TransGEM: a molecule generation model based on Transformer with gene expression data.

MOTIVATION: It is difficult to generate new molecules with desirable bioactivity through ligand-based de novo drug design, and receptor-based de novo drug design is constrained by disease target information availability. The combination of artificial intelligence and phenotype-based de novo drug design can generate new bioactive molecules, independent from disease target information. Gene expression profiles can be used to characterize biological phenotypes. The Transformer model can be utilized to capture the associations between gene expression profiles and molecular structures due to its remarkable ability in processing contextual information. RESULTS: We propose TransGEM (Transformer-based model from gene expression to molecules), which is a phenotype-based de novo drug design model. A specialized gene expression encoder is used to embed gene expression difference values between diseased cell lines and their corresponding normal tissue cells into TransGEM model. The results demonstrate that the TransGEM model can generate molecules with desirable evaluation metrics and property distributions. Case studies illustrate that TransGEM model can generate structurally novel molecules with good binding affinity to disease target proteins. The majority of genes with high attention scores obtained from TransGEM model are associated with the onset of the disease, indicating the potential of these genes as disease targets. Therefore, this study provides a new paradigm for de novo drug design, and it will promote phenotype-based drug discovery. AVAILABILITY AND IMPLEMENTATION: The code is available at https://github.com/hzauzqy/TransGEM.

Metabolites of pathogenic microorganisms database (MPMdb) and its seed metabolite applications.

Seed metabolites are the combination of essential compounds required by an organism across various potential environmental conditions. The seed metabolites screening framework based on the network topology approach can capture important biological information of species. This study aims to identify comprehensively the relationship between seed metabolites and pathogenic bacteria. A large-scale data set was compiled, describing the seed metabolite sets and metabolite sets of 124,192 pathogenic strains from 34 genera, by constructing genome-scale metabolic models. The enrichment analysis method was used to screen the specific seed metabolites of each species/genus of pathogenic bacteria. The metabolites of pathogenic microorganisms database (MPMdb) (http://qyzhanglab.hzau.edu.cn/MPMdb/) was established for browsing, searching, predicting, or downloading metabolites and seed metabolites of pathogenic microorganisms. Based on the MPMdb, taxonomic and phylogenetic analyses of pathogenic bacteria were performed according to the function of seed metabolites and metabolites. The results showed that the seed metabolites could be used as a feature for microorganism chemotaxonomy, and they could mirror the phylogeny of pathogenic bacteria. In addition, our screened specific seed metabolites of pathogenic bacteria can be used not only for further tapping the nutritional resources and identifying auxotrophies of pathogenic bacteria but also for designing targeted bactericidal compounds by combining with existing antimicrobial agents.IMPORTANCEMetabolites serve as key communication links between pathogenic microorganisms and hosts, with seed metabolites being crucial for microbial growth, reproduction, external communication, and host infection. However, the large-scale screening of metabolites and the identification of seed metabolites have always been the main technical bottleneck due to the low throughput and costly analysis. Genome-scale metabolic models have become a recognized research paradigm to investigate the metabolic characteristics of species. The developed metabolites of pathogenic microorganisms database in this study is committed to systematically predicting and identifying the metabolites and seed metabolites of pathogenic microorganisms, which could provide a powerful resource platform for pathogenic bacteria research.

Transporter proteins knowledge graph construction and its application in drug development.

Transporters are the main determinant for pharmacokinetics characteristics of drugs, such as absorption, distribution, and excretion of drugs in humans. However, it is difficult to perform drug transporter validation and structure analysis of membrane transporter proteins by experimental methods. Many studies have demonstrated that knowledge graphs (KG) could effectively excavate potential association information between different entities. To improve the effectiveness of drug discovery, a transporter-related KG was constructed in this study. Meanwhile, a predictive frame (AutoInt_KG) and a generative frame (MolGPT_KG) were established based on the heterogeneity information obtained from the transporter-related KG by the RESCAL model. Natural product Luteolin with known transporters was selected to verify the reliability of the AutoInt_KG frame, its ROC-AUC (1:1), ROC-AUC (1:10), PR-AUC (1:1), PR-AUC (1:10) are 0.91, 0.94, 0.91 and 0.78, respectively. Subsequently, the MolGPT_KG frame was constructed to implement efficient drug design based on transporter structure. The evaluation results showed that the MolGPT_KG could generate novel and valid molecules and that these molecules were further confirmed by molecular docking analysis. The docking results showed that they could bind to important amino acids at the active site of the target transporter. Our findings will provide rich information resources and guidance for the further development of the transporter-related drugs.

