abundance of roots was higher than that of other tissues; however, the OTUs abundance was similar among different deep soil samples. In the plant tissues, Cyanobacteria was the most abundant bacterial phylum, followed by Proteobacteria; however, the most abundant phyla were Proteobacteria and Acidobacteria in soil samples from three different layers. In addition, the diversity and richness of the microorganisms in the soil were very similar to those in roots but higher than those in other tissues of P. ludlowii. Predictive metagenome analysis revealed that endophytic bacteria play critical functional roles in P. ludlowii. This conclusion could facilitate the study of the ecological functions of endophytic bacteria and their interactions with P. ludlowii to analyze the reasons why this important medicinal plant is becoming endangered. [ABSTRACT FROM AUTHOR]

Rheum species present a significant economic value. Traditional Chinese medicine rhubarb is an important medicinal material in China. It has a long history of use, with a record of use as early as two thousand years ago. Here, we determined the complete chloroplast genome sequences of Rheum nobile and Rheum acuminatum and comprehensively compared them to two other available Rheum cp genomes at the genome scale. The results revealed cp genomes ranging in size from 159,051 to 161,707 bp with a similar typical quadripartite and circular structure. The genome organization, gene numbers, gene order, and GC contents of these four Rheum cp genomes were similar to those of many angiosperm cp genomes. Repeats and microsatellites were detected in the R. nobile and R. acuminatum cp genomes. The Mauve alignment revealed that there were no rearrangements in the cp genomes of the four Rheum species. Thirteen mutational hotspots for genome divergence were identified, which could be utilized as potential markers for phylogenetic studies and the identification of Rheum species. The phylogenetic relationships of the four species showed that the members of Rheum cluster into a single clade, indicating their close relationships. Our study provides valuable information for the taxonomic, phylogenetic, and evolutionary analysis of Rheum. [ABSTRACT FROM AUTHOR]

Herpetotriol, a typical lignan in Herpetospermum pedunculosum Wall's seeds that has long been used to treat icterhepatitis and indigestion and other related diseases in Tibet, is of potential hepatoprotection. This study aims to study the pharmacokinetics features of herpetotriol, including the blood drug concentration – time curve and tissue distribution. The ultrahigh-performance liquid chromatography with tandem mass spectrometry method was established to detect herpetotriol concentration in plasma and tissues, and the method showed good linearity from 10 to 2000 ng/mL (r ≥ 0.9972) and sensitivity (≥10 ng/mL). Our blood drug concentration – time curve indicated that herpetotriol was distributed quickly in rats with a Tmax value at about 0.083 h and eliminated rapidly with a clearance rate at 98.13 ± 8.05 and 137.04 ± 9.48 L·h–1·kg–1 with doses of 5 and 2.5 mg/kg, respectively. Although herpetotriol was detectable in all tested tissues, it has a higher concentration in liver than in heart, lung, spleen, and kidney, which is in line with its hepatoprotection. The pharmacokinetics features uncovered by the present study could provide more information for future pharmacological and toxicological study of herpetotriol. [ABSTRACT FROM AUTHOR]

Summary: Some medicinal plants of the Solanaceae produce pharmaceutical tropane alkaloids (TAs), such as hyoscyamine and scopolamine. Littorine is a key biosynthetic intermediate in the hyoscyamine and scopolamine biosynthetic pathways. However, the mechanism underlying littorine formation from the precursors phenyllactate and tropine is not completely understood.Here, we report the elucidation of littorine biosynthesis through a functional genomics approach and functional identification of two novel biosynthesis genes that encode phenyllactate UDP‐glycosyltransferase (UGT1) and littorine synthase (LS).UGT1 and LS are highly and specifically expressed in Atropa belladonna secondary roots. Suppression of either UGT1 or LS disrupted the biosynthesis of littorine and its TA derivatives (hyoscyamine and scopolamine). Purified His‐tagged UGT1 catalysed phenyllactate glycosylation to form phenyllactylglucose. UGT1 and LS co‐expression in tobacco leaves led to littorine synthesis if tropine and phenyllactate were added.This identification of UGT1 and LS provides the missing link in littorine biosynthesis. The results pave the way for producing hyoscyamine and scopolamine for medical use by metabolic engineering or synthetic biology. [ABSTRACT FROM AUTHOR]

Atropa belladonna is widely utilized for producing anticholinergic tropane alkaloids (TAs). Of them, scopolamine is the most valuable component due to its potent pharmacological effects and minimal side effects. However, the content of scopolamine in planta is extremely low, emphasizing the need to cultivate high-yield varieties of A. belladonna. Here it is reported that the genes encoding the rate-limiting upstream enzyme ornithine decarboxylase from A. belladonna (AbODC) and downstream enzyme hyoscyamine 6 β -hydroxylase from Hyoscyamus niger (HnH6H) were simultaneously overexpressed to genetically modify A. belladonna plants. Metabolite analysis in the AbODC overexpressing plants showed that the levels of hyoscyamine and anisodamine in both the roots and leaves of A. belladonna were significantly increased. However, there was no significant effect on the accumulation of scopolamine. On the other hand, overexpressing HnH6H alone was able to increase the contents of scopolamine to some extent. When AbODC and HnH6H were co-expressed, there was a greater increase in the contents of scopolamine in both roots and leaves, especially in the leaves, where the average content of scopolamine reached 6.67 mg/g dry weight (DW). Moreover, the increasing expression levels of ODC in transgenic plants greatly enhanced the biosynthesis of polyamines, thereby enhancing the cold tolerance trait of A. belladonna plants. These findings highlight the importance of co-overexpressing AbODC and HnH6H in A. belladonna and provide an effective promising approach to developing new varieties of A. belladonna with not only a high yield of scopolamine but also a high tolerance of cold stress. • AbODC overexpression boosts hyoscyamine yield and cold tolerance in A. belladonna. • HnH6H overexpression promotes scopolamine yield in A. belladonna. • Plants co-expressing ODC and H6H show cold tolerance and higher scopolamine yield. [ABSTRACT FROM AUTHOR]

