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Bacillus licheniformis MW3 degrades bird feathers. Feather keratin is rich in cysteine, which is metabolized to produce hazardous sulfide and sulfane sulfur. A challenge to B. licheniformis MW3 growing on feathers is to detoxify them. Here we identified a gene cluster in B. licheniformis MW3 to deal with these toxicity. The cluster contains 11 genes: the first gene yrkD encodes a repressor, the 8th and 9th genes nreB and nreC encode a two-component regulatory system, and the 10th and 11th genes encode sulfide: quinone reductase (SQR) and persulfide oxygenase (PDO). SQR and PDO collectively oxidize sulfide and sulfane sulfur to sulfite. YrkD sensed sulfane sulfur to derepress the 11 genes. The NreBC system sensed sulfide and further amplified the transcription of sqr and pdo. The two regulatory systems synergistically controlled the expression of the gene cluster, which was required for the bacterium to grow on feather. The findings highlight the necessity of removing sulfide and sulfane sulfur during feather degradation and may help with bioremediation of feather waste and sulfide pollution. The feather-degrading bacterium Bacillus licheniformis MW3 contains a sulfur-metabolizing gene cluster, in which one sulfide-sensor and one sulfane sulfur-sensor synergistically control gene expression, enabling the bacterium to grow on bird feathers. [ABSTRACT FROM AUTHOR]

Although intravenous immunoglobulin (IVIG) can effectively treat Kawasaki disease (KD), 10–20% of KD patients show no beneficial clinical response. Developing reliable criteria to discriminate non-responders is important for early planning of appropriate regimens. To predict the non-responders before IVIG treatment, gene expression dataset of 110 responders and 61 non-responders was obtained from Gene Expression Omnibus. After weighted gene co-expression network analysis, we found that modules positively correlated with the non-responders were mainly associated with myeloid cell activation. Transcripts up-regulated in the non-responders, IL1R2, GK, HK3, C5orf32, CXCL16, NAMPT and EMILIN2, were proven to play key roles via interaction with other transcripts in co-expression network. The crucial transcripts may affect the clinical response to IVIG treatment in acute KD. And these transcripts may serve as biomarkers and therapeutic targets for precise diagnosis and treatment of the non-responders. [ABSTRACT FROM AUTHOR]

Soybean is an economically important leguminous crop, and pod dehiscence of soybean could cause huge yield loss. In this study, we measured fruit-cracking forces and percentages of dehisced pods for ten soybean accessions, then separated them into two groups as shattering-sensitive (SS) and shattering-resistant (SR) soybeans. Pod transcriptomes from these two groups were analyzed, and 225 differentially expressed genes (DEGs) were identified between SS and SR soybeans. Some of these DEGs have been previously reported to be associated with pod dehiscence in soybean. The expression patterns of selected DEGs were validated by real-time quantitative reverse transcription PCR, which confirmed the expression changes found in RNA-seq analysis. We also de novo identified 246 soybean pod-long intergenic ncRNAs (lincRNAs), 401 intronic lncRNAs, and 23 antisense lncRNAs from these transcriptomes. Furthermore, genes and lincRNAs co-expression network analysis showed that there are distinct expression patterns between SS and SR soybeans in some co-expression modules. In conclusion, we systematically investigated potential genes and molecular pathways as candidates for differences in soybean pod dehiscence and will provide a useful resource for molecular breeding of soybeans. [ABSTRACT FROM AUTHOR]

Purpose: Colon cancer is the most common malignant tumor in the intestine. Abnormal Carboxylesterases 3 (CES3) expression had been reported to be correlated to multiple tumor progression. However, the association among CES3 expression and prognostic value and immune effects in colonic adenocarcinoma (COAD) were unclear. Patients and methods: The transcription and expression data of CES3 and corresponding clinical information was downloaded from The Cancer Genome Atlas (TCGA). The CES3 protein expression and the prognostic value were verified based on tissue microarray data. The Cancer immune group Atlas (TCIA), Tumor Immune Dysfunction and Exclusion (TIDE) algorithm and the GSE78220 immunotherapy cohort were used to forecast immunotherapy efficacy. Finally, a prognostic immune signature was constructed and verified. Results: Compared with normal colon tissues, the expression of mRNA and protein levels of CES3 were downregulated in tumor tissues. CES3 expression was associated with TIICs. Hihg-CES3 COAD patients had better efficacy of concurrent immunotherapy. CES3-related immune genes (CRIs) were identified and were then used to construct prognostic immune signature and had been successfully verified in GES39582. Conclusion: CES3 might be a potential immune-related gene and promising prognostic biomarker in COAD. [ABSTRACT FROM AUTHOR]

Background: Abnormality of PTEN gene and Wnt/β-catenin signaling have been strongly implicated in various malignant cancers. Recently, it has been noted that a functional interaction/cross-talk was found between the PTEN/PI3K/AKT and Wnt/β-catenin, which plays a key role in the development of cancers. However, few related studies on gastric cancer are available.Aim: We examined the expression of PTEN and β-catenin in gastric cancer tissues and detected whether down-regulation of PTEN promotes the migration and invasion in gastric cancer cells along with its underlying mechanism.Materials and Methods: Immunocytochemistry, a wound healing assay, a Matrigel invasion assay, an immunofluorescence staining were performed to detect expression of PTEN and β-catenin in gastric cancer and adjacent normal tissues, cell migration, cell invasion, and the effects of PTEN knockdown on β-catenin in cells, respectively. Further, MMP-2 and MMP-9 activities were analyzed by zymography assay. The changes in related proteins were further quantified by western blotting.Results: Low expression of PTEN was found in majority of gastric cancer tissues, which showed significant associations with differentiation grade in gastric cancer patients. Further, a negative correlation was revealed between PTEN and β-catenin protein expression in gastric cancer tissues (r = - 0.546, P < 0.01). Additionally, PTEN knockdown promoted the migration and invasion of cells and caused an obvious increase in p-AKT, p-GSK-3β, β-catenin, E-cadherin, MMP-7, MMP-2, and MMP-9 in gastric cancer cells.Conclusion: Our results indicated PTEN gene might induce cell invasion and migration via regulating AKT/GSK-3β/β-catenin signaling pathway, playing a vital role in the progression of gastric cancer. [ABSTRACT FROM AUTHOR]