The Holothuria leucospilota genome elucidates sacrificial organ expulsion and bioadhesive trap enriched with amyloid-patterned proteins.

Some tropical sea cucumbers of the family Holothuriidae can efficiently repel or even fatally ensnare predators by sacrificially ejecting a bioadhesive matrix termed the Cuvierian organ (CO), so named by the French zoologist Georges Cuvier who first described it in 1831. Still, the precise mechanisms for how adhesiveness genetically arose in CO and how sea cucumbers perceive and transduce danger signals for CO expulsion during defense have remained unclear. Here, we report the first high-quality, chromosome-level genome assembly of Holothuria leucospilota, an ecologically significant sea cucumber with prototypical CO. The H. leucospilota genome reveals characteristic long-repeat signatures in CO-specific outer-layer proteins, analogous to fibrous proteins of disparate species origins, including spider spidroin and silkworm fibroin. Intriguingly, several CO-specific proteins occur with amyloid-like patterns featuring extensive intramolecular cross-ß structures readily stainable by amyloid indicator dyes. Distinct proteins within the CO connective tissue and outer surface cooperate to give the expelled matrix its apparent tenacity and adhesiveness, respectively. Genomic evidence offers further hints that H. leucospilota directly transduces predator-induced mechanical pressure onto the CO surface through mediation by transient receptor potential channels, which culminates in acetylcholine-triggered CO expulsion in part or in entirety. Evolutionarily, innovative events in two distinct regions of the H. leucospilota genome have apparently spurred CO's differentiation from the respiratory tree to a lethal defensive organ against predators.

Pacific white shrimp (Litopenaeus vannamei) vitelline membrane outer layer protein 1 (VMO1) is produced in the hepatopancreas and transported into ovarian oocytes during vitellogenesis.

In birds, vitelline membrane outer layer protein 1 (VMO1) is an exogenous protein that can be absorbed by eggs as a barrier to prevent the mixing of yolk and egg white. However, researches on VMO1 are limited in birds but not other non-avian species until now. In this study, we first identified a novel Vmo1 cDNA (Lv-Vmo1) in Pacific white shrimp (Litopenaeus vannamei), the most important cultured shrimp in the world. We further analyzed its gene organization, phylogenetic relationship and protein structure. The Lv-Vmo1 transcript was specifically expressed in the hepatopancreas without sexual dimorphism. During ovarian development in female, the hepatopancreatic Lv-Vmo1 mRNA levels showed a significant increase. By in situ hybridization, Lv-Vmo1 mRNA was present in three cell types of the hepatopancreas but neither oocytes nor follicle cells of the ovary. In contrast, immunofluorescence revealed that Lv-VMO1 protein was distributed in the cytoplasms of both hepatopancreatic cells and ovarian oocytes. Western blot showed that Lv-VMO1 protein was produced in the hepatopancreas and transported to the ovary via hemolymph circulation. Identification of a species-specific egg-entry guide protein is the key to the receptor-mediated ovarian transduction of cargo, a novel gene editing approach in oviparous animals. This study lays the mechanism for exogenous transport into penaeid shrimp eggs by VMO1, as a foundation for achieving exogenous protein-mediated incorporation into oocytes.

