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Influenza A virus (IAV) is a leading cause of human respiratory infections and poses a major public health concern. IAV replication can affect the expression of DNA methyltransferases (DNMTs), and the subsequent changes in DNA methylation regulate gene expression and may lead to abnormal gene transcription and translation, yet the underlying mechanisms of virus-induced epigenetic changes from DNA methylation and its role in virus–host interactions remain elusive. Here in this paper, we showed that DNMT1 expression could be suppressed following the inhibition of miR-142-5p or the PI3K/AKT signaling pathway during IAV infection, resulting in demethylation of the promotor region of the 2′-5′-oligoadenylate synthetase-like (OASL) protein and promotion of its expression in A549 cells. OASL expression enhanced RIG-I-mediated interferon induction and then suppressed replication of IAV. Our study elucidated an innate immunity mechanism by which up-regulation of OASL contributes to host antiviral responses via epigenetic modifications in IAV infection, which could provide important insights into the understanding of viral pathogenesis and host antiviral defense. [ABSTRACT FROM AUTHOR]

Hypertension (HTN) is associated with gut dysbiosis and the depletion of butyrate-producing bacteria in animal models and people. Furthermore, fecal material transfer from donor hypertensive patients increases blood pressure in normotensive recipient animals and ameliorates HTN-associated pathophysiology. These observations have implications in the impaired interactions between the gut and gut microbiota in HTN. Although this concept is supported in animal models, little is known about human HTN. Therefore, our objective for this study was to compare gene expression with transcriptomics and its potential to influence microbiota in subjects with normal and high blood pressure (HBP). Colon samples from reference subjects with normal blood pressure (REF) and HBP were used for RNA-seq to analyze their transcriptomes. We observed the significant downregulation of gene sets governing immune responses (e.g., SGK1 and OASL), gut epithelial function (e.g., KRT20 and SLC9A3R1), gut microbiota (e.g., PPARG and CIDEC) and genes associated with cardiovascular and gut diseases (e.g., PLAUR and NLN) in HBP subjects; the expression of genes within these pathways correlated with blood pressure. Potential drug targets in the gut epithelium were identified using the Drug Gene International Database for possible use in HTN. They include peroxisome proliferator-activated receptor gamma (PPRG), active serum/glucocorticoid regulated kinase 1 (SGK1) and 3 beta-hydroxysteroid isomerase type II inhibitor (HSD3B). Finally, butyrate, a microbiota-derived short-chain fatty acid, restored the disrupted expression of certain functional genes in colonic organoids from HBP subjects. Patients with HBP exhibit a unique transcriptome that could underlie impaired gut–microbiota interactions. Targeting these interactions could provide a promising new therapeutic intervention for hypertension management. [ABSTRACT FROM AUTHOR]

Roselle (Hibiscus sabdariffa L.) is an annual herbaceous plant of the genus Hibiscus in family Malvaceae. Roselle calyxes are rich in anthocyanins, which play important roles in human health. However, limited information is available on anthocyanin biosynthesis in the roselle calyx. In this study, transcriptomic and metabolomic analyses were performed to identify the key genes involved in anthocyanin biosynthesis in the roselle calyx. Three roselle cultivars with different calyx colors, including FZ-72 (red calyx, R), Baitao K (green calyx, G), and MG5 (stripped calyx, S), were used for metabolomic analyses with UPLC-Q-TOF/MS and RNA-seq. Forty-one compounds were quantified, including six flavonoids and 35 anthocyanins. The calyx of FZ-72 (red calyx) had the highest contents of anthocyanin derivatives such as delphinidin-3-O-sambubioside (955.11 μg/g) and cyanidin-3-O-sambubioside (531.37 μg/g), which were responsible for calyx color, followed by those in MG5 (stripped calyx) (851.97 and 330.06 μg/g, respectively). Baitao K (green calyx) had the lowest levels of these compounds. Furthermore, RNA-seq analysis revealed 114,415 differentially expressed genes (DEGs) in the calyxes at 30 days after flowering (DAF) for the corresponding cultivars FZ-72 (R), Baitao K (G), and MG5(S). The gene expression levels in the calyxes of the three cultivars were compared at different flowering stages, revealing 11,555, 11,949, and 7177 DEGs in R vs. G, R vs. S, and G vs. S, respectively. Phenylpropanoid and flavonoid biosynthesis pathways were found to be enriched. In the flavonoid pathway, 29, 28, and 27 genes were identified in G vs. R, G vs. S, and S vs. R, respectively. In the anthocyanin synthesis pathway, two, two, and one differential genes were identified in the three combinations; these differential genes belonged to the UFGT gene family. After joint analysis of the anthocyanin content in roselle calyxes, nine key genes belonging to the CHS, CHI, UFGT, FLS, ANR, DFR, CCoAOMT, SAT, and HST gene families were identified as strongly related to anthocyanin synthesis. These nine genes were verified using qRT-PCR, and the results were consistent with the transcriptome data. Overall, this study presents the first report on anthocyanin biosynthesis in roselle, laying a foundation for breeding roselle cultivars with high anthocyanin content. [ABSTRACT FROM AUTHOR]