Background: It was reported that the nuclear receptor coactivator amplified in breast cancer1 (AIB1) could regulate cancer cell invasion and migration in a nuclear receptor signaling-independent manner. Meanwhile, the process of epithelial mesenchymal transition (EMT) is critical for tumor invasion and metastasis. The present study aimed to determine the role of AIB1 and EMT markers in human pancreatic adenocarcinoma. Methods: AIB1, ZO-1, E-cadherin, vimentin, and N-cadherin protein expression in 76 pancreatic adenocarcinomas were assessed using immunohistochemistry and analyzed for clinicopathological significance. Results: The frequency of AIB1 overexpression in pancreatic adenocarcinomas with lymph node metastasis is 68 % (19/28), which is significantly higher than in pancreatic adenocarcinomas without lymph node metastasis (42 %; 20/48). In addition, the frequency of low expression of E-cadherin in pancreatic carcinomas with lymph node metastasis (68 %; 19/28) was significantly higher than in tumors without lymph node metastasis (44 %; 21/48). Correlation analysis demonstrated that the overexpression of AIB1 was inversely correlated with low expression of E-cadherin in pancreatic adenocarcinomas. Conclusion: Overexpression of AIB1 might promote invasion and metastasis of cancer cells and is associated with down-regulation of E-cadherin in pancreatic adenocarcinomas. [ABSTRACT FROM AUTHOR]

Purpose: Kashin-Beck disease (KBD) is an endemic degenerative osteoarthritis associated with extracellular matrix degradation. The aim of this investigation was to evaluate the role of targeting genes in the pathogenesis of KBD and primary osteoarthritis (OA) involved in extracellular matrix degradation. Methods: Agilent 44 K human whole-genome oligonucleotide microarrays were used to detect the gene expression in KBD and OA cartilage. The mRNA and protein expressions of CSGalNAcT-1 and Hapln-1 in chondrocytes were verified by reverse transcription polymerase chain reaction (RT-PCR) and western blot, and their expression in cartilage were verified with immunocytochemical analysis. Meanwhile, CSGalNAcT-1 and Hapln-1 protein levels in the selenium intervention group of KBD with different concentrations (0.25, 0.1and 0.05 μg/ml) were detected by western blot. Results: CSGalNAcT-1 and Hapln-1 were down-regulated in KBD and OA at both mRNA and protein levels, and were increased in Se(Selenium) groups compared to KBD free-Se group. However, Wnt 3a, β-catenin and Runx-2 were up-regulated in OA and KBD at protein levels. Additionally, immunohistochemical staining showed that CSGalNAcT-1 and Hapln-1 were reduced in all zones of KBD and OA articular cartilage, but not significantly reduced in the up zone of OA articular cartilage. Conclusions: The CSGalNAcT-1 and Hapln-1 were down-regulated in both KBD and OA cartilage. CSGalNAcT-1 may be involved in the damage of articular cartilage of KBD and OA by regulating Hapln-1 in the Wnt/β-catenin signalling pathway. It was indicated that CSGalNAcT-1 and Hapln-1 may play important roles in the pathogenesis of KBD and OA. [ABSTRACT FROM AUTHOR]

Imperatorin is a type of coumarin compound with antibacterial and antiviral activities. In the present study, we examined the anti-inflammatory effects of imperatorin in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages by investigating its impact on the production and expression of cytokines and the major signal-transduction pathways. We found that imperatorin downregulated LPS-induced levels of TNF-α, IL-1β, and IL-6 in RAW 264.7 macrophages in a concentration-dependent manner, and it significantly inhibited expression of TNF-α and IL-6 ( P < 0.05 or P < 0.01). The phosphorylation of mitogen-activated protein kinases and nuclear factor-kappaB (NF-κB) p65 protein were analyzed by western blotting. In RAW 264.7 macrophages treated with 1 mg/L of LPS, imperatorin significantly inhibited p38 and Jun N-terminal kinase phosphorylation protein expression. However, there was no significant change in p-ERK. Furthermore, imperatorin also inhibited NF-κB translocation into the nucleus through blockage of IκBα phosphorylation and degradation. [ABSTRACT FROM AUTHOR]

The TBC1 domain family member 10B (EPI64B/TBC1D10B), a member of the RabGAP EPI64 subfamily, contains a TBC domain that confers GTPase-activating protein activity. Even though overexpression of TBC1D10B has been reported to promote tumor invasion and metastasis in gastric adenocarcinoma, the prognostic value of TBC1D10B and its correlation with DNA methylation and immune infiltration in hepatocellular carcinoma are still not known. Transcriptional expression profiles of TBC1D10B between hepatocellular carcinoma tissues and normal tissues were downloaded from The Cancer Genome Atlas and Gene Expression Omnibus. The Clinical Proteomic Tumor Analysis Consortium and the Human Protein Atlas were used to assess the TBC1D10B protein expression. The biological functions of TBC1D10B were evaluated by the Metascape database and by Gene Set Enrichment Analysis (GSEA). Receiver operating characteristic (ROC) curve analysis was used to distinguish hepatocellular carcinoma from adjacent normal tissues. The effect of TBC1D10B on survival was estimated using the Kaplan–Meier method. DNA methylation in the TBC1D10B gene was assessed using the online MEXPRESS and MethSurv tools. The association between TBC1D10B mRNA expression and immune cell infiltration was investigated by the TIMER2 web server, tumor immune estimation resource and single-sample GSEA. This study found that TBC1D10B is highly expressed in hepatocellular carcinoma and that increased TBC1D10B mRNA expression is associated with female sex, lower Body Mass Index, high level of alpha fetal protein, and worse clinical stages. The mRNA and protein levels of TBC1D10B were verified in cells. Functional annotation indicated enrichment with negative regulation of the cell cycle, extracellular matrix, and corresponding pathways in the high-TBC1D10B phenotype. The ROC curve analysis showed that, with a cutoff level of 2.912, the accuracy, sensitive, and specificity in differentiate TBC1D10B hepatocellular carcinoma from adjacent controls were 0.931, 0.920, and 0.802, respectively. Kaplan–Meier survival analysis showed that hepatocellular carcinoma patients with high TBC1D10B had a worse prognosis than those with low TBC1D10B, especially in patients with a weight below 70 kg, height above 170 cm, and histological G2 and G3. We also found that the methylation of TBC1D10B was associated with the prognosis in patients with hepatocellular carcinoma. Moreover, correlation analysis indicated that TBC1D10B mRNA expression was positively correlated with infiltration levels of most immune cells, but negatively correlated with Th17 and cytotoxic cells infiltration. Our study indicates that increased TBC1D10B expression in hepatocellular carcinoma may play a role in tumorigenesis by regulating the cell cycle and extracellular matrix. TBC1D10B may be a novel prognostic and predictive marker and immune therapeutic target in hepatocellular carcinoma patients. [ABSTRACT FROM AUTHOR]