Reconstructed Genome-Scale Metabolic Model Characterizes Adaptive Metabolic Flux Changes in Peripheral Blood Mononuclear Cells in Severe COVID-19 Patients.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) poses a mortal threat to human health. The elucidation of the relationship between peripheral immune cells and the development of inflammation is essential for revealing the pathogenic mechanism of COVID-19 and developing related antiviral drugs. The immune cell metabolism-targeting therapies exhibit a desirable anti-inflammatory effect in some treatment cases. In this study, based on differentially expressed gene (DEG) analysis, a genome-scale metabolic model (GSMM) was reconstructed by integrating transcriptome data to characterize the adaptive metabolic changes in peripheral blood mononuclear cells (PBMCs) in severe COVID-19 patients. Differential flux analysis revealed that metabolic changes such as enhanced aerobic glycolysis, impaired oxidative phosphorylation, fluctuating biogenesis of lipids, vitamins (folate and retinol), and nucleotides played important roles in the inflammation adaptation of PBMCs. Moreover, the main metabolic enzymes such as the solute carrier (SLC) family 2 member 3 (SLC2A3) and fatty acid synthase (FASN), responsible for the reactions with large differential fluxes, were identified as potential therapeutic targets. Our results revealed the inflammation regulation potentials of partial metabolic reactions with differential fluxes and their metabolites. This study provides a reference for developing potential PBMC metabolism-targeting therapy strategies against COVID-19.

Detecting plasma hsa_circ_0061276 in patients with gastric cancer by reverse transcription-digital polymerase chain reaction.

Background: The role of circular RNAs (circRNAs) in the occurrence of gastric cancer is still unclear. Therefore, the diagnostic value and mechanisms underlying hsa_circ_0061276 in the occurrence of gastric cancer were explored. Methods: Reverse transcription-droplet digital polymerase chain reaction was used to detect the copy number of hsa_circ_0061276 in plasma from healthy individuals, as well as from patients with gastric precancerous lesions or early-stage or advanced gastric cancer. Plasmids overexpressing or knocking down hsa_circ_0061276 expression were transfected into gastric cancer cells. The effects on the growth, migration, and cell cycle distribution of gastric cancer cells were then analyzed. Finally, miRanda and RNAhybrid were used to explore the binding sites between hsa_circ_0061276 and microRNAs (miRNAs). A double luciferase reporter gene assay was used to confirm the miRNA sponge effect. Results: The results show that plasma hsa_circ_0061276 copy number showed a trend of a gradual decrease when comparing healthy controls to the early cancer group and advanced gastric cancer group. Overexpression of hsa_circ_0061276 inhibited the growth and migration of gastric cancer cells. Through bioinformatic analyses combined with cellular experiments, it was found that hsa_circ_0061276 inhibited the growth of gastric cancer by binding to hsa-miR-7705. Conclusion: Hsa_circ_0061276 may be a new biomarker for gastric cancer. The tumor suppressor role of hsa_circ_0061276 on gastric cancer likely occurs through a sponge effect on miRNAs such as hsa-miR-7705.

Agent Clustering Strategy Based on Metabolic Flux Distribution and Transcriptome Expression for Novel Drug Development.

The network module-based method has been used for drug repositioning. The traditional drug repositioning method only uses the gene characteristics of the drug but ignores the drug-triggered metabolic changes. The metabolic network systematically characterizes the connection between genes, proteins, and metabolic reactions. The differential metabolic flux distribution, as drug metabolism characteristics, was employed to cluster the agents with similar MoAs (mechanism of action). In this study, agents with the same pharmacology were clustered into one group, and a total of 1309 agents from the CMap database were clustered into 98 groups based on differential metabolic flux distribution. Transcription factor (TF) enrichment analysis revealed the agents in the same group (such as group 7 and group 26) were confirmed to have similar MoAs. Through this agent clustering strategy, the candidate drugs which can inhibit (Japanese encephalitis virus) JEV infection were identified. This study provides new insights into drug repositioning and their MoAs.

FN3 Domain Displaying Double Epitopes: A Cost-Effective Strategy for Producing Substitute Antigens.