Cadmium (Cd) is one of the toxic heavy metals found widely in the environment. Skin is an important target organ of Cd exposure. However, the adverse effects of Cd on human skin are still not well known. In this study, normal human skin keratinocytes (HaCaT cells) were studied for changes in cell viability, morphology, DNA damage, cycle, apoptosis, and the expression of endoplasmic reticulum (ER) stress-related genes (XBP-1, BiP, ATF-4, and CHOP) after exposure to Cd for 24 h. We found that Cd decreased cell viability in a concentration-dependent manner, with a median lethal concentration (LC50) of 11 µM. DNA damage induction was evidenced by upregulation of the level of γ-H2AX. Furthermore, Cd induced G0/G1 phase cell cycle arrest and apoptosis in a dose-dependent manner and upregulated the mRNA levels of ER stress biomarker genes (XBP-1, BiP, ATF4, and CHOP). Taken together, our results showed that Cd induced cytotoxicity and DNA damage in HaCaT cells, eventually resulting in cell cycle arrest in the G0/G1 phase and apoptosis. In addition, ER stress may be involved in Cd-induced HaCaT apoptosis. Our data imply the importance of reducing Cd pollution in the environment to reduce its adverse impacts on human skin. [ABSTRACT FROM AUTHOR]

The development of high-throughput omics technologies has enabled the quantification of vast amounts of genes and gene products in the whole genome. Pathway enrichment analysis (PEA) provides an intuitive solution for extracting biological insights from massive amounts of data. Topology-based pathway analysis (TPA) represents the latest generation of PEA methods, which exploit pathway topology in addition to lists of differentially expressed genes and their expression profiles. A subset of these TPA methods, such as BPA, BNrich, and PROPS, reconstruct pathway structures by training Bayesian networks (BNs) from canonical biological pathways, providing superior representations that explain causal relationships between genes. However, these methods have never been compared for their differences in the PEA and their different topology reconstruction strategies. In this study, we aim to compare the BN reconstruction strategies of the BPA, BNrich, PROPS, Clipper, and Ensemble methods and their PEA and performance on tumor and non-tumor classification based on gene expression data. Our results indicate that they performed equally well in distinguishing tumor and non-tumor samples (AUC > 0.95) yet with a varying ranking of pathways, which can be attributed to the different BN structures resulting from the different cyclic structure removal strategies. This can be clearly seen from the reconstructed JAK-STAT networks by different strategies. In a nutshell, BNrich, which relies on expert intervention to remove loops and cyclic structures, produces BNs that best fit the biological facts. The plausibility of the Clipper strategy can also be partially explained by intuitive biological rules and theorems. Our results may offer an informed reference for the proper method for a given data analysis task. [ABSTRACT FROM AUTHOR]

Polymerase chain reaction (PCR) is a widely used technique in gene expression analysis, diagnostics, and various molecular biology applications. However, the accuracy and sensitivity of PCR can be compromised by primer–template mismatches, potentially leading to erroneous results. In this study, we strategically designed 111 primer–template combinations with varying numbers, types, and locations of mismatches to meticulously assess their impact on qPCR performance while two distinctly different types of DNA polymerases were used. Notably, when a single-nucleotide mismatch occurred at the 3' end of the primer, we observed significant decreases in the analytical sensitivity (0–4%) with Invitrogen™ Platinum™ Taq DNA Polymerase High Fidelity, while the analytical sensitivity remained unchanged with Takara Ex Taq Hot Start Version DNA Polymerase. Leveraging these findings, we designed a highly specific PCR to amplify Babesia while effectively avoiding the genetically close Theileria. Through elucidating the critical interplay between types of DNA polymerases and primer–template mismatches, this research provides valuable insights for improving PCR accuracy and performance. These findings have important implications for researchers aiming to achieve robust qPCR results in various molecular biology applications. [ABSTRACT FROM AUTHOR]

However, the honey bees exhibited a reduction in consuming the sucrose containing not less than 1 ug/mL of CdCl SB 2 sb ; although non-significant, there exists the possibility that sucrose containing more than 1 ug/mL of Cd may affect honey bee sucrose responsiveness that is essential for olfactory learning in honey bees [[38]]. We, therefore, investigated whether honey bees exposed to chronic Cd exhibited altered olfactory learning, and we further compared the patterns of brain gene expression between cadmium-exposed bees and control bees using RNA-seq analysis. Our studies showed that the head weight was significantly lower in cadmium-exposed bees when compared to controls, suggesting chronic Cd exposure may have adverse effects on the total protein content of the mandibular and hypopharyngeal glands in the heads of adult honey bees, thus resulting in a decrease in head weight in Cd-exposed bees. [Extracted from the article]