The appropriate reference gene is a prerequisite for accurate normalization of gene expression level, and research on suitable reference genes in clam Cyclina sinensis is scarce. To improve the situation, we selected five commonly used housekeeping genes, including β-actin, Elongation factor 1-α (EF1-α), Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), 40S ribosomal protein S18 (RPS18), and Tubulin α (TUB-α), then evaluated their expression stability in different adult tissues and under different experimental treatments (salinity stress and Vibrio parahaemolyticus infection). Their expression stability was analyzed by three frequently used programs, geNorm, NormFinder, and BestKeeper. This analysis indicated that multiple genes should be used for normalization, and we concluded that the reference gene combination GAPDH-RPS18-β-actin, should be used for qRT-PCR analysis in different tissues of C. sinensis under normal physiological conditions. For the clams under salinity stress and Vibrio infection, EF1-α-GAPDH-RPS18 was recommended as the gene combination for qRT-PCR normalization. TUB-α was generally poorly ranked by all programs, and should not be used in future studies. This study should provide fundamental support for accurate quantitative gene expression analysis of this species. [ABSTRACT FROM AUTHOR]

In this study, the cellulase gene celD from Clostridium thermocellum was cloned into expression vectors pET-20b(+) and pHsh. While high expression can be achieved by means of both these expression systems, only the pHsh expression system gives soluble proteins. By weakening the mRNA secondary structure and replacing the rare codons for the N-terminal amino acids of the target protein, the expression level of CelD was increased from 4.1 ± 0.3 to 6.4 ± 0.4 U ml in LB medium. Recombinant CelD was purified by heat treatment followed by Ni-NTA affinity. The purified CelD exhibited the highest activity at pH 5.4 and 60°C, and retained more than 50% activity after incubation at 70°C for 1 h. The cellulase activity of CelD was significantly enhanced by Ca but inhibited by EDTA. The favorable properties of CelD offer the potential for genetic modification of strains for biomass degradation. Presently, one of the major bottlenecks for industrial cellulase users is the high cost of enzyme production. The high level expression of soluble enzymes from the pHsh expression system offers a novel approach for the production of cellulases to be used in various agro-industrial processes such as chemical, food and textile. [ABSTRACT FROM AUTHOR]

As a tightly controlled cell death process, apoptosis eliminates unwanted cells and plays a vital role in multicellular organisms. Previous study have demonstrated that apoptosis occurred in Spodoptera frugiperda cultured Sf9 cells, which triggered by diverse apoptotic stimuli, including azadirachtin, camptothecin and ultraviolet. Due to its simplicity, high sensitivity and reliable specificity, RT-qPCR has been used widespread for analyzing expression levels of target genes. However, the selection of reference genes influences the accuracy of results profoundly. In this study, eight genes were selected for analyses of their suitability as references for normalizing RT-PCR data in Sf9 cells treated with apoptotic agents. Five algorithms, including NormFinder, BestKeeper, Delta Ct method, geNorm, and RefFinder, were used for stability ranking. Based on comprehensively analysis, the expression stability of selected genes varied in cells with different apoptotic stimuli. The best choices for cells under different apoptosis conditions were listed: EF2 and EF1α for cells treated with azadirachtin; RPL13 and RPL3 for cells treated with camptothecin; EF1α and β-1-TUB for cells irradiated under ultraviolet; and EF1α and EF2 for combinational analyses of samples. Our results not only facilitate a more accurate normalization for RT-qPCR data, but also provide the reliable assurance for further studies of apoptotic mechanisms under different stimulus in Sf9 cells. [ABSTRACT FROM AUTHOR]

Sulfur (S) is an essential macronutrient required by plants. Plants absorb and transport S through sulfate transporters (SULTRs). In this study, we cloned 8 SULTR genes (CsSULTR1;1/1;2/2;1/3;1/3;2/3;3/3;5/4;1) from tea plant (Camellia sinensis), all of which contain a typical sulfate transporter and antisigma factor antagonist (STAS) conserved domain. Phylogenetic tree analysis further divided the CsSULTRs into four main groups. Many cis-acting elements related to hormones and environmental stresses were found within the promoter sequence of CsSULTRs. Subcellular localization results showed that CsSULTR4;1 localized in the vacuolar membrane and that other CsSULTRs localized to the cellular membrane. The tissue-specific expression of the 8 CsSULTR genes showed different expression patterns during the active growing period and dormancy period. In particular, the expression of CsSULTR1;1 was highest in the roots, but that of CsSULTR1;2 was lowest in the dormancy period. The expression of CsSULTR1;1/1;2/2;1/3;2 was stimulated under different concentrations of selenium (Se) and S; moreover, CsSULTR1;2/2;1/3;3/3;5 was upregulated in response to different valences of Se. [ABSTRACT FROM AUTHOR]

MYB transcription factors play a key role in anthocyanin biosynthesis. However, there are few reports on MYB transcription factors that can regulate anthocyanin biosynthesis in potato. In this study, the StMYB113 gene was isolated from potato and characterized. Transient activation experiments showed that the StMYB113 gene could promote anthocyanin biosynthesis in tobacco leaves by enhancing the expression of genes encoding anthocyanin biosynthesis-related enzymes. In addition, overexpression of the StMYB113 gene promoted anthocyanin accumulation in potato tuber skin. Furthermore, transient luciferase analysis showed that StMYB113 enhanced the promoter activities of StF3H and StDFR genes significantly. These results indicated that the StMYB113 gene could promote anthocyanin accumulation in potato tuber skin by directly activating the expression of StF3H and StDFR. The promoter activity of StMYB113 was significantly higher in light than in dark conditions, suggesting that the expression of StMYB113 is induced by light. These findings illustrate the role and regulation of the StMYB113 gene in potato anthocyanin biosynthesis and enrich the regulatory theory of potato anthocyanin biosynthesis. [ABSTRACT FROM AUTHOR]