Construction of substitute antigens based on alternative scaffold proteins is a promising strategy in bioassay technology. In this study, we proposed a strategy for constructing substitute antigens derived from 10th human fibronectin type III (FN3) using two peptide epitopes of terminal pro-brain natriuretic peptide (NT-proBNP) as an example. The base sequences encoding the two antigenic epitopes of NT-proBNP were recombined into the FG loop region and the C-terminus of FN3, fused by 4 GS or polyN linker. The fusion proteins (named FN3-epitopes-4GS and FN3-epitopes-polyN, respectively) were expressed and purified cost-effectively using an Escherichia coli expression system. The immunoreactivity of recombinant substitutes was preliminarily confirmed by western blot analysis using epitope-specific antibodies. The sandwich enzyme-linked immunosorbent assay demonstrated that either FN3-epitopes-polyN or FN3-epitopes-4GS was highly sensitive, and FN3-epitopes-polyN exhibited better kinetics to specific antibodies than FN3-epitopes-4GS, showing a linear dose-response relationship in the concentration range of 0.06-12.85 ng/ml, which suggest that the polyN linker was more suitable for constructing the FN3-based substitute antigens compared to the 4 GS linker. Furthermore, the serum stability test and differential scanning calorimetry analysis showed that the recombinant FN3-epitopes-polyN maintained the original stability of FN3. Therefore, it was confirmed that FN3 could be engineered to construct a stable biomacromolecular substitute for displaying double epitopes of antigen proteins, such as NT-proBNP. In summary, a cost-effective strategy to produce NT-proBNP substitute antigens with good immunoreactivity and physicochemical stability was established in this work, which may provide potential uses for the production of other substitute antigens in the future.

Molecular Identification of Anion Exchange Protein 3 in Pacific White Shrimp (Litopenaeus vannamei): mRNA Profiles for Tissues, Ontogeny, Molting, and Ovarian Development and Its Potential Role in Stress-Induced Gill Damage.

Bicarbonate (HCO3 -) transport mechanisms play an essential role in the acid-base homeostasis of aquatic animals, and anion exchange protein 3 (AE3) is a membrane transport protein that exchanges Cl-/HCO3 - across the cell membrane to regulate the intracellular pH. In this study, the full-length cDNA of AE3 (Lv-AE3) was obtained from the Pacific white shrimp (Litopenaeus vannamei). The Lv-AE3 cDNA is 4,943 bp in length, contains an open reading frame of 2,850 bp, coding for a protein of 949 amino acids with 12 transmembrane domains. Lv-AE3 shows high sequence homology with other AE3 at the protein level. Lv-AE3 mRNA was ubiquitously detected in all tissues selected, with the highest expression level in the gill, followed by the ovary, eyestalk and brain. By in situ hybridization, Lv-AE3-positive cells were shown predominant localization in the secondary gill filaments. The expression levels of Lv-AE3 were further investigated during the essential life processes of shrimp, including ontogeny, molting, and ovarian development. In this case, the spatiotemporal expression profiles of Lv-AE3 in L. vannamei were highly correlated with the activities of water and ion absorption; for example, increased mRNA levels were present after hatching, during embryonic development, after ecdysis during the molt cycle, and in the stage IV ovary during gonadal development. After low/high pH and low/high salinity challenges, the transcript levels of Lv-AE3 were reduced in the gill, while the cell apoptosis rate increased. In addition, knockdown of Lv-AE3 mRNA expression induced cell apoptosis in the gill, indicating a potential link between Lv-AE3 and gill damage. Altogether, this study thoroughly investigated the relationship between the mRNA expression profiles of Lv-AE3 and multiple developmental and physiological processes in L. vannamei, and it may benefit the protection of crustaceans from fluctuated aquatic environments.

Development of specific and selective bactericide by introducing exogenous metabolite of pathogenic bacteria.