Pterocephalus hookeri was used as a traditional Chinese medicine for the treatment of rheumatoid arthritis. Sweroside was a main iridoid isolated from P. hookeri. The present study aimed to investigate the anti-inflammatory effect mechanism of sweroside. In RAW264.7 cells induced by lipopolysaccharide (LPS), the abnormal proliferation, the NO content increase, and the downregulated Sirtuin1 (SIRT1) expression were observed. Sweroside could alleviate the inflammation by inhibiting cell proliferation through arresting the cell cycle at the G0/G1 phase, by suppressing pro-inflammatory cytokines and by promoting anti-inflammatory cytokines in LPS-induced RAW264.7 cells. Further mechanism research indicated that sweroside could activate the SIRT1, then suppress the nuclear factor-kappa B (NF-κB) and promote the Forkhead transcription factor O1 (FOXO1) signaling pathways. The present study indicated that sweroside may be the main anti-inflammatory constituent of P. hookeri and a promising candidate for anti-inflammation therapy. [ABSTRACT FROM AUTHOR]

Basic helix-loop-helix (bHLH) proteins are the second largest family of transcription factors (TFs) involved in developmental and physiological processes in plants. In this study, 205 putative bHLH TF genes were identified in the genome of Artemisia annua and expression of 122 of these was determined from transcriptomes used to construct the genetic map of A. annua. Analysis of gene expression association allowed division of the 122 bHLH TFs into five groups. Group V, containing 15 members, was tightly associated with artemisinin biosynthesis genes. Phylogenetic analysis indicated that two bHLH TFs, AabHLH106 and AabHLH112, were clustered with Arabidopsis ICE proteins. AabHLH112 was induced by low temperature, while AabHLH106 was not. We therefore chose AabHLH112 for further examination. AabHLH112 was highly expressed in glandular secretory trichomes, flower buds, and leaves. Dual-luciferase assays demonstrated that AabHLH112 enhanced the promoter activity of artemisinin biosynthesis genes and AaERF1 , an AP2/ERF TF that directly and positively regulates artemisinin biosynthesis genes. Yeast one-hybrid assays indicated that AabHLH112 could bind to the AaERF1 promoter, but not to the promoters of artemisinin biosynthesis genes. Overexpression of AabHLH112 significantly up-regulated the expression levels of AaERF1 and artemisinin biosynthesis genes and consequently promoted artemisinin production. [ABSTRACT FROM AUTHOR]

A bioassay-guided chemical investigation on the ethyl acetate extract of the stems of Herpetospermum pedunculosum led to the isolation and identification of 22 lignans including 6 previously undescribed ones, herpetosiols A-F. Their structures including stereochemistries were elucidated by analysis of NMR, HRMS and ECD data. The in vitro cytotoxic activities of all isolates were studied against human gastric carcinoma SGC7901, lung carcinoma A549, breast carcinoma MDA-MB-231 and hepatocellular carcinoma HepG2 cell lines. Among them, eight lignans exhibited anti-proliferative effects against four tumor cell lines with IC 50 ranging from 1.7 ± 0.1 to 32.6 ± 1.1 μM. Hedyotol-B displayed potent inhibitory effect with IC 50 values of 1.7 ± 0.1 μM against SGC7901 and 6.1 ± 0.5 μM against A549, respectively. Image 1 • Six undescribed lignans were obtained from Herpetospermum pedunculosum. • The stereochemistry was elucidated by ECD and 2D-NMR. • Eight compounds selectively inhibited the proliferation of four cancer cell lines. [ABSTRACT FROM AUTHOR]

Przewalskia tangutica is a traditional medicinal plant from Tibet used for the analgesic effect from the tropane alkaloids (TAs) produced by the plant. Its roots have the highest yield of hyoscyamine in all plant species and so have been overharvested becoming an endangered medicinal plant species. Metabolic engineering is a good way to improve the yield of TAs in plants. In our study, two functionally distinct tropinone reductases genes, PtTRI and PtTRII, were cloned from P. tangutica and the functional divergence were characterized. The enzyme kinetics of PtTRI and PtTRII were investigated. The phylogenetic analysis classified them into different clades: PtTRI and PtTRII were in the clade of tropine‐forming reductase and pseudotropine‐forming reductase, respectively. We found PtTRI to be expressed in the roots but less in leaves, whereas PtTRII was expressed in the roots at higher levels than in the leaves. The kinetic parameters (Km, Vmax, and Kcat) were analyzed using purified recombinant enzymes at their optimum pH. Enzymatic analysis results showed that tropinone is a better substrate for PtTRII compared with PtTRI, suggesting that PtTRII might be a potential gene target for TA biosynthesis engineering. Compared with the reported TRIs, PtTRI exhibited a higher affinity for tropinone. [ABSTRACT FROM AUTHOR]