PBK (PDZ-binding kinase) is a protein-coding gene that encodes a serine/threonine protein kinase associated with the dual-specific mitogen-activated protein kinase (MAPKK) family. Overexpression of this gene is closely linked to tumor development. In this study, we aimed to investigate the role of PBK in lung adenocarcinoma (LUAD) progression, prognosis, and immune evasion. We conducted a pan-cancer analysis of PBK to examine its expression and prognostic value. In the LUAD cohort, we analyzed PBK expression, prognosis, mutational features, and immune infiltration in groups with different PBK expression levels. We constructed a PBK-associated genomic model, integrated it into a nomogram, and compared high and low-risk subgroups. In our pan-cancer analysis, PBK was significantly upregulated, particularly in LUAD patients, and displayed poor prognosis. The high PBK expression group had many deletion mutations but still showed gene upregulation. Immune infiltration analysis indicated that PBK-triggered immune escape in the high expression group might relate to antigen presentation, dendritic cell, and CD8+ T cell infiltration. We constructed a 5-gene prognostic model and a nomogram to quantify individual survival probabilities. The PBK-associated gene prognostic model reliably predicted patient prognosis and drug response. Our findings offer new insights into PBK-induced immune escape and targeted therapy during LUAD development, providing valuable suggestions for clinical treatment approaches. [ABSTRACT FROM AUTHOR]

Differential gene expression (DGE) analysis has been widely employed to identify genes expressed differentially with respect to a trait of interest using RNA sequencing (RNA-Seq) data. Recent RNA-Seq data with large samples pose challenges to existing DGE methods, which were mainly developed for dichotomous traits and small sample sizes. Especially, existing DGE methods are likely to result in inflated false positive rates. To address this gap, we employed a linear mixed model (LMM) that has been widely used in genetic association studies for DGE analysis of quantitative traits. We first applied the LMM method to the discovery RNA-Seq data of dorsolateral prefrontal cortex (DLPFC) tissue (n = 632) with four continuous measures of Alzheimer's Disease (AD) cognitive and neuropathologic traits. The quantile–quantile plots of p-values showed that false positive rates were well calibrated by LMM, whereas other methods not accounting for sample-specific mixed effects led to serious inflation. LMM identified 37 potentially significant genes with differential expression in DLPFC for at least one of the AD traits, 17 of which were replicated in the additional RNA-Seq data of DLPFC, supplemental motor area, spinal cord, and muscle tissues. This application study showed not only well calibrated DGE results by LMM, but also possibly shared gene regulatory mechanisms of AD traits across different relevant tissues. [ABSTRACT FROM AUTHOR]

Wool fibre diameter (WFD) is one of the wool traits with higher economic impact. However, the main genes specifically regulating WFD remain unidentified. In this current work we have used Agilent Sheep Gene Expression Microarray and proteomic technology to investigate the gene expression patterns of body side skin, bearing more wool, in Aohan fine wool sheep, a Chinese indigenous breed, and compared them with that of small tail Han sheep, a sheep bread with coarse wool. Microarray analyses showed that most of the genes likely determining wool diameter could be classified into a few categories, including immune response, regulation of receptor binding and growth factor activity. Certain gene families might play a role in hair growth regulation. These include growth factors, immune cytokines, solute carrier families, cellular respiration and glucose transport amongst others. Proteomic analyses also identified scores of differentially expressed proteins. [ABSTRACT FROM AUTHOR]

Differential gene expression (DGE) analysis has been widely employed to identify genes expressed differentially with respect to a trait of interest using RNA sequencing (RNA-Seq) data. Recent RNA-Seq data with large samples pose challenges to existing DGE methods, which were mainly developed for dichotomous traits and small sample sizes. Especially, existing DGE methods are likely to result in inflated false positive rates. To address this gap, we employed a linear mixed model (LMM) that has been widely used in genetic association studies for DGE analysis of quantitative traits. We first applied the LMM method to the discovery RNA-Seq data of dorsolateral prefrontal cortex (DLPFC) tissue (n = 632) with four continuous measures of Alzheimer's Disease (AD) cognitive and neuropathologic traits. The quantile–quantile plots of p-values showed that false positive rates were well calibrated by LMM, whereas other methods not accounting for sample-specific mixed effects led to serious inflation. LMM identified 37 potentially significant genes with differential expression in DLPFC for at least one of the AD traits, 17 of which were replicated in the additional RNA-Seq data of DLPFC, supplemental motor area, spinal cord, and muscle tissues. This application study showed not only well calibrated DGE results by LMM, but also possibly shared gene regulatory mechanisms of AD traits across different relevant tissues. [ABSTRACT FROM AUTHOR]

Purpose: Previous evidence indicated anti-ageing potential of docosahexaenoic acid (DHA), but the underlying mechanism remains unclear. We investigated protective effect of DHA on telomere attrition and lipid disturbance in male mice with premature ageing caused by telomerase deficiency. Methods: Wild-type (WT) and fourth-generation telomerase-deficient (G4 Terc−/−, Terc knockout, KO) male mice (C57BL/6, 2 months old) were fed control diet (WT-C and KO-C groups) or DHA-enriched diet containing 0.80% DHA by weight (WT-DHA and KO-DHA groups) for 10 months. The ageing phenotypes and metabolic level [carbon dioxide emission, oxygen consumption, and respiratory exchange ratio (RER)] were assessed at the end of the experiment. Telomere length in various tissues and the hepatic gene and protein expression for regulating lipid synthesis and lipolysis were measured. Data were tested using one- or two-factor ANOVA. Results: In KO male mice, DHA prevented weight loss, corrected high RER, and reduced fat loss. Telomere shortening was reduced by 22.3%, 25.5%, and 13.5% in heart, liver, and testes of the KO-DHA group compared with those in the KO-C group. The KO-DHA group exhibited higher gene transcription involved in glycerol-3-phosphate pathway [glycerol-3-phosphate acyltransferase (Gpat)], lower gene expression of β-oxidation [carnitine palmitoyltransferase 1a (Cpt1a)], and upregulation of proteins in lipid synthesis [mammalian target of rapamycin complex 1 (mTORC1) and sterol responsive element binding protein 1 (SREBP1)] in liver than the KO-C group. Conclusion: Long-term DHA intervention attenuates telomere attrition and promotes lipid synthesis via the tuberous sclerosis complex 2 (TSC2)-mTORC1-SREBP1 pathway in KO male mice. [ABSTRACT FROM AUTHOR]