The widespread and repeated use of broad-spectrum bactericides has led to an increase in resistance. Developing novel broad-spectrum bactericides cannot solve the resistance problem, and may even aggravate it. The design of specific and selective bactericides has become urgent. A specific bactericidal design strategy was proposed by introducing exogenous metabolites in this study. This strategy was used to optimize two known antibacterial agents, luteolin (M) and Isoprothiolane (D), against Xoo. Based on the prodrug principles, target compound MB and DB were synthesized by combing M or D with exogenous metabolites, respectively. Bactericidal activity test results demonstrated that while the antibacterial ability of target compounds was significantly improved, their selectivity was also well enhanced by the introducing of exogenous metabolites. Comparing with the original compound, the antibacterial activity of target compound was significantly increased 92.0% and 74.5%, respectively. The optimized target compounds were more easily absorbed, and the drug application concentrations were much lower than those of the original agents, which would greatly reduce environmental pollution and relieve resistance risk. Our proposed strategy is of great significance for exploring the specific and selective bactericides against other pathogens.

Formation of Multispecies Biofilms and Their Resistance to Disinfectants in Food Processing Environments: A Review.

ABSTRACT: In food processing environments, various microorganisms can adhere and aggregate on the surface of equipment, resulting in the formation of multispecies biofilms. Complex interactions among microorganisms may affect the formation of multispecies biofilms and resistance to disinfectants, which are food safety and quality concerns. This article reviews the various interactions among microorganisms in multispecies biofilms, including competitive, cooperative, and neutral interactions. Then, the preliminary mechanisms underlying the formation of multispecies biofilms are discussed in relation to factors, such as quorum-sensing signal molecules, extracellular polymeric substances, and biofilm-regulated genes. Finally, the resistance mechanisms of common contaminating microorganisms to disinfectants in food processing environments are also summarized. This review is expected to facilitate a better understanding of interspecies interactions and provide some implications for the control of multispecies biofilms in food processing.

The tRNA-derived fragment 5026a inhibits the proliferation of gastric cancer cells by regulating the PTEN/PI3K/AKT signaling pathway.

BACKGROUND: Recently, tRNA-derived fragments (tRFs) have been shown to serve important biological functions. However, the role of tRFs in gastric cancer has not been fully elucidated. This study aimed to identify the tumor suppressor role of tRF-5026a (tRF-18-79MP9P04) in gastric cancer. METHODS: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was first used to detect tRF-5026a expression levels in gastric cancer tissues and patient plasma. Next, the relationship between tRF-5026a levels and clinicopathological features in gastric cancer patients was assessed. Cell lines with varying tRF-5026a levels were assessed by measuring tRF-5026a using qRT-PCR. After transfecting cell lines with a tRF-5026a mimic or inhibitor, cell proliferation, colony formation, migration, apoptosis, and cell cycle were evaluated. The expression levels of related proteins in the PTEN/PI3K/AKT pathway were also analyzed by Western blotting. Finally, the effect of tRF-5026a on tumor growth was tested using subcutaneous tumor models in nude mice. RESULTS: tRF-5026a was downregulated in gastric cancer patient tissues and plasma samples. tRF-5026a levels were closely related to tumor size, had a certain diagnostic value, and could be used to predict overall survival. tRF-5026a was also downregulated in gastric cancer cell lines. tRF-5026a inhibited the proliferation, migration, and cell cycle progression of gastric cancer cells by regulating the PTEN/PI3K/AKT signaling pathway. Animal experiments showed that upregulation of tRF-5026a effectively inhibited tumor growth. CONCLUSIONS: tRF-5026a (tRF-18-79MP9P04) is a promising biomarker for gastric cancer diagnostics and has tumor suppressor effects mediated through the PTEN/PI3K/AKT signaling pathway.

Neurological deterioration as a result of improper neck position detected by intraoperative neurophysiological monitoring in a cervical stenosis patient: A case report.