Previous studies indicated that extensive genetic variations could be generated due to polyploidy, which is considered to be closely associated with the manifestation of polyploid heterosis. Our previous studies confirmed that triploid loquats demonstrated significant heterosis, other than the ploidy effect, but the underlying mechanisms are largely unknown. This study aimed to overcome the narrow genetic distance of loquats, increase the genetic variation level of triploid loquats, and systematically illuminate the heterosis mechanisms of triploid loquats derived from two cross combinations. Here, inter-simple sequence repeats (ISSRs) and simple sequence repeats (SSRs) were adopted for evaluating the genetic diversity, and transcriptome sequencing (RNA-Seq) was performed to investigate gene expression as well as pathway changes in the triploids. We found that extensive genetic variations were produced during the formation of triploid loquats. The polymorphism ratios of ISSRs and SSRs were 43.75% and 19.32%, respectively, and almost all their markers had a PIC value higher than 0.5, suggesting that both ISSRs and SSRs could work well in loquat assisted breeding. Furthermore, our results revealed that by broadening the genetic distance between the parents, genetic variations in triploids could be promoted. Additionally, RNA-Seq results suggested that numerous genes differentially expressed between the triploids and parents were screened out. Moreover, KEGG analyses revealed that "photosynthetic efficiency" and "glyco-metabolism" were significantly changed in triploid loquats compared with the parents, which was consistent with the results of physiological indicator analyses, leaf micro-structure observations, and qRT-PCR validation. Collectively, our results suggested that extensive genetic variations occurred in the triploids and that the changes in the "photosynthetic efficiency" as well as "glyco-metabolism" of triploids might have further resulted in heterosis manifestation in the triploid loquats. [ABSTRACT FROM AUTHOR]

HIV-1 Tat Interactive Protein 2 (HTATIP2) is a tumor suppressor, of which reduced or absent expression is associated with increased susceptibility to tumorigenesis and enhanced tumor invasion and metastasis. However, whether the absent expression of HTATIP2 is a tumor-promoting factor that acts through improving tumor adaptation to hypoxia is unclear. Here, we established a stable HTATIP2-knockdown A549 human lung adenocarcinoma cell line (A549shHTATIP2) using lentiviral-delivered HTATIP2-targeting short hairpin RNA (shRNA), employed a double subcutaneous xenograft model and incorporated photoacoustic imaging and metabolomics approaches to elucidate the impact of the absent HTATIP2 expression on tumor response to hypoxic stress. Results from the in vivo study showed that A549shHTATIP2 tumors exhibited accelerated growth but decreased intratumoral oxygenation and angiogenesis and reduced sensitivity to sorafenib treatment as compared with their parental counterparts. Moreover, results of the immunoblot and real-time PCR analyses revealed that the HIF2α protein and mRNA levels in vehicle-treated A549shHTATIP2 tumors were significantly increased (p < 0.01 compared with the parental control tumors). Despite the strong HIF2α-c-Myc protein interaction indicated by our co-immunoprecipitation data, the increase in the c-Myc protein and mRNA levels was not significant in the A549shHTATIP2 tumors. Nonetheless, MCL-1 and β-catenin protein levels in A549shHTATIP2 tumors were significantly increased (p < 0.05 compared with the parental control tumors), suggesting an enhanced β-catenin/c-Myc/MCL-1 pathway in the absence of HTATIP2 expression. The finding of significantly decreased E-cadherin (p < 0.01 compared with vehicle-treated A549shHTATIP2 tumors) and increased vimentin (p < 0.05 compared with sorafenib-treated A549 tumors) protein levels in A549shHTATIP2 tumors implicates that the absence of HTATIP2 expression increases the susceptibility of A549 tumors to sorafenib-activated epithelial-mesenchymal transition (EMT) process. Comparison of the metabolomic profiles between A549 and A549shHTATIP2 tumors demonstrated that the absence of HTATIP2 expression resulted in increased tumor metabolic plasticity that enabled tumor cells to exploit alternative metabolic pathways for survival and proliferation rather than relying on glutamine and fatty acids as a carbon source to replenish TCA cycle intermediates. Our data suggest a mechanism by which the absent HTATIP2 expression modulates tumor adaptation to hypoxia and promotes an aggressive tumor phenotype by enhancing the HIF2α-regulated β-catenin/c-Myc/MCL-1 signaling, increasing the susceptibility of tumors to sorafenib treatment-activated EMT process, and improving tumor metabolic plasticity. [ABSTRACT FROM AUTHOR]