As novel master regulators of initiation and progression, long non-coding RNAs (lncRNAs) can response to therapy in a wide variety of malignances. This study aimed to explore the regulatory mechanisms in non-small cell lung cancer (NSCLC) by constructing lncRNA-miRNA-mRNA networks. The whole transcriptome sequence was performed in five NSCLC tissues and their corresponding paired adjacent normal tissues. MicroRNAs (miRNAs) and mRNAs expression profiles were obtained from TCGA database. "EdgeR" R package was used for gene expression comparisons and the genes with | log2FC |> 2 and false discovery rate (FDR) adjusted P<0.05 were considered significant. Totally, 559 differentially expressed lncRNAs (DELs) (235 up-regulated, 324 down-regulated) were identified via whole transcriptome sequence analysis. Subsequently, 14 up-regulated lncRNAs were obtained in the intersection of our RNA-seq data and TCGA dataset. The dysregulations of the selected lncRNAs were evaluated through GEPIA. Three candidate lncRNAs (LINC01426, TYMSOS, VPS9D1-AS1) were finally selected for further study. Through bioinformatics prediction, with the three lnRNAs and their associated miRNAs (n=14) and mRNAs (n=218), a lncRNA-miRNA-mRNA network was constructed. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway associated with the targeted genes were proceeded. In addition, the protein–protein interaction (PPI) network construction was performed by STRING and lncRNA-miRNA-mRNA regulatory network was visualized via Cytoscape. The top ten significant hub-genes in the PPI network are identified based on their node degrees. Moreover, the lncRNA-miRNA and miRNA-mRNA interactions were predicted using the StarBase and survival analyses were performed in Kaplan-Meier plotter website. We concluded that TYMSOS/hsa-miR-101-3p/CEP55 and TYMSOS/hsa-miR-195-5p/CHEK1 ceRNA regulatory aixs may play vital roles in the occurrence and development of NSCLC, and may provide novel therapeutic targets in the future. [ABSTRACT FROM AUTHOR]

Insulin/IGF-1 signaling (IIS) pathway is known to control growth, development and reproduction. Insulin-like peptide mediated body size plasticity in Drosophila melanogaster has been reported. Here, our studies showed that IIS pathway and nutrition regulate growth and maturation of the male accessory gland (MAG) in the red flour beetle, Tribolium castaneum. The size of MAG increased from day 1 to day 5 post-adult emergence (PAE). This increase in the size of MAG is contributed by an increase in cell size, but not cell number. The growth of MAG was impaired after double-stranded RNA (dsRNA)-mediated knockdown in the expression of genes coding for ILP3, InR, Chico, PI3k, AKT, and GATA1 involved in IIS pathway. Interestingly, starvation showed similar effects on the growth and maturation of MAG. The phenotypes observed in animals where IIS signaling pathway genes were knocked down are similar to the phenotypes observed after starving beetles for 5 days PAE. These data suggest that nutrition signals working through IIS pathway regulate maturation of MAG by promoting the growth of MAG cells. [ABSTRACT FROM AUTHOR]

Isodon rubescens (Hemsley) H. Hara (Lamiaceae) is a traditional Chinese medicine plant that has been used to treat various human diseases. Oridonin is one of the main active ingredients, and the route of its molecular biosynthesis remains to be determined. The study of gene expression patterns can provide clues toward the understanding of its biological functions. The selection of suitable reference genes for normalizing target gene expression is the first steps in any quantitative real-time PCR (RT-qPCR) gene expression study. Therefore, validation of suitable reference genes is necessary for obtaining reliable results in RT-qPCR analyses of I. rubescens. Here, 12 candidate reference genes were chosen, and their expression stability in different tissues of I. rubescens and in leaves under different abiotic stresses (NaCl, dehydration, SA, MeJA, and ABA) was evaluated using the ∆Ct, NormFinder, GeNorm, BestKeeper, and RankAggreg statistical tools. Analysis using the comprehensive tools of RankAggreg algorithm showed that GADPH, 18S and eIF were stably expressed in different tissues; UBQ, Apt, and HIS; Cycl, UBQ, and PP2A; GADPH, 18S, and eIF; eIF, UBQ, and PP2A; TUB, Cycl, and UBQ; were the best three candidate reference genes for the samples of Dehydration, NaCl, SA, MeJA, and ABA treatment, respectively. While for the concatenated sets of ND (NaCl and dehydration) and SMA (SA, MeJA, and ABA), UBQ, HIS, and TUA; UBQ, eIF and Apt were the three appropriate candidate reference genes, respectively. In addition, the expression patterns of HMGR in different tissues and under different treatments were used to confirm the reliability of the selected reference genes, indicating that the use of an inappropriate reference gene as the internal control will cause results with a large deviation. This work is the first study on the expression stability of reference genes in I. rubescens and will be particularly useful for gene functional research in this species. [ABSTRACT FROM AUTHOR]

Key message: Abscisic acid induced the expression of AsKIN during the recovery period of garlic cryopreservation. AsKIN was identified as a gene involved in cold and osmotic stress resistance. Cryopreservation has been proven to be effective in removing viruses from garlic. However, oxidative damage in cryopreservation has a significant impact on the survival after preservation. Abscisic acid (ABA) has been shown to reduce oxidative stress and promote the survival after cryopreservation. However, it is not clear which genes play important roles in this process. In this study, we added ABA to the dehydration step and analyzed the transcriptomic divergences between the ABA-treated group and the control group in three cryogenic steps (dehydration, unloading and recovery). By short time-series expression miner (STEM) analysis and weighted gene co-expression network analysis (WGCNA), the recovery step was identified as the period of significant changes in gene expression levels in cryopreservation. The addition of ABA promoted the upregulated expression of microtubule-related genes in the recovery step. We further identified AsKIN as a hub gene in the recovery step and verified its function. The results showed that overexpression of AsKIN enhanced the tolerance of Arabidopsis to oxidative stress in cryopreservation, influenced the expression of genes in response to cold and osmotic stress and promoted plant growth after stress. The AsKIN gene is likely to be involved in the plant response to cold stress and osmotic stress. These results reveal the molecular mechanisms of ABA in cryopreservation and elucidate the potential biological functions of the kinesin-14 subfamily. [ABSTRACT FROM AUTHOR]