Mirabilis himalaica is an important Tibetan medicinal plant in China. However, it has become a rare and class I endangered Tibetan medicine plant. Therefore, the use of callus to propagate germplasm resources is of great significance. We found that the flavonoid content of M. himalaica callus increased continuously with the extension of UV-B treatment. Multi-omics profiles were used to reveal the co-expression patterns of gene networks of flavonoid metabolism in M. himalaica callus during UV-B radiation. Results showed that five medicinal metabolics, including geranin, eriodictyol, astragalin, isoquercetin, pyrotechnic acid, and one anthocyanin malvide-3-O-glucoside were identified. The transcriptome data were divided into 46 modules according to the expression pattern by WGCNA (weighted gene co-expression network analysis), of which the module Turquoise had the strongest correlation with six target metabolites. We found that seven structural genes and twenty-five transcription factors were related to the metabolism of flavonoid synthesis, among which the structural genes CHI , C4H and UGT79B6 had strong co-expression relationships with the 6 target metabolites. WRKY42, WRKY7, bHLH128 and other transcription factors had strong co-expression relationships with multiple structural genes. Consequently, these findings suggest callus grown under UV-B treatment could be an effective alternative medical resource of M. himalaica , which is valuable for conservation and usage of this wild and endangered plant. • Six key medicinal metabolic were identified in M. himalaica callus by UV-B radiation. • WGCNA was used to analyze the relationship between gene modules and metabolites. • CHI , C4H and UGT79B6 had strong co-expression relationship with 6 target metabolites. • WRKY42, WRKY7, bHLH128 had strong co-expression relationships with structural genes. [ABSTRACT FROM AUTHOR]

Rhodiola crenulata is a Tibetan native herbal plant belonging to the family of Crassulaceae, which produces the pharmaceutical icariside D2 with the activities of inhibiting angiotensin-converting enzyme and killing leukemia cancer cells. In this study, we functionally characterized a novel UDP-glycosyltransferase (RcUGT1) that converted tyrosol to specifically produce icariside D2 from R. crenulata at molecular and biochemical levels. RcUGT1 was highly expressed in flowers and roots, while the icariside D2 content was much higher in stems than that in other organs, suggesting the potential translocation of icariside D2 from flowers and roots to stems. The high production of icariside D2 in stems provided a reasonable suggestion to farmers to harvest stems instead of roots for icariside D2 production. Enzymatic assays of recombinant RcUGT1 indicated that it converted tyrosol to specifically form icariside D2, with the values of Km 0.97±0.10 mM, Vmax 286±8.26 pKat/mg, Kcat 0.01552 s−1, and Kcat/Km 159.55 s−1 M−1. Functional identification of RcUGT1 facilitated the icariside D2 production through metabolic engineering in plants or synthetic biology in microbes. [ABSTRACT FROM AUTHOR]

N -methylputrescine is the precursor of nicotine and pharmaceutical tropane alkaloids such as hyoscyamine. Putrescine N -methyltransferase (PMT) catalyzes the N -methylation of putrescine to form N -methylputrescine. While the role of PMT in nicotine biosynthesis is clear, knowledge of PMT in the biosynthesis of tropane alkaloids (TAs) and the regulation of polyamines remains limited. We characterized a PMT gene from Hyoscyamus niger , designated HnPMT that was specifically expressed in roots, especially in the secondary roots and dramatically induced by methyl jasmonate (MeJA). The GUS gene was specifically expressed in Arabidopsis roots or in the vascular tissues, including pericycles and endodermis, of the H. niger hairy root cultures, when it was driven by the 5′-flanking promoter region of HnPMT . The recombinant HnPMT was purified for enzymatic assays. HnPMT converted putrescine to form N -methylputrescine, as confirmed by LC-MS. The kinetics analysis revealed that HnPMT had high affinity with putrescine but low catalytic activity, suggesting that it was a rate-limiting enzyme. When HnPMT was suppressed in the H. niger plants by using the VIGS approach, the contents of N -methylputrescine and hyoscyamine were markedly decreased, but the contents of putrescine, spermidine and a mixture of spermine and thermospermine were significantly increased; this suggested that HnPMT was involved in the biosynthesis of tropane alkaloids and played a competent role in regulating the biosynthesis of polyamines. Functional identification of HnPMT facilitated the understanding of TA biosynthesis and thus implied that the HnPMT-catalyzed step might be a target for metabolic engineering of the TA production in H. niger . [ABSTRACT FROM AUTHOR]

