Promiscuous Oxidosqualene Cyclases from Neoalsomitra integrifoliola Catalyzing the Formation of Tetracyclic, Pentacyclic, and Heterocyclic Triterpenes.

Six oxidosqualene cyclases (NiOSC1-NiOSC6) from Neoalsomitra integrifoliola were characterized for the biosynthesis of diverse triterpene scaffolds, including tetracyclic and pentacyclic triterpenes from the 2,3-oxidosqualene (1) and oxacyclic triterpenes from the 2,3:22,23-dioxidosqualene (2). NiOSC1 showed high efficiency in the production of naturally rare (20R)-epimers of oxacyclic triterpenes. Mutagenesis results revealed that the NiOSC1-F731G mutant significantly increased the yields of (20R)-epimers compared to the wild type. Homology modeling and molecular docking elucidated the origin of the (20R)-configuration in the epoxide addition step.

Biosynthetic pathway of prescription cucurbitacin IIa and high-level production of key triterpenoid intermediates in engineered yeast and tobacco.

Cucurbitacin IIa is a triterpenoid isolated exclusively from Hemsleya plants and a non-steroidal anti-inflammatory drug that functions as the main ingredient of prescription Hemslecin capsules and tablets in China. Synthetic biology provides new strategies for production of such valuable cucurbitacins at a large scale; however, the biosynthetic pathway of cucurbitacin IIa has been unknown, and the heterologous production of cucurbitacins in galactose medium has been expensive and low yielding. In this study, we characterized the functions of genes encoding two squalene epoxidases (HcSE1-2), six oxidosqualene cyclases (HcOSC1-6), two CYP450s (HcCYP87D20 and HcCYP81Q59), and an acyltransferase (HcAT1) in cucurbitacin IIa biosynthesis by heterologous expression in Saccharomyces cerevisiae and Nicotiana benthamiana. We achieved high-level production of the key cucurbitacin precursor 11-carbonyl-20ß-hydroxy-Cuol from glucose in yeast via modular engineering of the mevalonate pathway and optimization of P450 expression levels. The resulting yields of 46.41 mg/l 11-carbonyl-20ß-hydroxy-Cuol and 126.47 mg/l total cucurbitacin triterpenoids in shake flasks are the highest yields yet reported from engineered microbes. Subsequently, production of 11-carbonyl-20ß-hydroxy-Cuol by transient gene expression in tobacco resulted in yields of 1.28 mg/g dry weight in leaves. This work reveals the key genes involved in biosynthesis of prescription cucurbitacin IIa and demonstrates that engineered yeast cultivated with glucose can produce high yields of key triterpenoid intermediates. We describe a low-cost and highly efficient platform for rapid screening of candidate genes and high-yield production of pharmacological triterpenoids.

Integrated metabolome and transcriptome analyses reveal the molecular mechanism underlying dynamic metabolic processes during taproot development of Panax notoginseng.

BACKGROUND: Panax notoginseng (Burk) F. H. Chen is one of the most famous Chinese traditional medicinal plants. The taproot is the main organ producing triterpenoid saponins, and its development is directly linked to the quality and yield of the harvested P. notoginseng. However, the mechanisms underlying the dynamic metabolic changes occurring during taproot development of P. notoginseng are unknown. RESULTS: We carried out metabolomic and transcriptomic analyses to investigate metabolites and gene expression during the development of P. notoginseng taproots. The differentially accumulated metabolites included amino acids and derivatives, nucleotides and derivatives, and lipids in 1-year-old taproots, flavonoids and terpenoids in 2- and 3-year-old taproots, and phenolic acids in 3-year-old taproots. The differentially expressed genes (DEGs) are related to phenylpropanoid biosynthesis, metabolic pathway and biosynthesis of secondary metabolites at all three developmental stages. Integrative analysis revealed that the phenylpropanoid biosynthesis pathway was involved in not only the development of but also metabolic changes in P. notoginseng taproots. Moreover, significant accumulation of triterpenoid saponins in 2- and 3-year-old taproots was highly correlated with the up-regulated expression of cytochrome P450s and uridine diphosphate-dependent glycosyltransferases genes. Additionally, a gene encoding RNase-like major storage protein was identified to play a dual role in the development of P. notoginseng taproots and their triterpenoid saponins synthesis. CONCLUSIONS: These results elucidate the molecular mechanism underlying the accumulation of and change relationship between primary and secondary metabolites in P. notoginseng taproots, and provide a basis for the quality control and genetic improvement of P. notoginseng.