CDK8/19 kinases, which mediate transcriptional reprogramming, have become an active target for cancer drug discovery. Several small-molecule CDK8/19 inhibitors showed in vivo efficacy and two have entered clinical trials, with no significant toxicities reported. However, Clarke et al. (eLife 2016; 5; e20722) found severe systemic toxicity associated with two potent CDK8/19 inhibitors, Cmpd3 (CCT251921) and Cmpd4 (MSC2530818), and suggested that their toxicity was due to on-target effects. Here, we compared five CDK8/19 inhibitors: Cmpd3, Cmpd4, Senexin B, 16-didehydro-cortistatin A (dCA) and 15w, in different assays. Only Cmpd4 showed striking toxicity in developing zebrafish. In cell-based assays for CDK8 and CDK19 inhibition, Cmpd3, Cmpd4, dCA and 15w showed similar low-nanomolar potency and efficacy against CDK8 and CDK19, while Senexin B was less potent. Only dCA produced sustained inhibition of CDK8/19-dependent gene expression. While toxicity of different compounds did not correlate with their effects on CDK8 and CDK19, kinome profiling identified several off-target kinases for both Cmpd3 and Cmpd4, which could be responsible for their toxicity. Off-target activities could have been achieved in the study of Clarke et al. due to high in vivo doses of Cmpd3 and Cmpd4, chosen for the ability to inhibit STAT1 S727 phosphorylation in tumor xenografts. We show here that STAT1 S727 phosphorylation is induced by various cytokines and stress stimuli in CDK8/19-independent manner, indicating that it is not a reliable pharmacodynamic marker of CDK8/19 activity. These results illustrate the need for careful off-target analysis and dose selection in the development of CDK8/19 inhibitors. [ABSTRACT FROM AUTHOR]

Forkhead Box M1 (FOXM1) is an oncogenic transcription factor implicated in the pathogenesis of solid and hematologic cancers. In this study, we examined the significance of FOXM1 in NPM-ALK-positive anaplastic large cell lymphoma (NPM-ALK + ALCL), with a focus on how it interacts with NPM-ALK, which is a key oncogenic driver in these tumors. FOXM1 was expressed in NPM-ALK + ALCL cell lines (5/5), patient samples (21/21), and tumors arising in NPM-ALK transgenic mice (4/4). FOXM1 was localized in the nuclei and confirmed to be transcriptionally active. Inhibition of FOXM1 in two NPM-ALK + ALCL cells using shRNA and pharmalogic agent (thiostrepton) resulted in reductions in cell growth and soft-agar colony formation, which were associated with apoptosis and cell-cycle arrest. FOXM1 is functionally linked to NPM-ALK, as FOXM1 enhanced phosphorylation of the NPM-ALK/STAT3 axis. Conversely, DNA binding and transcriptional activity of FOXM1 was dependent on the expression of NPM-ALK. Further studies showed that this dependency hinges on the binding of FOXM1 to NPM1 that heterodimerizes with NPM-ALK, and the phosphorylation status of NPM-ALK. In conclusion, we identified FOXM1 as an important oncogenic protein in NPM-ALK+ ALCL. Our results exemplified that NPM-ALK exerts oncogenic effects in the nuclei and illustrated a novel role of NPM1 in NPM-ALK pathobiology. [ABSTRACT FROM AUTHOR]

The loquat (Eriobotrya japonica L.) is a special evergreen tree, and its fruit is of high medical and health value as well as having stable market demand around the world. In recent years, research on the accumulation of nutrients in loquat fruit, such as carotenoids, flavonoids, and terpenoids, has become a hotspot. The SBP-box gene family encodes transcription factors involved in plant growth and development. However, there has been no report on the SBP-box gene family in the loquat genome and their functions in carotenoid biosynthesis and fruit ripening. In this study, we identified 28 EjSBP genes in the loquat genome, which were unevenly distributed on 12 chromosomes. We also systematically investigated the phylogenetic relationship, collinearity, gene structure, conserved motifs, and cis-elements of EjSBP proteins. Most EjSBP genes showed high expression in the root, stem, leaf, and inflorescence, while only five EjSBP genes were highly expressed in the fruit. Gene expression analysis revealed eight differentially expressed EjSBP genes between yellow- and white-fleshed fruits, suggesting that the EjSBP genes play important roles in loquat fruit development at the breaker stage. Notably, EjSBP01 and EjSBP19 exhibited completely opposite expression patterns between white- and yellow-fleshed fruits during fruit development, and showed a close relationship with SlCnr involved in carotenoid biosynthesis and fruit ripening, indicating that these two genes may participate in the synthesis and accumulation of carotenoids in loquat fruit. In summary, this study provides comprehensive information about the SBP-box gene family in the loquat, and identified two EjSBP genes as candidates involved in carotenoid synthesis and accumulation during loquat fruit development. [ABSTRACT FROM AUTHOR]