Stem cell transplantation is one of most valuable methods in the treatment of myocardial infarction, and adipose-derived stem cells (ASCs) are becoming a hot topic in medical research. Previous studies have shown that ASCs can be differentiated into cardiomyocyte-like cells, but the efficiency and survival rates are low. We investigated the role and mechanism of microRNA-1 (miR-1) in the differentiation of ASCs into cardiomyocyte-like cells. ASCs and cardiomyocytes were isolated from neonatal rats. We constructed lentivirus for overexpressing miR-1 and used DAPT, an antagonist of the Notch1 pathway, for in vitro analyses. We performed cocultures with ASCs and cardiomyocytes. The differentiation efficiency of ASCs was detected by cell-specific surface antigens. Our results showed that miR-1 can promote the expression of Notch1 and reduce the expression of Hes1, a Notch pathway factor, and overexpression of miR-1 can promote the differentiation of ASCs into cardiomyocyte-like cells, which may occur by regulating Notch1 and Hes1. [ABSTRACT FROM AUTHOR]

This study compared the effects of hot air drying (HAD) and vacuum drying (VD) on the physicochemical qualities of lily (Lilium lancifolium Thunb.) bulbs. The results showed that VD treatment effectively reduced the browning of lily scales with a total color difference (ΔE) 42–73% lower than those of HAD group. Two drying treatments enhanced the total phenolic content (TPC) of the lily bulbs effectively. TPC increased with rising drying temperatures, and under isothermal conditions, the TPC of VD group was 1.1–1.6-fold higher than that of HAD group. Five typical phenolic compounds were identified in lily bulbs. As the predominant phenolic fraction, regaloside A of VD group was 51–291% above HAD group. Regaloside B was temperature sensitive and would be destroyed by high temperature. In parallel, VD treatment elevated the antioxidant activity overall, which could be contributed by phenolics and some Maillard reaction products. Furthermore, drying significantly improved the antiproliferative activity of lily bulbs, with samples treated with 120 °C HAD and 60 °C VD exhibiting the strongest antiproliferative capacity. The present investigation demonstrated that VD treatment effectively improved the physicochemical quality of lily bulbs and would provide a theoretical basis for optimizing drying conditions to obtain higher-quality lily bulbs. [Display omitted] • Vacuum drying treatment effectively inhibited the browning of lily bulbs. • Regaloside A was the major phenolic in lily bulbs and increased by vacuum drying. • Regaloside B was temperature sensitive and would be damaged by high temperature. • 7 typical volatiles were identified in lily bulbs after hot air and vacuum drying. • Vacuum drying raised the antioxidant and antiproliferative features of lily bulbs. [ABSTRACT FROM AUTHOR]

In recent years, although several related genes had been cloned and characterized, the role of aldehyde dehydrogenase 1 ( ALDH1), the newly cloned gene involved in artemisinin biosynthesis pathway, is still not clear. In this study, a 2,100-bp ALDH1 promoter region fused with GUS reporter gene was stably transferred into Arabidopsis thaliana. Histochemical staining showed the methyl jasmonate (MeJA) and wounding treatment induced the GUS gene expression specifically in the trichomes of transgenic A. thaliana, consistent with the results that the expression level of ALDH1 gene was increased in the A. annua under MeJA and wounding treatments. Two RAA motifs (AP2/ERF binding site) but no W box (WRKY binding site) motif were identified in the ALDH1 promoter by the analysis through PLACE and plantCARE. Through the dual luciferase reporter assay, we revealed that both AaORA and AaERF2, rather than AaWRKY1, could activate the expression of ALDH1 promoter. Our study shed light on the in-depth understanding of the role of ALDH1 in artemisinin biosynthesis. [ABSTRACT FROM AUTHOR]

Polygonatum cirrhifolium is a rare and endangered Tibetan medicinal plant. In this paper, resource status of P. cirrhifolium from 13 counties of Tibet Plateau was Investigated by combining field Investigation with laboratory analysis, and sustainable utilization countermeasures were analyzed. Results showed that (1) distribution area sequence of P. cirrhifolium in each county wets Gongbo'gvamda County > Chayu County > Lang County > Milin County > Maizhokunggar County > Zuogong County > Mangkang County > Jiali County > Dlngqing County > Duilongdeqing County > Baqing County = Suo County = Nimu County. Its total distribution area was 458 723.81 hm², and It was not distributed in Baqing County, Nimu County and Suo County. (2) Average dry weight per plant of P. cirrhifolium medicinal part In each county had the bellow sequence; Duilongdeqing County > Gongbo'gvamda County > Maizhokunggar County > Zuogong County > Dlngqing County > Lang County > Mangkang County > Milin County >Jiali County >Chayu County > Baqing County = Nimu County = Suo County, and it reached 46.48 g/plant in Duilongdeqing County. (3) Biomass of P. cirrhifolium medicinal part in each county had the bellow sequence; Duilongdeqing County >Gongbo'gvamda County >Maizhokunggar County > Lang County > Dlngqing County > Zuogong County > Jiali County > Mangkang County > Milin County > Chayu County > Nimu County = Suo County = Baqing County, and It reached 300.52 kg/hm² in Duilongdeqing County. (4) Reserve amount sequence of P. cirrhifolium in each county was Gongbo'gvamda County > Lang County > Maizhokunggar County > Chayu County > Zuogong County > Dingqing County > Milin County > Jiali County > Mangkang County > Duilongdeqing County > Baqing County = Nimu County = Suo County, and total reserve amount was 42 293.961. (5) At medicinal function aspect, P. cirrhifolium Is sweet, and has the effects of keeping fit, nourishing vitality, regenerating teeth and delaying senility. (6) For resource status of P. cirrhifolium, It is suggested establishing resource protection region and constructing germplasm resource bank; establishing plantation park and perfecting cultivation and planting techniques; deeply processing and extending Industry chain of Tibetan medicine; updating concept and rationally developing and using. [ABSTRACT FROM AUTHOR]