TRPM2 enhances ischemic excitotoxicity by associating with PKCγ.

N-methyl-D-aspartate receptor (NMDAR)-mediated glutamate excitotoxicity significantly contributes to ischemic neuronal death and post-recanalization infarction expansion. Despite tremendous efforts, targeting NMDARs has proven unsuccessful in clinical trials for mitigating brain injury. Here, we show the discovery of an interaction motif for transient receptor potential melastatin 2 (TRPM2) and protein kinase Cγ (PKCγ) association and demonstrate that TRPM2-PKCγ uncoupling is an effective therapeutic strategy for attenuating NMDAR-mediated excitotoxicity in ischemic stroke. We demonstrate that the TRPM2-PKCγ interaction allows TRPM2-mediated Ca2+ influx to promote PKCγ activation, which subsequently enhances TRPM2-induced potentiation of extrasynaptic NMDAR (esNMDAR) activity. By identifying the PKCγ binding motif on TRPM2 (M2PBM), which directly associates with the C2 domain of PKCγ, an interfering peptide (TAT-M2PBM) is developed to disrupt TRPM2-PKCγ interaction without compromising PKCγ function. M2PBM deletion or TRPM2-PKCγ dissociation abolishes both TRPM2-PKCγ and TRPM2-esNMDAR couplings, resulting in reduced excitotoxic neuronal death and attenuated ischemic brain injury.

Eutypellaolides A-J, Sesquiterpene diversity expansion of the polar fungus Eutypella sp. D-1.

Eight new 12,8-eudesmanolide sesquiterpenes, eutypellaolides A-H (1-8), and two new eudesmane-type sesquiterpenes, eutypellaolides I-J (9-10), along with four known 12,8-eudesmanolide compounds 11-14, were isolated from the culture extract of the polar fungus Eutypella sp. D-1 by one strain many compounds (OSMAC) approach. The structures of these compounds were determined through comprehensive spectroscopic data and experimental and calculated ECD analysis. Antibacterial, immunosuppressive, and PTP1B inhibition activities of these compounds were evaluated. Compounds 1 and 11 exhibited strong inhibitory activities against Bacillus subtilis and Staphylococcus aureus, with each showing an MIC value of 2 µg/mL. Compound 9 displayed weak immunosuppressive activity against ConA-induced T-cell proliferation with an inhibitory rate of 61.7% at a concentration of 19.8 µM. Compounds 5, 11, and 14 exhibited weak PTP1B inhibition activities with IC50 values of 44.8, 43.2, and 49.5 µM, respectively.

Proteomics analysis of histone deacetylase inhibitor-resistant solid tumors reveals resistant signatures and potential drug combinations.

Histone deacetylase inhibitors (HDACis) are important drugs for cancer therapy, but the indistinct resistant mechanisms of solid tumor therapy greatly limit their clinical application. In this study we conducted HDACi-perturbated proteomics and phosphoproteomics analyses in HDACi-sensitive and -resistant cell lines using a tandem mass tag (TMT)-based quantitative proteomic strategy. We found that the ribosome biogenesis proteins MRTO4, PES1, WDR74 and NOP16 vital to tumorigenesis might regulate the tumor sensitivity to HDACi. By integrating HDACi-perturbated protein signature with previously reported proteomics and drug sensitivity data, we predicted and validated a series of drug combination pairs potentially to enhance the sensitivity of HDACi in diverse solid tumor. Functional phosphoproteomic analysis further identified the kinase PDK1 and ROCK as potential HDACi-resistant signatures. Overall, this study reveals the potential HDACi-resistant signatures and may provide promising drug combination strategies to attenuate the resistance of solid tumor to HDACi.