Simple Summary: This study explores the role of circTPST2, a circular RNA, in the chemotherapy sensitivity of head and neck squamous cell carcinoma (HNSCC). Elevated levels of circTPST2 were observed in HNSCC tissues, and functional experiments revealed its inhibitory effect on cisplatin sensitivity in HNSCC cells. By interacting with miR-770-5p and the Nucleolin pathway, circTPST2 emerged as a crucial regulator influencing chemotherapy responsiveness. The findings suggest that circTPST2 may serve as a potential marker for chemotherapy regimen selection in HNSCC patients, providing insights to enhance treatment efficacy and patient outcomes. Chemoresistance poses a significant challenge in the treatment of advanced head and neck squamous cell cancer (HNSCC). The role and mechanism of circular RNAs (circRNAs) in HNSCC chemoresistance remain understudied. We conducted circRNA microarray analysis to identify differentially expressed circRNAs in HNSCC. The expression of circRNAs from the tyrosylprotein sulfotransferase 2 (TPST2) gene and miRNAs was evaluated through qPCR, while the circular structure of circTPST2 was verified using Sanger sequencing and RNase R. Through Western blotting, biotin-labeled RNA pulldown, RNA immunoprecipitation, mass spectrometry, and rescue experiments, we discovered miR-770-5p and nucleolin as downstream targets of circTPST2. Functional tests, including CCK8 assays and flow cytometry, assessed the chemoresistance ability of circTPST2, miR-770-5p, and Nucleolin. Additionally, FISH assays determined the subcellular localization of circTPST2, miR-770-5p, and Nucleolin. IHC staining was employed to detect circTPST2 and Nucleolin expression in HNSCC patients. circTPST2 expression was inversely correlated with cisplatin sensitivity in HNSCC cell lines. Remarkably, high circTPST2 expression correlated with lower overall survival rates in chemotherapeutic HNSCC patients. Mechanistically, circTPST2 reduced chemosensitivity through sponge-like adsorption of miR-770-5p and upregulation of the downstream protein Nucleolin in HNSCC cells. The TCGA database revealed improved prognosis for patients with low circTPST2 expression after chemotherapy. Moreover, analysis of HNSCC cohorts demonstrated better prognosis for patients with low Nucleolin protein expression after chemotherapy. We unveil circTPST2 as a circRNA associated with chemoresistance in HNSCC, suggesting its potential as a marker for selecting chemotherapy regimens in HNSCC patients. Further exploration of the downstream targets of circTPST2 advanced our understanding and improved treatment strategies for HNSCC. [ABSTRACT FROM AUTHOR]

Simple Summary: Bladder cancer (BLCA) is one of the most prevalent urinary cancers and novel non-invasive BLCA tumor biomarkers with high sensitivity and specificity are needed to support early diagnosis. Exosomal long non-coding RNAs (lncRNAs) are promising diagnostic biomarkers for BLCA. Our study suggests that the combined evaluation of lncRNA RMRP levels in urinary and plasma exosomes is an excellent potential non-invasive diagnostic biomarker for BLCA patients. Additionally, targeting the RMRP/miR-206/G6PD axis is a promising strategy for BLCA treatment. Bladder cancer (BLCA) is one of the cancers that is highly sensitive to specific non-invasive tumor biomarkers that facilitate early diagnosis. Exosome-derived long non-coding RNAs (lncRNAs) hold promise as diagnostic biomarkers for BLCA. In this study, we employed RNA-sequencing to compare the expression patterns of lncRNAs in urine exosomes from three BLCA patients and three healthy individuals. RMRP displayed the most significant differential expression. Elevated RMRP expression levels were observed in urinary and plasma exosomes from BLCA patients compared with those from healthy individuals. RMRP exhibited significant associations with certain BLCA patient clinicopathological features, including tumor stage, poor prognosis, and tumor grade. Combined diagnosis using RMRP in urine and plasma exosomes demonstrated a superior diagnostic performance with receiver operating characteristic curve analysis. RMRP was found to be related to BLCA tumor progression and the cell migration and invasion processes via the miR-206/G6PD axis both in vitro and in vivo. Mechanistically, RMRP serves as an miR-206 sponge, as suggested by dual-luciferase reporter assays and RNA immunoprecipitation. Our study suggests that the combined diagnosis of RMRP in urinary and plasma exosomes can serve as an excellent non-invasive diagnostic biomarker for BLCA patients. Additionally, targeting the RMRP/miR-206/G6PD axis holds promise as a therapeutic strategy for BLCA. [ABSTRACT FROM AUTHOR]