SLC16A family members play crucial roles in tumorigenesis and tumor progression. However, the exact role of distinct members in the SLC16A family in human pancreatic cancer remains unclear. Integrated bioinformatics analysis for the identification of therapeutic targets for certain cancers based on transcriptomics, proteomics and high-throughput sequencing could help us obtain novel information and understand potential underlying molecular mechanisms. In the present study, we investigated SLC16A family members in pancreatic cancer through accumulated data from GEO (Gene Expression Omnibus), TCGA (The Cancer Genome Atlas) and other available databases. The expression profile, clinical application significance and prognostic value of the SLC16A family for patients with pancreatic cancer were explored. SLC16A1, SLC16A3 and SLC16A13 exhibited biomarker potential for prognosis, and we further identified their related genes and regulatory networks, revealing core molecular pathways that require further investigation for pancreatic cancer. [ABSTRACT FROM AUTHOR]

In this study, we investigated low-frequency and rare variants associated with blood pressure (BP) by focusing on a linkage region on chromosome 16p13. We used whole genome sequencing (WGS) data obtained through the NHLBI Trans-Omics for Precision Medicine (TOPMed) program on 395 Cleveland Family Study (CFS) European Americans (CFS-EA). By analyzing functional coding variants and non-coding rare variants with CADD score > 10 residing within the chromosomal region in families with linkage evidence, we observed 25 genes with nominal statistical evidence (burden or SKAT p < 0.05). One of the genes is RBFOX1, an evolutionarily conserved RNA-binding protein that regulates tissue-specific alternative splicing that we previously reported to be associated with BP using exome array data in CFS. After follow-up analysis of the 25 genes in ten independent TOPMed studies with individuals of European, African, and East Asian ancestry, and Hispanics (N = 29,988), we identified variants in SLX4 (p = 2.19 × 10−4) to be significantly associated with BP traits when accounting for multiple testing. We also replicated the associations previously reported for RBFOX1 (p = 0.007). Follow-up analysis with GTEx eQTL data shows SLX4 variants are associated with gene expression in coronary artery, multiple brain tissues, and right atrial appendage of the heart. Our study demonstrates that linkage analysis of family data can provide an efficient approach for detecting rare variants associated with complex traits in WGS data. [ABSTRACT FROM AUTHOR]

Origin of replication (ori in theta-replicating plasmids or dso in rolling circle replicating plasmids) initiates plasmid replication in a broad range of bacteria. These two kinds of plasmids were both identified in Streptococcus, a genus composed of both human commensal bacteria and pathogens with the ability to cause severe community-acquired infections, including meningitides, septicemia, and respiratory tract diseases. Given the important roles of Streptococcus in the exchange of genetic elements with other symbiotic microbes, the genotypes and phenotypes of both Streptococcus spp. and other symbiotic species could be changed during colonization of the host. Therefore, an improved plasmid system is required to study the functional, complicated, and changeable genomes of Streptococcus. In this study, a dual-replicon shuttle vector system named pDRE was constructed to achieve heterologous gene expression. The vector system contained theta replicon for Escherichia coli. The origin of rolling circle replicon was synthesized according to pMV158 in Gram-positive bacteria. By measuring the products of inserted genes at multiple cloning sites, the ability of this vector system in the replication and expression of heterologous genes was assessed in four Streptococcus and three other Gram-positive bacteria: Bacillus subtilis, Lactococcus lactis, and Staphylococcus aureus. The results showed that the newly constructed vector could simultaneously replicate and express heterologous genes in a broad range of Gram-positive and Gram-negative bacteria, thus providing a potentially powerful genetic tool for further functional analysis. [ABSTRACT FROM AUTHOR]

Macrophages are central players in inflammation, which leads to liver injury. It has been reported that continuous macrophage activation initiates this process. Our previous data show that the anti-inflammatory factor, 15-deoxy-Δ-prostaglandin J (15d-PGJ), inhibits bone marrow (BM)-derived macrophage (BMM) migration and inflammatory cytokine production. However, the underlying mechanism of 15d-PGJ inhibited BMM activation is still unclear. Here, we evaluate the role of 15d-PGJ/PPARγ axis in BMM activation. 15d-PGJ reduced activated BMM population in injured livers. Inflammatory cytokine expressions (MIP-1β, TNF-α, NOS2) were depressed by 15d-PGJ in macrophages isolated from treated livers. In vitro, 15d-PGJ inhibited BMM activation via PPARγ. Moreover, miR-27b-3p, miR-181a-1-3p, and miR-326-5p target MIP-1β, TNF-α, and NOS2 mRNA, respectively. The miRNA expressions were decreased in damaged livers, macrophages isolated from injured livers, and activated BMMs, which were renewed by 15d-PGJ/PPARγ axis. In activated BMMs, the miRNA inhibitors attenuated inhibitory effect of PPARγ agonist (troglitazone or ciglitazone), while replenishing the lack of miRNAs induced by PPARγ deficiency using miRNA mimics caused a decline of inflammatory cytokines. In conclusion, these data suggest that 15d-PGJ/PPARγ axis regulates BMM activation via promoting miR-27b-3p, miR-181a-1-3p, and miR-326-5p expressions. Key messages: [ABSTRACT FROM AUTHOR]