RATIONALE: Intraoperative neurophysiological monitoring (IONM) is widely used in spinal surgeries to prevent iatrogenic spinal cord injury (SCI). Most surgeons focus on avoiding neurological compromise intraoperatively, while ignoring the possibility of nerve damage preoperatively, such as neck positioning. Thus, this study aims to report a case with transient neurological deterioration due to improper neck position detected by IONM during cervical surgery. PATIENT CONCERNS: A 63-year-old male patient had been suffering from hypoesthesia of the upper and lower extremities for three years. DIAGNOSES: Severe cervical stenosis (C5-C7) and cervical ossification of a posterior longitudinal ligament. INTERVENTIONS: The cervical stenosis patient underwent an anterior cervical corpectomy decompression and fusion (ACDF) surgery with the assistance of IONM. When the lesion segment was exposed, the SSEP and MEP suddenly elicited difficulty indicating that the patient may have developed SCI. All the technical causes of IONM events were eliminated, and the surgeon suspended operation immediately and suspected that the IONM alerts were caused by cervical SCI due to the improper position of the neck. Subsequently, the surgeon repositioned the neck of the patient by using a thinner shoulders pad. OUTCOMES: At the end of the operation, the MEP and SSEP signals gradually returned to 75% and 80% of the baseline, respectively. Postoperatively, the muscle strength of bilateral biceps decreased from grade IV to grade III. Besides, the sensory disturbance of both upper extremities aggravated. However, the muscle power and hypoesthesia were significantly improved after three months of neurotrophic therapy and rehabilitation training, and no complications of nerve injury were found at the last follow-up visit. LESSONS: IONM, consisting of SSEP and MEP, should be applied throughout ACDF surgery from the neck positioning to suture incisions. Besides, in the ward 1to 2 days before operation, it is necessary for conscious patients with severe cervical stenosis to simulate the intraoperative neck position. If the conscious patients present signs of nerve damage, they can adjust the neck position immediately until the neurological symptoms relieve. Therefore, intraoperatively, the unconscious patient can be placed in a neck position that was confirmed preoperatively to prevent SCI.

Global profile of tRNA-derived small RNAs in gastric cancer patient plasma and identification of tRF-33-P4R8YP9LON4VDP as a new tumor suppressor.

Transfer RNA (tRNA)-derived small RNAs (tsRNAs) have been found to play important roles in the occurrence and development of cancers. However, the tsRNA profile in gastric cancer is unknown. In this study, we aimed to identify the global tsRNA profile in plasma from gastric cancer patients and elucidate the role of tRF-33-P4R8YP9LON4VDP in gastric cancer. Differentially expressed tsRNAs in the plasma of gastric cancer patients and healthy controls were investigated using RNA sequencing. The expression levels of tRF-33-P4R8YP9LON4VDP in the plasma of gastric cancer patients, healthy controls and gastric cancer cell lines were first detected by quantitative reverse transcription-polymerase chain reaction. The effects of tRF-33-P4R8YP9LON4VDP overexpression or downregulation in gastric cancer cells on proliferation, migration, apoptosis, and cell cycle were analyzed using the Cell Counting Kit-8, scratch assay, Transwell assay, and flow cytometry, respectively. There were 21 upregulated and 46 downregulated tsRNAs found in plasma from gastric cancer patients. The significantly upregulated tsRNAs included tRF-18-S3M83004, tRF-31-PNR8YP9LON4VD, tRF-19-3L7L73JD, tRF-33-P4R8YP9LON4VDP, tRF-31-PER8YP9LON4VD, tRF-18-MBQ4NKDJ, and tRF-31-PIR8YP9LON4VD. The significantly downregulated tsRNAs included tRF-41-YDLBRY73W0K5KKOVD, tRF-18-07QSNHD2, tRF-28-86J8WPMN1E0J, tRF-29-86V8WPMN1EJ3, tRF-31-6978WPRLXN4VE, tRF-30-MIF91SS2P46I, tRF-26-MI7O3B1NR8E, tRF-30-RRJ89O9NF5W8, tRF-26-XIP2801MK8E, and tRF-35-V0J8O9YEKPRS93, In vitro studies showed that tRF-33-P4R8YP9LON4VDP inhibited proliferation of gastric cancer cells. In conclusion, tsRNAs such as tRF-33-P4R8YP9LON4VDP could serve as a novel diagnostic biomarker and target for gastric cancer therapeutics.

Whole genome sequencing and genome annotation of Dermacoccus abyssi strain HZAU 226 isolated from spoiled eggs.