Combined ATAC-seq, RNA-seq, and GWAS analysis reveals glycogen metabolism regulatory network in Jinjiang oyster ( Crassostrea ariakensis).

Glycogen serves as the principal energy reserve for metabolic processes in aquatic shellfish and substantially contributes to the flavor and quality of oysters. The Jinjiang oyster ( Crassostrea ariakensis) is an economically and ecologically important species in China. In the present study, RNA sequencing (RNA-seq) and assay for transposase-accessible chromatin using sequencing (ATAC-seq) were performed to investigate gene expression and chromatin accessibility variations in oysters with different glycogen contents. Analysis identified 9 483 differentially expressed genes (DEGs) and 7 215 genes with significantly differential chromatin accessibility (DCAGs) were obtained, with an overlap of 2 600 genes between them. Notably, a significant proportion of these genes were enriched in pathways related to glycogen metabolism, including "Glycogen metabolic process" and "Starch and sucrose metabolism". In addition, genome-wide association study (GWAS) identified 526 single nucleotide polymorphism (SNP) loci associated with glycogen content. These loci corresponded to 241 genes, 63 of which were categorized as both DEGs and DCAGs. This study enriches basic research data and provides insights into the molecular mechanisms underlying the regulation of glycogen metabolism in C. ariakensis.

Integrated metabolome and transcriptome analysis reveals the regulatory mechanism of low nitrogen-driven biosynthesis of saponins and flavonoids in Panax notoginseng.

BACKGROUND: Nitrogen (N) is an important macronutrient involved in the biosynthesis of primary and secondary metabolites in plants. However, the metabolic regulatory mechanism of low-N-induced triterpenoid saponin and flavonoid accumulation in rhizomatous medicinal Panax notoginseng (Burk.) F. H. Chen remains unclear. METHODS: To explore the potential regulatory mechanism and metabolic basis controlling the response of P. notoginseng to N deficiency, the transcriptome and metabolome were analysed in the roots. RESULTS: The N content was significantly reduced in roots of N0-treated P. notoginseng (0 kg·N·667 m-2). The C/N ratio was enhanced in the N-deficient P. notoginseng. N deficiency promotes the accumulation of amino acids (L-proline, L-leucine, L-isoleucine, L-norleucine, L-arginine, and L-citrulline) and sugar (arabinose, xylose, glucose, fructose, and mannose), thus providing precursor metabolites for the biosynthesis of flavonoids and triterpenoid saponins. Downregulation of key structural genes (PAL, PAL3, ACC1, CHS2, PPO, CHI3, F3H, DFR, and FGT), in particular with the key genes of F3H, involved in the flavonoid biosynthesis pathway possibly induced the decrease in flavonoid content with increased N supply. Notoginsenoside R1, ginsenoside Re, Rg1, Rd, F1, R1 + Rg1 + Rb1 and total triterpenoid saponins were enhanced in the N0 groups than in the N15 (15 kg·N·667 m-2) plants. Higher phosphoenolpyruvate (an intermediate of glycolyticwith pathway metabolism) and serine (an intermediate of photorespiration) levels induced by N deficiency possibly promote saponin biosynthesis through mevalonic acid (MVA) and methylerythritol (MEP) pathways. Genes (MVD2, HMGS, HMGR1, HMGR2, DXR, and HMGR1) encoding the primary enzymes HMGS, HMGR, DXR, and MVD in the MVA and MEP pathways were significantly upregulated in the N0-treated P. notoginseng. The saponin biosynthesis genes DDS, DDS, CYP716A52, CYP716A47, UGT74AE2, and FPS were upregulated in the N-deficient plants. Upregulation of genes involved in saponin biosynthesis promotes the accumulation of triterpenoid saponins in the N0-grown P. notoginseng. CONCLUSIONS: N deficiency enhances primary metabolisms, such as amino acids and sugar accumulation, laying the foundation for the synthesis of flavonoids and triterpenoid saponins in P. notoginseng. F3H, DDS, FPS, HMGR, HMGS and UGT74AE2 can be considered as candidates for functional characterisation of the N-regulated accumulation of triterpenoid saponins and flavonoids in future.