Circadian rhythms are a collection of endogenously driven biochemical, physiological, and behavioral processes that oscillate in a 24-h cycle and can be entrained by external cues. Circadian clock molecules are responsible for the expression of regulatory components that modulate, among others, the cell's metabolism and energy consumption. In clinical practice, the regulation of clock mechanisms is relevant to biotransformation of therapeutics. Accordingly, xenobiotic metabolism and detoxification, the two processes that directly influence drug effectiveness and toxicity, are direct manifestations of the daily oscillations of the cellular and biochemical processes taking place within the gastrointestinal, hepatic/biliary, and renal/urologic systems. Consequently, the impact of circadian timing should be factored in when developing therapeutic regimens aimed at achieving maximum efficacy, minimum toxicity, and decreased adverse effects in a patient. However, and despite a strong mechanistic foundation, only 0.16 % of ongoing clinical trials worldwide exploit the concept of 'time-of-day' administration to develop safer and more effective therapies. In this article, we (1) emphasize points of control at which circadian biology intersects critical processes governing treatment interventions; (2) explore the extent to which chronotherapeutics are incorporated into clinical trials; (3) recognize roadblocks; and (4) recommend approaches to precipitate the integration of chronobiological concepts into clinical practice. [ABSTRACT FROM AUTHOR]

Kashin-Beck disease (KBD) is a chronic and deformed endemic osteoarthritis, without fully known etiology. As compared to primary osteoarthritis (POA), its pathogenesis still exists controversial. We performed this study to discriminate the difference in genes expression involved in apoptosis from KBD cartilages and POA cartilages. Microarray analysis and Ingenuity Pathway Analysis (IPA) were used to identify the molecular mechanisms/canonical pathways implicated in KBD arthritis. Immunohistochemistry staining was carried out to detect tissue distribution of PDCD5 and EGR1 in the knee cartilages from KBD and POA. The 23 up-regulated apoptosis-related genes with >2-fold change were identified. 'Role of Macrophages, Fibroblasts, and Endothelial Cells in Rheumatoid Arthritis' signaling was screened as the most relevant pathway through IPA. The 8 differentially regulated genes were verified by qRT-PCR analysis. Programmed cell death 5 in middle and deeper zones and early growth response 1 in superficial and middle zones showed protein overexpressed in KBD cartilage samples comparing to those in POA cartilage samples. Data indicates a higher expression level of apoptosis-related functional genes in KBD articular cartilage compared with POA articular cartilage. [ABSTRACT FROM AUTHOR]

Mouse models have been widely utilized to elucidate the basic principles and regulatory mechanisms of primordial follicle activation. Outside their natural environment, the growth of follicles might be affected by unknown factors in vitro and the elimination of regulation in vivo. Currently, in vitro culture and transplantation of ovaries under the kidney capsule are two commonly used incubation methods. However, the limited number of studies that have been published compare various incubation systems and reveal differences between ovaries that are incubated and grown in vivo. We compare the number of primordial, primary and secondary follicles in cultured, transplanted and in-vivo-grown ovaries. We investigate the expression levels of four genes, including zona pellucida 3 (ZP3), growth and differentiation factor-9 (GDF-9), proliferating cell nuclear antigen (PCNA) and anti-Müllerian hormone (AMH). Our results suggest that in vitro culture accelerates follicle activation, delays the transition from primary to secondary follicles and affects the expression patterns of ZP3, GDF-9, PCNA and AMH. A larger number of secondary follicles in ovaries cultured in alpha-minimal essential medium (α-MEM) had intact zona pellucida compared with those grown in Dulbecco's modified Eagle medium containing Ham's F-12 nutrient mixture (D/F12), suggesting that α-MEM is a better basal medium. The transplanted ovaries demonstrated the most similar characteristics to the in-vivo-grown ovaries, indicating that transplantation provided an optimal environment for ovarian incubation. This study has thus established the similarities and differences between in-vivo-grown and incubated ovaries, demonstrated that transplantation can mostly mimic the environment of ovarian growth in vivo and determined the optimal basal culture medium between α-MEM and D/F12. [ABSTRACT FROM AUTHOR]

Purpose: Hirschsprung's disease (HSCR) is characterized by absence of the enteric nervous system in a variable portion of the distal gut. The endothelin receptor type B ( EDNRB) gene has been localized to the chromosome 13q22 region and encodes a G-protein coupled receptor, is generally accepted as a crucial gene for HSCR. This study is to identify the epigenetic changes of EDNRB in the pathogenesis of HSCR. Methods: We investigated the expression levels of EDNRB in 58 HSCR patients and 25 unrelated controls, using reverse transcriptase polymerase chain reaction (RT-PCR) and western blot assay. Moreover, using the methylation-specific polymerase chain reaction, we examined the methylation status of the promoter region of EDNRB. Results: Aberrant high expression level of EDNRB was detected in HSCR patients compared with the control group ( P = 0.023). Besides, western blot assay confirmed the up-regulation of EDNRB in the post transcription level in the aganglionosis segment of HSCR patients. Furthermore, there was a significantly lower ratio of methylation level of EDNRB in HSCR. Conclusions: Our study demonstrates that epigenetic inactivation of the EDNRB gene may play a role in the development of HSCR. [ABSTRACT FROM AUTHOR]

Background: Osteopontin (OPN) is associated with the Th1 immune response in inflammatory bowel diseases (IBD). While OPN has been shown to play an important role in maintaining the epithelial barrier, its role in IBD remains unclear. Aim: The aim of this study was to assess OPN function in patients with IBD and in the mouse colitis model. Methods: Osteopontin expression in colonic samples from IBD patients was determined by a semi-quantitative immunohistochemical staining method. Colitis in BALB/c mice was induced by 5 % dextran sodium sulfate (DSS), followed by treatment with salazosulfapyridine (SASP) and infliximab, respectively. The plasma OPN concentration was measured by an enzyme-linked immunosorbent assay. The expression of OPN in colonic tissues was detected by reverse transcriptase PCR, real-time PCR and Western blot, and the localization of OPN was determined by a semi-quantitative immunohistochemical staining method. The immune function of OPN was investigated by measuring the production of cytokines, and the amount of cytokines produced was then used to determine OPN immune functions. Results: Osteopontin expression in intestinal epithelial cells was significantly lower in IBD patients than in controls, while its expression in lamina propria exudative cells was significantly higher in IBD patients than in controls. In DSS-induced mice, OPN expression in plasma and colonic tissues increased significantly, and this increase was significantly reduced after the mice were treated with SASP and infliximab. OPN promoted the Th1 immune response and strengthened inflammation in the mouse colitis model. Conclusions: Our results indicate that OPN plays an important role in the immune response and is also involved in the mucosal protective mechanism in IBD. [ABSTRACT FROM AUTHOR]