Dermacoccus abyssi strain HZAU 226 is a spoilage bacterium isolated from eggs. So far, there are still few genomic resources available on the Dermacoccus abyssi. Here, we reported the complete genome sequence of Dermacoccus abyssi strain HZAU 226. High-quality DNA was extracted using the Qiagen kit, then single-molecule sequencing was performed by GridION sequencer. The raw data was quality-controlled and assembled to obtain the final genome, which consisted of a complete genome of 2,992,060 bp circular chromosome and a 64,524 bp plasmid. The structural and functional annotations of the genome were achieved through the analysis of different available databases, including antibiotic resistance genes, secondary metabolite synthesis genes and stress-related genes. Meanwhile, comparative genomic analyses of the strains were also performed. This is the first report on the complete genome of Dermacoccus abyssi, which will provide genomic resources for the study of spoilage bacteria in eggs.

Low expression of hsa_circ_0001811 in gastric cancer and its role in clinical diagnosis.

BACKGROUND: As far as gastric cancer is concerned, there is a lack of specific early diagnostic biomarkers in clinical diagnosis. Circular RNAs (circRNAs) have been found to be stable in gastric cancer tissues and plasma, so they have the potential to become new type of diagnostic biomarkers for gastric cancer. MATERIALS AND METHODS: The hsa_circ_0001811 expressions in gastric cancer tissues and paired non-cancer tissues, preoperative and postoperative plasma of patients with gastric cancer, and plasma of healthy volunteers were detected using quantitative reverse transcription-polymerase chain reaction. The receiver operating characteristic (ROC) curves were drawn; and the combined ROC curves were used to analyze their diagnostic values. The correlation between the plasma or tissue hsa_circ_0001811 levels and the clinicopathological factors of gastric cancer was further analyzed. RESULTS: Hsa_circ_0001811 was found to be lowly expressed in gastric cancer tissues and plasma from patients with gastric cancer. The area under the ROC curve (AUC) in the gastric cancer tissues and the plasma of patients with gastric cancer was 0.658 and 0.747, respectively. The AUC increased to 0.824 after combination of both. The expression level of hsa_circ_0001811 in gastric cancer tissue correlated with carcinoembryonic antigen (CEA) (P = .0347), tissue differentiation (P = .0138), and lymph node metastasis (P = .0234), while plasma hsa_circ_0001811 level was related to carbohydrate antigen (CA19-9) (P = .0278), lymph node metastasis (P = .0469), distant metastasis (P = .0384), and age (P = .0085). CONCLUSIONS: The above results indicate that hsa_circ_0001811 may become a new biomarker for clinical diagnosis of gastric cancer.

Interleukin-2 enhancer binding factor 2 (ILF2) in pacific white shrimp (Litopenaeus vannamei): Alternatively spliced isoforms with different responses in the immune defenses against vibrio infection.

Alternative splicing is an essential molecular mechanism that increase the protein diversity of a species to regulate important biological processes. As a transcription factor, Interleukin-2 enhancer binding factor 2 (ILF2) regulates the functions of interleukin-2 (IL-2) at the levels of transcription, splicing and translation, and plays other critical roles in the immune system. ILF2 is well-documented in vertebrates, while little is currently known in crustacean species such as the Pacific white shrimp (Litopenaeus vannamei). In the present study, five cDNA for spliced isoforms of Lv-ILF2 were identified, in which four of them are the full-length long isoforms (Lv-ILF2-L1, Lv-ILF2-L2, Lv-ILF2-L3 and Lv-ILF2-L4) and one of them is a truncated short isoform (Lv-ILF2-S). The whole sequence of ILF2 gene from L. vannamei was obtained, which is 11,680 bp in length with 9 exons separated by 8 introns. All five isoforms contain a domain associated with zinc fingers (DZF). Two alternative splicing types (alternative 5' splice site and alternative 3' splice site) were identified in the five isoforms. The Lv-ILF2 mRNA showed a broad distribution in all detected tissues, and the Lv-ILF2-L transcript levels were higher than those of Lv-ILF2-S in corresponding tissues. The mRNA levels of Lv-ILF2-S in the hepatopancreas, heart, muscle and stomach, but not in the eyestalk, were significantly increased after challenges with Vibrio harveyi or lipopolysaccharide (LPS), while no significant changes were observed for the transcript levels of Lv-ILF2-L in these tissues under the same immune stimulants. On the contrary, the transcript levels of neither Lv-ILF2-S nor Lv-ILF2-L were affected by challenges of polyinosinic: polycytidylic acid [Poly (I:C)]. In addition, after knockdown of the Lv-ILF2 mRNA level by siRNA, the mortality of shrimp and the hepatopancreatic bacterial numbers were significantly increased under V. harveyi challenge, indicating that Lv-ILF2 might participate in the immune defenses against V. harveyi invasion. Collectively, our study here supplied the first evidence for a novel splicing mechanism of ILF2 transcripts, and provided a functional link between the Lv-ILF2 isoforms and the capacity against pathogenic Vibrio in penaeid shrimp.