Phyllofenones F-M, Scalarane Sesterterpenes from the Marine Sponge Phyllospongia foliascens.

Eight new scalarane sesterterpenes, phyllofenones F-M (1-8), together with two known analogues, carteriofenones B and A (9-10), were isolated from the marine sponge Phyllospongia foliascens collected from the South China Sea. The structures of these compounds were determined based on extensive spectroscopic and quantum chemical calculation analysis. The antibacterial and cytotoxic activity of these compounds was evaluated. Among them, only compounds 4 and 6 displayed weak inhibitory activity against Staphylococcus aureus and Escherichia coli, with MIC values of 16 µg/mL and 8 µg/mL, respectively. Compounds 1-10 exhibited cytotoxic activity against the HeLa, HCT-116, H460, and SW1990 cancer cell lines, with IC50 values ranging from 3.4 to 19.8 µM.

Reconstruction of engineered yeast factory for high yield production of ginsenosides Rg3 and Rd.

Panax notoginseng is one of the most valuable traditional Chinese herbs. The main active ingredients, dammarane-type ginsenosides, show multiple pharmacological activities. Recently, the key UDP-dependent glycosyltransferases (UGTs) involved in the biosynthesis of common ginsenosides have been widely studied. However, only a few UGTs that catalyze ginsenoside formation have been reported. This study further investigated the new catalytic function of 10 characterized UGTs from the public database. PnUGT31(PnUGT94B2) and PnUGT53 (PnUGT71B8)exhibited promiscuous sugar-donor specificity of UDP-glucose and UDP-xylose, which could catalyze the glycosylation of C20-OH sites and elongation of the sugar chain at the C3 and/or C20 sites. We further analyzed the expression patterns in P. notoginseng and predicted the catalytic mechanisms of PnUGT31 and PnUGT53 using molecular docking simulations. Moreover, different gene modules were built to increase the yield of ginsenosides in engineered yeast. The metabolic flow of the proginsenediol (PPD) synthetic pathway was enhanced by LPPDS gene modules based on the engineered strain. The resulting yeast was constructed to produce 1.72 g/L PPD in a shaking flask, but cell growth was significantly inhibited. EGH and LKG gene modules were constructed to achieve high-level production of dammarane-type ginsenosides. The production of G-Rg3 controlled by LKG modules increased 3.84 times (254.07 mg/ L), whereas the G-Rd titer reached 56.68 mg/L after 96 h in shaking flask culture under the control of all modules, both of which yielded the highest values for known microbes.

Cytosporin Derivatives from Arctic-Derived Fungus Eutypella sp. D-1 via the OSMAC Approach.

A chemical investigation of the Arctic-derived fungus Eutypella sp. D-1 based on the OSMAC (one strain many compounds) approach resulted in the isolation of five cytosporin polyketides (compounds 1-3 and 11-12) from rice medium and eight cytosporins (compounds 2 and 4-11) from solid defined medium. The structures of the seven new compounds, eutypelleudesmane A (1), cytosporin Y (2), cytosporin Z (3), cytosporin Y1 (4), cytosporin Y2 (5), cytosporin Y3 (6), and cytosporin E1 (7), were elucidated by analyzing their detailed spectroscopic data. Structurally, cytosporin Y1 (4) may be a key intermediate in the biosynthesis of the isolated cytosporins, rather than an end product. Compound 1 contained a unique skeleton formed by the ester linkage of two moieties, cytosporin F (12) and the eudesmane-type sesquiterpene dihydroalanto glycol. Additionally, the occurrence of cyclic carbonate moieties in compounds 6 and 7 was found to be rare in nature. The antibacterial, immunosuppressive, and cytotoxic activities of all compounds derived from Eutypella sp. D-1 were evaluated. Unfortunately, only compounds 3, 6, 8, and 10-11 displayed immunosuppressive activity, with inhibitory rates of 62.9%, 59.5%, 67.8%, 55.8%, and 68.7%, respectively, at a concentration of 5 µg/mL.