Previous publications reported that the artemisinin level was increased in Artemisia annua following a night-frost period. However, the molecular mechanism was not clear. In this study, we found that exogenous jasmonate (JA) effectively enhanced the freezing tolerance of A. annua. The JA biosynthetic genes ( LOX1, LOX2, allene oxide cyclase [ AOC], and jasmonate resistant 1 [ JAR1]) were induced by cold stress, leading to an increase in endogenous JA in cold-treated A. annua. Increased endogenous JA enhanced the expression of three JA-responsive transcription factors, ethylene response factor 1, ethylene response factor 2, and octadecanoid-responsive AP2/ERF, all of which were reported to transcriptionally activate the expression of artemisinin biosynthetic genes, such as amorpha-4,11-diene synthase ( ADS), CYP71AV1, DBR2, and aldehyde dehydrogenase 1 ( ALDH1). Furthermore, the expression levels of the four artemisinin biosynthetic genes were also significantly increased under cold stress. Consequently, the levels of artemisinin and related secondary metabolites, such as dihydroartemisinic acid, artemisinin B, and artemisinic acid, were increased in A. annua under cold stress. Our study points to a molecular mechanism in which the production of artemisinin is regulated by cold stress in A. annua. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effects and mechanisms of quercetin on pulmonary arterial endothelial cell (PAEC) transdifferentiation into smooth muscle-like cells. TGF-β1-induced PAEC transdifferentiation models were applied to evaluate the pharmacological actions of quercetin. PAEC proliferation was detected with CCK8 method and BurdU immunocytochemistry. Meanwhile, the identification and transdifferentiation of PAECs were determined by FVIII immunofluorescence staining and α-SMA protein expression. The related mechanism was elucidated based on the levels of Akt and Erk1/2 signal pathways. As a result, quercetin effectively inhibited the TGF-β1-induced proliferation and transdifferentiation of the PAECs and activation of Akt/Erk1/2 cascade in the cells. In conclusion, quercetin is demonstrated to be effective for pulmonary arterial hypertension (PAH) probably by inhibiting endothelial transdifferentiation possibly via modulating Akt and Erk1/2 expressions. [ABSTRACT FROM AUTHOR]

Atropa belladonna is one of the most important plant sources for producing pharmaceutical tropane alkaloids (TAs). T1 progeny of transgenic A. belladonna , in which putrescine N -methyltransferase (EC. 2.1.1.53) from Nicotiana tabacum (NtPMT) and hyoscyamine 6β-hydroxylase (EC. 1.14.11.14) from Hyoscyamus niger (HnH6H) were overexpressed, were established to investigate TA biosynthesis and distribution in ripe fruits, leaves, stems, primary roots and secondary roots under field conditions. Both NtPMT and HnH6H were detected at the transcriptional level in transgenic plants, whereas they were not detected in wild-type plants. The transgenes did not influence the root-specific expression patterns of endogenous TA biosynthetic genes in A. belladonna . All four endogenous TA biosynthetic genes ( AbPMT , AbTRI , AbCYP80F1 and AbH6H ) had the highest/exclusive expression levels in secondary roots, suggesting that TAs were mainly synthesized in secondary roots. T1 progeny of transgenic A. belladonna showed an impressive scopolamine-rich chemotype that greatly improved the pharmaceutical value of A. belladonna . The higher efficiency of hyoscyamine conversion was found in aerial than in underground parts. In aerial parts of transgenic plants, hyoscyamine was totally converted to downstream alkaloids, especially scopolamine. Hyoscyamine, anisodamine and scopolamine were detected in underground parts, but scopolamine and anisodamine were more abundant than hyoscyamine. The exclusively higher levels of anisodamine in roots suggested that it might be difficult for its translocation from root to aerial organs. T1 progeny of transgenic A. belladonna , which produces scopolamine at very high levels (2.94–5.13 mg g −1 ) in field conditions, can provide more valuable plant materials for scopolamine production. [ABSTRACT FROM AUTHOR]