(1) Background: MicroRNAs (miRNAs) are involved in a variety of biological processes, such as cell proliferation, cell differentiation, and organ development. Recent studies have shown that plant miRNAs may enter the diet and play physiological and/or pathophysiological roles in human health and disease; however, little is known about plant miRNAs in chickens. (2) Methods: Here, we analyzed miRNA sequencing data, with the use of five Chinese native chicken breeds and six different tissues (heart, liver, spleen, lung, kidney, and leg muscle), and used Illumina sequencing to detect the expression of plant miRNAs in the pectoralis muscles at fourteen developmental stages of Tibetan chickens. (3) Results: The results showed that plant miRNAs are detectable in multiple tissues and organs in different chicken breeds. Surprisingly, we found that plant miRNAs, such as tae-miR2018, were detectable in free-range Tibetan chicken embryos at different stages. The results of gavage feeding experiments also showed that synthetic tae-miR2018 was detectable in caged Tibetan chickens after ingestion. The analysis of tae-miR2018 showed that its target genes were related to skeletal muscle organ development, regulation of mesodermal cell fate specification, growth factor activity, negative regulation of the cell cycle, and regulation of growth, indicating that exogenous miRNA may regulate the development of chicken embryos. Further cell cultures and exogenous miRNA uptake assay experiments showed that synthetic wheat miR2018 can be absorbed by chicken myoblasts. (4) Conclusions: Our study found that chickens can absorb and deposit plant miRNAs in various tissues and organs. The plant miRNAs detected in embryos may be involved in the development of chicken embryos. [ABSTRACT FROM AUTHOR]

Purpose: Due to poor prognosis and immunotherapy failure of skin cutaneous melanoma (SKCM), this study sought to find necroptosis-related biomarkers to predict prognosis and improve the situation with predicted immunotherapy drugs. Experimental Design: The Cancer Genome Atlas (TCGA) and The Genotype-Tissue Expression Program (GTEx) database were utilized to recognize the differential necroptosis-related genes (NRGs). Univariate Cox (uni-Cox) and least absolute shrinkage and selection operator (LASSO) Cox analysis were utilized for prognostic signature establishment. The signature was verified in the internal cohort. To assess the signature's prediction performance, the area under the curve (AUC) of receiver operating characteristic (ROC) curves, Kaplan-Meier (K-M) analyses, multivariate Cox (multi-Cox) regression, nomogram, and calibration curves were performed. The molecular and immunological aspects were also reviewed using single-sample gene set enrichment analysis (ssGSEA). Cluster analysis was performed to identify the different types of SKCM. Finally, the expression of the signature gene was verified by immunohistochemical staining. Results: On basis of the 67 NRGs, 4 necroptosis-related genes (FASLG, PLK1, EGFR, and TNFRSF21) were constructed to predict SKCM prognosis. The area's 1-, 3-, and 5-year OS under the AUC curve was 0.673, 0.649, and 0.677, respectively. High-risk individuals had significantly lower overall survival (OS) compared to low-risk patients. Immunological status and tumor cell infiltration in high-risk groups were significantly lower, indicating an immune system that was suppressed. In addition, hot and cold tumors could be obtained by cluster analysis, which is helpful for accurate treatment. Cluster 1 was considered a hot tumor and more susceptible to immunotherapy. Immunohistochemical results were consistent with positive and negative regulation of coefficients in signature. Conclusion: The results of this finding supported that NRGs could predict prognosis and help make a distinction between the cold and hot tumors for improving personalized therapy for SKCM. [ABSTRACT FROM AUTHOR]

Berberine hydrochloride (BBR) could inhibit the proliferation, migration, and invasion of various cancer cells. As the only enzyme for the de novo synthesis of ribonucleotides, RRM2 is closely related to the development of tumorigenesis. However, not much is currently known about the functional roles of RRM2 in breast cancer (BRCA), and whether BBR regulates the migration and invasion of BRCA cells by regulating the expression of RRM2 remains to be determined. We study the effects of BBR on BRCA cell proliferation in vitro and tumorigenesis in vivo by using colony formation assays, EdU assays, and xenograft models. Transcriptome sequencing, the random forest algorithm, and KEGG analysis were utilized to explore the therapeutic target genes and relative pathways. The expression of RRM2 in BRCA patients was analyzed with The Cancer Genome Atlas (TCGA) dataset, the GEPIA website tool, the Gene Expression Omnibus (GEO) database, and the UALCAN database. The survival probability of BRCA patients could be predicted by survival curve and nomogram analysis. Molecular docking was used to explore the affinity between BBR and potential targets. Gain- and loss-of-function methods were employed to explore the biological process in RRM2 participants. We comprehensively investigated the pharmacological characteristics of BBR on BRCA cell lines and discovered that BBR could inhibit the proliferation of BRCA cells in vitro and in vivo. Combining transcriptome sequencing and KEGG analysis, we found that BBR mainly affected the biological behavior of BRCA cells via HIF-1α and AMPK signal pathways. Additionally, by using bioinformatics and molecular docking, we demonstrated that RRM2 plays an oncogenic role in BRCA samples and that it acts as the hub gene of BBR on BRCA cells. Knockdown and overexpression studies indicated that RRM2 promoted BRCA cell migration as well as invasion in vitro by affecting the epithelial-to-mesenchymal transition (EMT). Our study demonstrated the significance of BBR regulating HIF-1α and AMPK signaling pathways in BRCA cells. Moreover, we revealed the carcinogenic role and potential mechanism of RRM2 as a core regulatory factor of BBR in BRCA in controlling BRCA invasion, migration, and EMT, suggesting that RRM2 may be a therapeutic target and prognostic biomarker for BRCA therapy. [ABSTRACT FROM AUTHOR]