Phyllospongianes A-E, Dinorscalarane Sesterterpenes from the Marine Sponge Phyllospongia foliascens.

Phyllospongianes A-E (1-5), five new scalarane derivatives featuring an unprecedented 6/6/6/5 tetracyclic dinorscalarane scaffold, along with the known probable biogenetic precursor, 12-deacetylscalaradial (6), were isolated from the marine sponge Phyllospongia foliascens. The structures of the isolated compounds were determined by analysis of spectroscopic data and electronic circular dichroism experiments. Compounds 1-5 are the first 6/6/6/5 tetracyclic scalarane derivatives to be reported within the scalarane family. Compounds 1, 2, and 4 exhibited antibacterial activity against Vibrio vulnificus, Vibrio parahemolyticus, Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, Bacillus subtilis, and Pseudomonas aeruginosa with MIC values ranging from 1 to 8 µg/mL. Furthermore, compound 3 exhibited significant cytotoxic activity on MDA-MB-231, HepG2, C4-2-ENZ, MCF-7, H460, and HT-29 cancer cell lines with IC50 values in the range between 0.7 and 13.2 µM.

Phylogeny-Related Variations in Venomics: A Test in a Subset of Habu Snakes (Protobothrops).

We conducted a comparative analysis to unveil the divergence among venoms from a subset of Old World habu snakes (Protobothrops) in terms of venomic profiles and toxicological and enzymatic activities. A total of 14 protein families were identified in the venoms from these habu snakes, and 11 of them were shared among these venoms. The venoms of five adult habu snakes were overwhelmingly dominated by SVMP (32.56 ± 13.94%), PLA2 (22.93 ± 9.26%), and SVSP (16.27 ± 4.79%), with a total abundance of over 65%, while the subadult P. mangshanensis had an extremely low abundance of PLA2 (1.23%) but a high abundance of CTL (51.47%), followed by SVMP (22.06%) and SVSP (10.90%). Apparent interspecific variations in lethality and enzymatic activities were also explored in habu snake venoms, but no variations in myotoxicity were found. Except for SVSP, the resemblance of the relatives within Protobothrops in other venom traits was estimated to deviate from Brownian motion evolution based on phylogenetic signals. A comparative analysis further validated that the degree of covariation between phylogeny and venom variation is evolutionarily labile and varies among clades of closely related snakes. Our findings indicate a high level of interspecific variation in the venom proteomes of habu snakes, both in the presence or absence and the relative abundance of venom protein families, and that these venoms might have evolved under a combination of adaptive and neutral mechanisms.

Identification and characterization of new proteins crucial for bacterial spore resistance and germination.

2Duf, named after the presence of a transmembrane (TM) Duf421 domain and a small Duf1657 domain in its sequence, is likely located in the inner membrane (IM) of spores in some Bacillus species carrying a transposon with an operon termed spoVA 2mob. These spores are known for their extreme resistance to wet heat, and 2Duf is believed to be the primary contributor to this trait. In this study, we found that the absence of YetF or YdfS, both Duf421 domain-containing proteins and found only in wild-type (wt) B. subtilis spores with YetF more abundant, leads to decreased resistance to wet heat and agents that can damage spore core components. The IM phospholipid compositions and core water and calcium-dipicolinic acid levels of YetF-deficient spores are similar to those of wt spores, but the deficiency could be restored by ectopic insertion of yetF, and overexpression of YetF increased wt spore resistance to wet heat. In addition, yetF and ydfS spores have decreased germination rates as individuals and populations with germinant receptor-dependent germinants and increased sensitivity to wet heat during germination, potentially due to damage to IM proteins. These data are consistent with a model in which YetF, YdfS and their homologs modify IM structure to reduce IM permeability and stabilize IM proteins against wet heat damage. Multiple yetF homologs are also present in other spore forming Bacilli and Clostridia, and even some asporogenous Firmicutes, but fewer in asporogenous species. The crystal structure of a YetF tetramer lacking the TM helices has been reported and features two distinct globular subdomains in each monomer. Sequence alignment and structure prediction suggest this fold is likely shared by other Duf421-containing proteins, including 2Duf. We have also identified naturally occurring 2duf homologs in some Bacilli and Clostridia species and in wt Bacillus cereus spores, but not in wt B. subtilis. Notably, the genomic organization around the 2duf gene in most of these species is similar to that in spoVA 2mob, suggesting that one of these species was the source of the genes on this operon in the extremely wet heat resistant spore formers.