Brugmansia arborea is a woody plant species that produces tropane alkaloids (TAs). The gene encoding tropine-forming reductase or tropinone reductase I (BaTRI) in this plant species was functionally characterised. The full-length cDNA of BaTRI encoded a 272-amino-acid polypeptide that was highly similar to tropinone reductase I from TAs-producing herbal plant species. The purified 29 kDa recombinant BaTRI exhibited maximum reduction activity at pH 6.8–8.0 when tropinone was used as substrate; it also exhibited maximum oxidation activity at pH 9.6 when tropine was used as substrate. The K m, V max and K cat values of BaTRI for tropinone were 2.65 mM, 88.3 nkat mg −1 and 2.93 S −1 , respectively, at pH 6.4; the K m, V max and K cat values of TRI from Datura stramonium (DsTRI) for tropinone were respectively 4.18 mM, 81.20 nkat mg −1 and 2.40 S −1 at pH 6.4. At pH 6.4, 6.8 and 7.0, BaTRI had a significantly higher activity than DsTRI. Analogues of tropinone, 4-methylcyclohexanone and 3-quinuclidinone hydrochloride, were also used to investigate the enzymatic kinetics of BaTRI. The K m, V max and K cat values of BaTRI for tropine were 0.56 mM, 171.62 nkat.mg −1 and 5.69 S −1 , respectively, at pH 9.6; the K m, V max and K cat values of DsTRI for tropine were 0.34 mM, 111.90 nkat mg −1 and 3.30 S −1 , respectively, at pH 9.6. The tissue profiles of BaTRI differed from those in TAs-producing herbal plant species. BaTRI was expressed in all examined organs but was most abundant in secondary roots. Finally, tropane alkaloids, including hyoscyamine, anisodamine and scopolamine, were detected in various organs of B. arborea by HPLC. Interestingly, scopolamine constituted most of the tropane alkaloids content in B. arborea , which suggests that B. arborea is a scopolamine-rich plant species. The scopolamine content was much higher in the leaves and stems than in other organs. The gene expression and TAs accumulation suggest that the biosynthesis of hyoscyamine, especially scopolamine, occurred not only in the roots but also in the aerial parts of B. arborea . [ABSTRACT FROM AUTHOR]

Bioactivity-guided study led to the isolation of a natural phenylpropionate derivative, ( E )-3-(4-hydroxy-2-methoxyphenyl)-propenoic acid 4-hydroxy-3-methoxyphenyl ester from the roots of Mirabilis himalaica . Cellular analysis showed that compound 1 specifically inhibited the cancer cell growth through the S phase arrest. Mechanistically, compound 1 was able to induce the apoptosis in HepG2 cells through mitochondrial apoptosis pathway in which Bcl-2 and p53 were required. Interestingly, the cellular phenotype of compound 1 were shown specifically in cancer cells originated from hepatocellular carcinoma (HepG2) while compromised influence by compound 1 were detected within the normal human liver cells (L-02). Consistently, the in vivo inhibitory effects of compound 1 on tumor growth were validated by the in xenograft administrated with HepG2 cells. Our results provided a novel compound which might serve as a promising candidate and shed light on the therapy of the hepatocellular carcinoma. [ABSTRACT FROM AUTHOR]

Quantitative PCR (qPCR) is a powerful tool for measuring gene expression levels. Accurate and reproducible results are dependent on the correct choice of reference genes for data normalization. Atropa belladonna is a commercial plant species from which pharmaceutical tropane alkaloids are extracted. In this study, eight candidate reference genes, namely 18S ribosomal RNA ( 18S), actin ( ACT), cyclophilin ( CYC), elongation factor 1α ( EF-1α), β-fructosidase ( FRU), glyceraldehyde-3-phosphate dehydrogenase ( GAPDH), phosphoglycerate kinase ( PGK), and beta-tubulin ( TUB), were selected and their expression stabilities studied to determine their suitability for normalizing gene expression in A. belladonna. The expression stabilities of these genes were analyzed in the root, stem, and leaf under cold, heat, NaCl, UV-B, methyl jasmonate, salicylic acid, and abscisic acid treatments using geNorm, NormFinder, and BestKeeper. The statistical algorithms indicated that PGK was a reliable gene for normalizing gene expression under most of the experimental conditions. The pairwise value analysis showed that two genes were sufficient for proper expression normalization, except when analyzing gene expression in heat-treated roots. However, the choice of the second reference gene depended on specific conditions. Finally, the relative expression level of the PMT gene of A. belladonna was detected to validate the selection of PGK a reliable reference gene. In summary, our results should guide the selection of appropriate reference genes for gene expression studies in A. belladonna under different organs and abiotic stress conditions. [ABSTRACT FROM AUTHOR]

The 2- C-methyl- D-erythritol-4-phosphate ( MEP) pathway genes encoding DXR and MECS from Taxus species and STR from Catharanthus roseus were used to genetically modify the ajmalicine biosynthetic pathway in hairy root cultures of C. roseus. As expected, the STR-overexpressed root cultures showed twofold higher accumulation of ajmalicine than the control. It was important to discover that overexpression of the single DXR or MECS gene from the MEP pathway also remarkably enhanced ajmalicine biosynthesis in transgenic hairy root cultures, and this suggested that engineering the MEP pathway by overexpression of DXR or MECS promoted the metabolic flux into ajmalicine biosynthesis. The transgenic hairy root cultures with co-overexpression of DXR and STR or MECS and STR had higher levels of ajmalicine than those with overexpression of a single gene alone such as DXR, MECS, and STR. It could be concluded that transgenic hairy root cultures harboring both DXR/ MECS and STR possessed an increased flux in the terpenoid indole alkaloid biosynthetic pathway that enhanced ajmalicine yield, which was more efficient than cultures harboring only one of the three genes. [ABSTRACT FROM AUTHOR]