We analyzed whole-genome bisulfite sequencing (WGBS) and RNA sequencing data of two young (1 year old) and two adult (9 years old) rhesus macaques (Macaca mulatta) to characterize the genomic DNA methylation profile of the thymus and explore the molecular mechanism of age-related changes in the thymus. Combining the two-omics data, we identified correlations between DNA methylation and gene expression and found that DNA methylation played an essential role in the functional changes of the aging thymus, especially in immunity and coagulation. The hypomethylation levels of C3 and C5AR2 and the hypermethylation level of C7 may lead to the high expressions of these genes in adult rhesus macaque thymuses, thus activating the classical complement pathway and the alternative pathway and enhancing their innate immune function. Adult thymuses had an enhanced coagulation pathway, which may have resulted from the hypomethylation and upregulated expressions of seven coagulation-promoting factor genes (F13A1, CLEC4D, CLEC4E, FCN3, PDGFRA, FGF2 and FGF7) and the hypomethylation and low expression of CPB2 to inhibit the degradation of blood clots. Furthermore, the functional decline in differentiation, activation and maturation of T cells in adult thymuses was also closely related to the changes in methylation levels and gene expression levels of T cell development genes (CD3G, GAD2, ADAMDEC1 and LCK) and the thymogenic hormone gene TMPO. A comparison of the age-related methylated genes among four mammal species revealed that most of the epigenetic clocks were species-specific. Furthermore, based on the genomic landscape of allele-specific DNA methylation, we identified several age-related clustered sequence-dependent allele-specific DNA methylated (cS-ASM) genes. Overall, these DNA methylation patterns may also help to assist with understanding the mechanisms of the aging thymus with the epigenome. [ABSTRACT FROM AUTHOR]

Circular RNAs (circRNAs) are involved in diverse biological processes. Avian influenza virus (AIV) can cross the species barrier to infect humans. Here, we employed RNA sequencing technology to profile circRNA, microRNA, and mRNA expression in human lung carcinoma cells in response to AIV or human influenza A virus (IAV) infection at viral replication. The analysis revealed that the expression of 475 common circRNAs were significantly regulated. The 381 and 1163 up-regulated circRNAs were induced by AIV at 8 and 16 h, respectively. Subsequently, gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses were also conducted for the AIV-specific up-regulated circRNAs. Moreover, the circRNAs were characterized, of which six were verified by quantitative real-time PCR. We further confirmed that expression of the selected circRNAs only increased following AIV infection. Knocking down the selected circRNAs promoted AIV proliferation, and overexpression of three of the candidate circRNAs restricted AIV replication and proliferation. We further analyzed that AIV-specific up-regulated circRNA mechanisms might function through the ceRNA network to affect the "Endocytosis" pathway and the "Cell cycle process". These data provide the first expression profile of AIV-specific up-regulated circRNAs and shed new light on the pathogenesis of AIV infection. Our findings also suggest that these circRNAs serve an important role in AIV infection. [ABSTRACT FROM AUTHOR]

Simple Summary: Autophagy is a cell survival and recycling mechanism which protects cancer cells upon therapeutic drug treatment. Here we investigated the impact of autophagy inhibition in a cancer of lymphoid origin, namely ALK-positive anaplastic large cell lymphoma (ALK + ALCL). We inhibited autophagy in two distinct cell subsets of ALK + ALCL, one of which we previously shown to possess more stem-like and tumorigenic properties. Our study found that blockage of autophagy in the stem-like subset resulted in marked drug-sensitization to crizotinib, a current therapeutic agent used to treat ALK + ALCL. We also found differential involvement of the Myconcogene in the autophagy process within the two subsets and identified its relative importance to the stem-like population. Our research suggests inhibition of autophagy alongside crizotinib preferentially targets stem-like cells, thus improving crizotinib therapy. Previously it was shown that autophagy contributes to crizotinib resistance in ALK-positive anaplastic large cell lymphoma (ALK + ALCL). We asked if autophagy is equally important in two distinct subsets of ALK + ALCL, namely Reporter Unresponsive (RU) and Reporter Responsive (RR), of which RR cells display stem-like properties. Autophagic flux was assessed with a fluorescence tagged LC3 reporter and immunoblots to detect endogenous LC3 alongside chloroquine, an autophagy inhibitor. The stem-like RR cells displayed significantly higher autophagic response upon crizotinib treatment. Their exaggerated autophagic response is cytoprotective against crizotinib, as inhibition of autophagy using chloroquine or shRNA against BECN1 or ATG7 led to a decrease in their viability. In contrast, autophagy inhibition in RU resulted in minimal changes. Since the differential protein expression of MYC is a regulator of the RU/RR dichotomy and is higher in RR cells, we asked if MYC regulates the autophagy-mediated cytoprotective effect. Inhibition of MYC in RR cells using shRNA significantly blunted crizotinib-induced autophagic response and effectively suppressed this cytoprotective effect. In conclusion, stem-like RR cells respond with rapid and intense autophagic flux which manifests with crizotinib resistance. For the first time, we have highlighted the direct role of MYC in regulating autophagy and its associated chemoresistance phenotype in ALK + ALCL stem-like cells. [ABSTRACT FROM AUTHOR]