A search and rescue robot search method based on flower pollination algorithm and Q-learning fusion algorithm.

Search algorithm plays an important role in the motion planning of the robot, it determines whether the mobile robot complete the task. To solve the search task in complex environments, a fusion algorithm based on the Flower Pollination algorithm and Q-learning is proposed. To improve the accuracy, an improved grid map is used in the section of environment modeling to change the original static grid to a combination of static and dynamic grids. Secondly, a combination of Q-learning and Flower Pollination algorithm is used to complete the initialization of the Q-table and accelerate the efficiency of the search and rescue robot path search. A combination of static and dynamic reward function is proposed for the different situations encountered by the search and rescue robot during the search process, as a way to allow the search and rescue robot to get better different feedback results in each specific situation. The experiments are divided into two parts: typical and improved grid map path planning. Experiments show that the improved grid map can increase the success rate and the FIQL can be used by the search and rescue robot to accomplish the task in a complex environment. Compared with other algorithms, FIQL can reduce the number of iterations, improve the adaptability of the search and rescue robot to complex environments, and have the advantages of short convergence time and small computational effort.

Investigation of targets and anticancer mechanisms of covalently acting natural products by functional proteomics.

Eriocalyxin B (EB), 17-hydroxy-jolkinolide B (HJB), parthenolide (PN), xanthatin (XT) and andrographolide (AG) are terpenoid natural products with a variety of promising antitumor activities, which commonly bear electrophilic groups (α,ß-unsaturated carbonyl groups and/or epoxides) capable of covalently modifying protein cysteine residues. However, their direct targets and underlying molecular mechanisms are still largely unclear, which limits the development of these compounds. In this study, we integrated activity-based protein profiling (ABPP) and quantitative proteomics approach to systematically characterize the covalent targets of these natural products and their involved cellular pathways. We first demonstrated the anti-proliferation activities of these five compounds in triple-negative breast cancer cell MDA-MB-231. Tandem mass tag (TMT)-based quantitative proteomics showed all five compounds commonly affected the ubiquitin mediated proteolysis pathways. ABPP platform identified the preferentially modified targets of EB and PN, two natural products with high anti-proliferation activity. Biochemical experiments showed that PN inhibited the cell proliferation through targeting ubiquitin carboxyl-terminal hydrolase 10 (USP10). Together, this study uncovered the covalently modified targets of these natural products and potential molecular mechanisms of their antitumor activities.

Crystal structure of the CoV-Y domain of SARS-CoV-2 nonstructural protein 3.

Replication of the coronavirus genome starts with the formation of viral RNA-containing double-membrane vesicles (DMV) following viral entry into the host cell. The multi-domain nonstructural protein 3 (nsp3) is the largest protein encoded by the known coronavirus genome and serves as a central component of the viral replication and transcription machinery. Previous studies demonstrated that the highly-conserved C-terminal region of nsp3 is essential for subcellular membrane rearrangement, yet the underlying mechanisms remain elusive. Here we report the crystal structure of the CoV-Y domain, the most C-terminal domain of the SARS-CoV-2 nsp3, at 2.4 Å-resolution. CoV-Y adopts a previously uncharacterized V-shaped fold featuring three distinct subdomains. Sequence alignment and structure prediction suggest that this fold is likely shared by the CoV-Y domains from closely related nsp3 homologs. NMR-based fragment screening combined with molecular docking identifies surface cavities in CoV-Y for interaction with potential ligands and other nsps. These studies provide the first structural view on a complete nsp3 CoV-Y domain, and the molecular framework for understanding the architecture, assembly and function of the nsp3 C-terminal domains in coronavirus replication. Our work illuminates nsp3 as a potential target for therapeutic interventions to aid in the on-going battle against the COVID-19 pandemic and diseases caused by other coronaviruses.