The seeds of Herpetospermum pedunculosum (Ser.) C.B. Clarke, a well-known Tibetan medicine in China, are rich in kinds of bioactive lignans. In this phytochemical investigation on H. pedunculosum , sixteen undescribed lignans, named as herpedulins A - P together with 24 known ones were isolated from the ethyl acetate extract of its seeds. Their structures including the absolute configurations were determined by HR MS, 1D and 2D NMR experiments, and comparison of their experimental ECD spectra with calculated ones or literature data. High content screening experiments revealed that 9 compounds could promote the expression of farnesoid X receptor in guggulsterone-induced human normal liver cells L02 cells significantly. Further molecular docking results demonstrated that herpedulin E, J and K exhibited best docking scores (9.70, 9.28 and 10.31, respectively). Hydrogen bonding and hydrophobic interactions might contribute to the main interaction of active compounds with FXR. [Display omitted] • Sixteen undescribed compounds were isolated from Herpetospermum pedunculosum. • The absolute configurations were determined via quantum chemical calculations. • 9 compounds could promote FXR expression in guggulsterone-induced L02 cells. • Molecular interactions of compounds with FXR were analyzed by in silico methods. [ABSTRACT FROM AUTHOR]

[Display omitted] • A tyrosine aminotransferase gene (RcTAT) was cloned from Rhodiola crenulata. • RcTAT catalyzes the formation of 4-hydroxyphenylpyruvate from tyrosine. • Suppression of RcTAT significantly enhances the salidroside production. Rhodiola crenulata is a traditional Tibetan medicinal plant mainly distributed in high altitude areas of Tibet. As the main extract of R. crenulata , salidroside has important medicinal activities in enhancing the body's immunity and resisting microwave radiation by influencing cell metabolism. Tyrosine is the starting amino acid precursor for salidroside biosynthesis. As an aromatic amino acid, tyrosine participates not only in salidroside biosynthesis, but also in other metabolite biosynthesis which might compete against salidroside biosynthetic pathway. In this study, we functionally identified a tyrosine aminotransferase in R. crenulata (RcTAT) that could convert tyrosine into 4-hydroxyphenylpyruvate (4-HPP). The Km and Vmax values of RcTAT for tyrosine were respectively 0.21 mM and 0.17 μmol/min.mg under 9.0 pH and 39 °C. RNAi-mediated suppression of RcTAT expression markedly increased salidroside production in hairy root culture of R. crenulata. In summary, RcTAT is a key gene involved in the metabolic pathway that competes against salidroside biosynthesis, and suppressing its expression is a promising way to elevate salidroside production in planta. [ABSTRACT FROM AUTHOR]

• Glyphosate is used for screening transgenic plants of Atropa belladonna. • Homozygous transgenic plants overexpressing HnH6H and G2-EPSPS are obtained. • Homozygous transgenic plants have high-yield scopolamine and glyphosate resistance. Atropa belladonna , one of the most important medicinal plants in China, is used to produce anticholinergic tropane alkaloids. Weeds severely reduce the agricultural productivity of A. belladonna in fields, and the low content of high-value scopolamine in planta limits its economic value. In this study, homozygous lines of A. belladonna , with glyphosate resistance and high yield of scopolamine, were developed using metabolic engineering technology for the first time. Two homozygous transgenic lines (T2GH03 and T2GH06), harboring G2-EPSPS and HnH6H , were established and tested in the field. Glyphosate resistance assays demonstrated that the two homozygous transgenic lines could resist commercial recommended dose of glyphosate. Alkaloid analysis showed that the transgenic lines produced more scopolamine than wild-type plants. T2GH03 and T2GH06 produced scopolamine at the levels of 5.45 mg/g dry weight (DW) and 7.04 mg/g DW respectively in leaves, and 1.06 mg/g DW to 1.07 mg/g DW respectively in roots. Compared with wild-type plants, the scopolamine contents of homozygous transgenic lines increased by over 10 folds in leaves and nearly 5 folds in roots. To sum up, Atropa belladonna's new varieties with glyphosate resistance and high-yield scopolamine are developed, which are highly valuable for industrial production of scopolamine. [ABSTRACT FROM AUTHOR]

• An arabinofuranan, AQP70-3, was isolated from A. quinata for the first time. • The structure and morphology of AQP70-3 were characterized. • The absolute configuration of monosaccharide of AQP70-3 was characterized. • AQP70-3 showed immunomodulatory effects in cell level. • AQP70-3 was confirmed to possess immunomodulatory effects in zebrafish model. In our continuous searching for natural active polysaccharides with immunomodulatory activity, an arabinofuranan (AQP70-3) was isolated and purified from the fruits of Akebia quinata (Houtt.) Decne. by using ion-exchange chromatography and gel permeation chromatography for the first time. AQP70-3 contained both α - l -Ara f and β - l -Ara f , and the absolute molecular weight was 1.06 × 104 g/mol. The backbone of AQP70-3 comprised →5)- α - l-A ra f -(1→, →3,5)- α - l-A ra f -(1→, and →2,5)- α - l-A ra f -(1→, with branches of →1)- β - l-A ra f and →3)- α - l-A ra f -(1→ residues. Biological assay suggested that AQP70-3 can stimulate phagocytic activity and promote the levels of nitric oxide (NO), interleukin (IL)-6, IL-1 β , and tumor necrosis factor- α (TNF- α) of RAW264.7 cells. Furthermore, AQP70-3 was found to increase the production of reactive oxygen species (ROS) and NO in zebrafish embryo model. [ABSTRACT FROM AUTHOR]