Diastereoselective synthesis and conformational analysis of 4,5-difluoropipecolic acids.

Stereoselectively-fluorinated analogs of pipecolic acid have been investigated through a combined theoretical and experimental approach. Three of the four possible diastereoisomers of 4,5-difluoropipecolic acid were successfully synthesized via deoxyfluorination chemistry, navigating a complex reaction network that included neighboring group participation, rearrangement, and elimination pathways. A DFT-based conformational study, supported by NMR J-based analysis, revealed that the different diastereoisomers of 4,5-difluoropipecolic acid preferentially adopt different puckers of the six-membered ring. These findings could have future relevance for the conformational control of biologically active peptides.

First echinoderm alpha-amylase from a tropical sea cucumber (Holothuria leucospilota): Molecular cloning, tissue distribution, cellular localization and functional production in a heterogenous E.coli system with codon optimization.

Holothuria leucospilota (Echinodermata: Holothuroidea) is a widespread tropical sea cucumber with strong value for the ecological restoration of coral reefs. Therefore, some studies regarding the artificial breeding and cultivation of H. leucospilota have been undertaken recently. However, the biological functions of the digestive system of this species have not been elucidated. In this study, a cDNA coding for α-amylase, an indicator of digestive maturity in animals, was identified from H. leucospilota and designated Hl-Amy. The full-length cDNA of the Hl-Amy gene, which is 1734 bp in length with an open reading frame (ORF) of 1578 bp, encodes a 525 amino acid (a.a.) protein with a deduced molecular weight of 59.34 kDa. According to the CaZy database annotation, Hl-Amy belongs to the class of GH-H with the official nomenclature of α-amylase (EC 3.2.1.1) or 4-α-D-glucan glucanohydrolase. The Hl-Amy protein contains a signal peptide at the N-terminal followed by a functional amylase domain, which includes the catalytic activity site. The mRNA expression of Hl-Amy was abundantly exhibited in the intestine, followed by the transverse vessel with a low level, but was hardly detected in other selected tissues. During embryonic and larval development, Hl-Amy was constitutively expressed in all stages, and the highest expression level was observed in the blastula. By in situ hybridization (ISH), positive Hl-Amy signals were observed in different parts of the three different intestinal segments (foregut, midgut and hindgut). The Hl-Amy recombinant protein was generated in an E. coli system with codon optimization, which is necessary for Hl-Amy successfully expressed in this heterogenous system. The Hl-Amy recombinant protein was purified by immobilized metal ion affinity chromatography (IMAC), and its activity of starch hydrolysis was further detected. The optimal temperatures and pH for Hl-Amy recombinant protein were 55°C and 6.0, respectively, with an activity of 62.2 U/mg. In summary, this current study has filled a knowledge gap on the biological function and expression profiles of an essential digestive enzyme in sea cucumber, which may encourage future investigation toward rationalized diets for H. leucospilota in artificial cultivation, and optimized heterogenous prokaryotic systems for producing recombinant enzymes of marine origins.