Structure of the Sec14 domain of Kalirin reveals a distinct class of lipid-binding module in RhoGEFs.

Gated entry of lipophilic ligands into the enclosed hydrophobic pocket in stand-alone Sec14 domain proteins often links lipid metabolism to membrane trafficking. Similar domains occur in multidomain mammalian proteins that activate small GTPases and regulate actin dynamics. The neuronal RhoGEF Kalirin, a central regulator of cytoskeletal dynamics, contains a Sec14 domain (KalbSec14) followed by multiple spectrin-like repeats and catalytic domains. Previous studies demonstrated that Kalirin lacking its Sec14 domain fails to maintain cell morphology or dendritic spine length, yet whether and how KalbSec14 interacts with lipids remain unknown. Here, we report the structural and biochemical characterization of KalbSec14. KalbSec14 adopts a closed conformation, sealing off the canonical ligand entry site, and instead employs a surface groove to bind a limited set of lysophospholipids. The low-affinity interactions of KalbSec14 with lysolipids are expected to serve as a general model for the regulation of Rho signaling by other Sec14-containing Rho activators.

Transfer performance of gated recurrent unit model for runoff prediction based on the comprehensive spatiotemporal similarity of catchments.

Accurate runoff prediction in data-poor catchments is important for water resource management, flood mitigation, environmental protection, and other tasks. One possible solution is to transfer a runoff prediction model constructed by using a machine learning model for gauged catchments to data-poor catchments. However, the transfer of runoff prediction model must consider the comprehensive spatiotemporal similarities between the catchments; otherwise, the transfer performance can be massively uncertain. Therefore, to improve the accuracy of runoff prediction and eliminate the uncertainty in identifying the differences between catchment environments, this paper proposes a novel measurement approach of comprehensive spatiotemporal similarity. This approach measures the similarities among catchments by fully considering which of the various catchments' spatiotemporal attributes can better represent the geographical similarity. Then, according to the similarities between the catchments, a runoff prediction model trained in gauged catchments is transformed for the most similar data-poor catchments to predict the runoff and the transfer performance is analyzed. To this end, a runoff prediction model is built using a gated recurrent unit (GRU) network based on the CAMELS catchments data set. A framework to extract the comprehensive spatiotemporal features of catchments is designed using three autoencoders. The catchments' similarities can be measured, further, and their spatiotemporal attributes determined once a measurement model of comprehensive spatiotemporal similarity is constructed. Finally, the transfer performance of the GRU runoff prediction model based on comprehensive spatiotemporal and other geographical similarities is evaluated and analyzed. The experimental results demonstrate that the proposed method outperforms comparable approaches.

Biosynthetic pathway of prescription bergenin from Bergenia purpurascens and Ardisia japonica.

Bergenin is a typical carbon glycoside and the primary active ingredient in antitussive drugs widely prescribed for central cough inhibition in China. The bergenin extraction industry relies on the medicinal plant species Bergenia purpurascens and Ardisia japonica as their resources. However, the bergenin biosynthetic pathway in plants remains elusive. In this study, we functionally characterized a shikimate dehydrogenase (SDH), two O-methyltransferases (OMTs), and a C-glycosyltransferase (CGT) involved in bergenin synthesis through bioinformatics analysis, heterologous expression, and enzymatic characterization. We found that BpSDH2 catalyzes the two-step dehydrogenation process of shikimic acid to form gallic acid (GA). BpOMT1 and AjOMT1 facilitate the methylation reaction at the 4-OH position of GA, resulting in the formation of 4-O-methyl gallic acid (4-O-Me-GA). AjCGT1 transfers a glucose moiety to C-2 to generate 2-Glucosyl-4-O-methyl gallic acid (2-Glucosyl-4-O-Me-GA). Bergenin production ultimately occurs in acidic conditions or via dehydration catalyzed by plant dehydratases following a ring-closure reaction. This study for the first time uncovered the biosynthetic pathway of bergenin, paving the way to rational production of bergenin in cell factories via synthetic biology strategies.