• A root-expressed SAUR gene (AbSAUR1) was cloned from Atropa belladonna. • Overexpression of AbSAUR1 significantly increased the biomass of Atropa belladonna. • Overexpression of AbSAUR1 did not reduce biosynthesis of tropane alkaloids. • The yield of TAs was significantly improved in AbSAUR1 -overexpressing plants. Tropane alkaloids (TAs), including hyoscyamine, anisodamine and scopolamine, are widely used as anticholinergic drugs in the clinic. At present, the production of TAs is completely dependent on extraction from medicinal plants of Solanaceae. Atropa belladonna , commonly known as deadly nightshade, is a major commercial source of TAs. Previous studies have focused on increasing TA content by overexpressing TA biosynthesis genes. However, research on increasing TA yield by increasing the biomass of A. belladonna has not been reported. In this study, a Small Auxin Up-regulated RNA (SAUR) gene from A. belladonna (AbSAUR1) had been cloned for the first time. Based on heat map and tissue expression analysis, it was demonstrated that AbSAUR1 was specifically expressed in roots. Compared to control plants, transgenic lines of A. belladonna overexpressing AbSAUR1 showed a number of phenotypic changes, including increased plant height, thickened stems, increased number of branches, leaves and roots. Meanwhile, the biomass of AbSAUR1 -overexpressing plants also increased significantly, with fresh weight and dry weight being 3.79- and 3.20-fold higher than control plants, respectively. On average, there was no significant change in TA content, but the production of hyoscyamine, anisodamine and scopolamine was slightly enhanced. The overall yield of TAs was significantly improved, reaching 3.55 times that of the control. In summary, the AbSAUR1 gene can be employed as an effective molecular tool to increase the yield of TAs by increasing the biomass of A. belladonna. [ABSTRACT FROM AUTHOR]

AaPIF3, a bHLH transcription factor phylogenetically associated with Arabidopsis PIF3, positively regulates artemisinin biosynthesis in Artemisia annua. Overexpression of AaPIF3 significantly promotes the production of artemisinin and its related metabolites. • AaPIF3 is a bHLH transcription factor that is phylogenetically related with Arabidopsis PIF3. • AaPIF3 is able to transactivate artemisinin biosynthesis genes, according to dual-luciferase assays. • Overexpression of AaPIF3 markedly increased the production of artemisinin in Artemisia annua. Artemisia annua is a Chinese traditional herbal plant that produces artemisinin, the potent anti-malarial drug. It is a common goal for the artemisinin industry to develop plants of A. annua with high yields of artemisinin. In this study, a bHLH transcription factor of A. annua (AaPIF3) was characterized and used to promote the production of artemisinin. AaPIF3 was highly expressed in flower buds and glandular secretory trichomes and its coding protein was localized to the nucleus. Dual-luciferase assays indicated that AaPIF3 was able to significantly enhance the promoter activities of artemisinin biosynthesis genes, including ADS , CYP71AV1 , DBR2 , and ALDH1 , suggesting that AaPIF3 positively regulated artemisinin biosynthesis. Overexpression of AaPIF3 markedly upregulated artemisinin biosynthesis genes at the transcriptional level, and consequently the production of artemisinin was significantly promoted in transgenic A. annua plants. On the contrary, suppression of AaPIF3 led to significantly decreased transcript levels of artemisinin biosynthesis genes, with overall production of artemisinin significantly reduced. In summary, AaPIF3 plays a positive role in regulating artemisinin biosynthesis and transgenic plants of A. annua with high yields of artemisinin have been successfully developed through overexpression of AaPIF3. [ABSTRACT FROM AUTHOR]

Mirabilis himalaica (Edgew.) Heimerl is one of the most important genuine medicinal plants in Tibet, in which the special plateau habitat has been associated with its excellent medicinal quality and efficacy. However, the mechanisms by which environmental factors affect biosynthesis of secondary metabolic components remain unclear in this species. In this study, RNA sequencing and iTRAQ (isobaric Tags for Relative and Absolute Quantification) techniques were used to investigate the critical molecular "events" of rotenoid biosynthesis responding to UV-B radiation, a typical plateau ecological factor presented in native environment-grown M. himalaica plants. A total of 3641 differentially expressed genes (DEGs) and 106 differentially expressed proteins (DEPs) were identified in M. himalaica between UV-B treatment and control check (CK). Comprehensive analysis of protein and transcript data sets resulted in 14 and 7 DEGs from the plant hormone signal transduction and phosphatidylinositol signaling system pathways, respectively, being significantly enriched. The result showed that the plant hormone signal transduction and phosphatidylinositol signaling system might be the key metabolic strategy of UV-B radiation to improve the biosynthesis of rotenoid in M. himalaica. At same time, most of the DEGs were associated with auxin and calcium signaling, inferring that they might drive the downstream transmission of these signal transduction pathways. Regarding those pathways, two chalcone synthase enzymes, which play key roles in the biosynthesis of rotenoid that were thought as the representative medicinal component of M. himalaica, were significantly upregulated in UV-B radiation. This study provides a theoretical basis for further exploration of the adaptation mechanism of M. himalaica to UV-B radiation, and references for cultivation standardization. [ABSTRACT FROM AUTHOR]