The cya gene encoding adenylate cyclase was cloned from Arthrobacter sp. CGMCC 3584 by thermal asymmetric interlaced PCR for the first time. It exhibited an open reading frame containing 1,125 bp and encoding 374 amino acids. Amino acid sequence analysis showed that this enzyme was a class III adenylate cyclase. Expression of the cya gene was carried out in Escherichia coli Rosetta, and purification was performed via Ni-NTA agarose gel column. SDS-PAGE indicated that the molecular mass of the recombinant adenylate cyclase was 45 kDa. The V and K were determined to be 5.06 μmol/min/mg and 7.56 mM, respectively. The optimum pH and temperature were 8.0 and 35 °C. Several divalent metal ions were found to activate the enzyme to different extents, and the maximal specific activity reached 3.04 μmol/min/mg when 50 mM Mg was added. This was the first report of the cloning of an adenylate cyclase gene from Arthrobacter sp. [ABSTRACT FROM AUTHOR]

There is accumulating evidence that cancer stem cells (CSCs) play an important role in tumor progression. Novel strategies targeting CSCs have been widely researched. In the present study, we explored whether such CSCs existed in human ovarian cancer (OVCA) cell line and whether anti-CD44 antibody had effects on such subpopulation. We isolated and identified spheroid cells from SKOV-3. Then we used A3D8, an anti-CD44 mAb to treat spheroid cells with so-called 'stemness'. Effects of A3D8 on spheroid cells' biological behaviors were examined. Our findings showed that there was a small subpopulation that had so-called 'stemness' in SKOV-3 cell line. Against spheroid cells, A3D8 can (1) inhibit cell proliferation; (2) change cell cycle distribution and expression of p21, CDK2 and cyclinA; (3) enhance cisplatin (DDP)-induced apoptosis; (4) promote cell differentiation; (5) inhibit clone formation efficiency; (6) reduce invasive efficacy; (7) inhibit tumorigenicity. Thus, to sum up points which we have just showed, spheroid cells isolated from SKOV-3 can be used as an appropriate in vitro model for relevant study of human ovarian CSCs. And our results reasoned that anti-CD44 therapy may become a potential promising strategy for OVCA treatment. [ABSTRACT FROM AUTHOR]

In this study, we investigated the correlation between p53 and bcl-2 in gambogic acid (GA)-induced apoptosis. MTT assay was employed to evaluate MCF-7 cell viability after GA treatment. Cell morphological changes were observed follow-up under the inverted microscope after GA withdrawal. To observe the cell apoptosis, DAPI staining was used. Meanwhile, p53 small interfering RNA (si-RNA) was adopted to knock p53 down. All expressions of p53 and bcl-2 were evaluated by Western blot analysis. MTT assay showed that GA inhibited MCF-7 cell growth effectively in a time-dependent manner. With 0.5 h GA treatment, p53 was significantly increased, whereas bcl-2 was reduced potently with 6 h GA treatment. After GA withdrawal, p53 expressions were maintained in high levels. Furthermore, bcl-2 decreasing was attenuated after co-treatment with PFT alpha, a p53 transcription blocker. Same result was observed after p53 knock-down by p53 si-RNA. Gambogic acid induced human breast cancer cells MCF-7 apoptosis by reducing bcl-2 expression via p53. [ABSTRACT FROM AUTHOR]

Single nucleotide polymorphisms (SNPs) and genes associated with susceptibility to hepatitis B virus (HBV) infection that have been identified by genome-wide association studies explain only a limited portion of the known heritability, indicating more genetic variants remain to be discovered. In this study, we adopted a new research strategy to identify more susceptibility genes and variants for HBV infection. We first performed genetic association analysis of 300 sib-pairs and 3,087 case-control samples, which revealed that 36 SNPs located in 31 genes showed nominal associations with HBV infection in both samples. Of these genes, we selected SEC24D for further molecular analysis according to the following two main lines of evidence. First, a time course analysis of the expression profiles from HBV-infected primary human hepatocytes (PHH) demonstrated that SEC24D expression increased markedly as time passed after HBV infection (P = 4.0 × 10−4). Second, SNP rs76459466 in SEC24D was adversely associated with HBV risk (ORmeta = 0.82; Pmeta = 0.002), which again indicated that SEC24D represents a novel susceptibility gene for HBV infection. Moreover, SEC24D appeared to be protective against HBV infection in vitro. Consistently, we found that SEC24D expression was significantly enhanced in non-infected liver tissues (P = 0.002). We conclude that SEC24D is a novel candidate gene linked to susceptibility to HBV infection. [ABSTRACT FROM AUTHOR]

To investigate the relationship between starch hydrolysis and Monascus pigments (MPs) production, the α-amylase gene (AOamyA) from Aspergillus oryzae was heterologously expressed in Monascus ruber CICC41233, and we obtained a positive transformant named Monascus ruber Amy9. In M. ruber Amy9, the α-amylase activities were 6.65- and 4.26-fold higher at 72 h and 144 h, respectively, than those in the parent strain with the glucose as solo carbon medium. Surprisingly, in the MPs fermentation medium with rice powder as solo material, M. ruber Amy9 completely degraded starch at 48 h, while 43.93 and 7.29 mg/mL starch remained at 48 and 144 h, respectively, in the parent strain. Monascus ruber Amy9 accelerated starch hydrolysis, which enhanced biomass and also increased total MPs by 132% after 144 h. Compared with M. ruber CICC41233, the relative gene expression levels, as determined by a quantitative real-time polymerase chain reaction analysis, of acl2 encoding ATP-citrate lyase subunit 2, pks encoding polyketide synthase, and fasB encoding the fatty acid synthase beta subunit increased by 33.14, 145.18, and 32.15%, respectively, after 144 h in M. ruber Amy9. The up-regulated expression of these key genes in MPs synthesis contributed to the large increase in MPs production. This interesting work provided us with a new idea and a new target for the study of the MPs production. [ABSTRACT FROM AUTHOR]