Simple Summary: Recently, there has been increased recognition of an association between the bacterial microbiome and cancer. In this study, we characterized the non-HPV head and neck squamous cell carcinoma (HNSCC) microbiome. We found a significant enrichment of Fusobacterium, depletion of Streptococcus, and the microbial signatures of twelve bacterial genera distinguishing HNSCC. With increased Fusobacterium—in particular, F. nucleatum—in our HNSCC cohort and its known association with prognosis in colorectal cancers (CRC), we sought to further characterize the association between clinical outcomes and F. nucleatum, and the host interaction with F. nucleatum. We identified a higher abundance of F. nucleatum in non-smokers and an improved survival, in contrast to CRC. An integrative analysis also identified that the enrichment of F. nucleatum was associated with host gene promoter methylation, suggesting that the bacterial mircobiome status may have a potential role as a prognostic biomarker and be involved in the pathogenesis of HNSCC. The role of oral microbiota in head and neck squamous cell carcinoma (HNSCC) is poorly understood. Here we sought to evaluate the association of the bacterial microbiome with host gene methylation and patient outcomes, and to explore its potential as a biomarker for early detection or intervention. Here we performed 16S rRNA gene amplicon sequencing in sixty-eight HNSCC patients across both tissue and oral rinse samples to identify oral bacteria with differential abundance between HNSCC and controls. A subset of thirty-one pairs of HNSCC tumor tissues and the adjacent normal tissues were characterized for host gene methylation profile using bisulfite capture sequencing. We observed significant enrichments of Fusobacterium and Peptostreptococcus in HNSCC tumor tissues when compared to the adjacent normal tissues, and in HNSCC oral rinses when compared to healthy subjects, while ten other bacterial genera were largely depleted. These HNSCC-related bacteria were discriminative for HNSCC and controls with area under the receiver operating curves (AUCs) of 0.84 and 0.86 in tissue and oral rinse samples, respectively. Moreover, Fusobacterium nucleatum abundance in HNSCC cases was strongly associated with non-smokers, lower tumor stage, lower rate of recurrence, and improved disease-specific survival. An integrative analysis identified that enrichment of F. nucleatum was associated with host gene promoter methylation, including hypermethylation of tumor suppressor genes LXN and SMARCA2, for which gene expressions were downregulated in the HNSCC cohort from The Cancer Genome Atlas. In conclusion, we identified a taxonomically defined microbial consortium associated with HNSCC that may have clinical potential regarding biomarkers for early detection or intervention. Host–microbe interactions between F. nucleatum enrichment and clinical outcomes or host gene methylation imply a potential role of F. nucleatum as a pro-inflammatory driver in initiating HNSCC without traditional risk factors, which warrants further investigation for the underlying mechanisms. [ABSTRACT FROM AUTHOR]

Wintersweet (Chimonanthus praecox) is a well-known traditional fragrant plant and a winter-flowering deciduous shrub that originated in China. The five different developmental stages of wintersweet, namely, flower-bud period (FB), displayed petal stage (DP), open flower stage (OF), later blooming period (LB), and wilting period (WP) were studied using a scanning electron microscope (SEM) to determine the distribution characteristics of aroma-emitting nectaries. Results showed that the floral scent was probably emitted from nectaries distributed on the adaxial side of the innermost and middle petals, but almost none on the abaxial side. The nectaries in different developmental periods on the petals differ in numbers, sizes, and characteristics. Although the distribution of nectaries on different rounds of petals showed a diverse pattern at the same developmental periods, that of the nectaries on the same round of petals showed some of regularity. The nectary is concentrated on the adaxial side of the petals, especially in the region near the axis of the lower part of the petals. Based on transcriptional sequence and phylogenetic analysis, we report one nectary development related gene CpCRC (CRABS CLAW), and the other four YABBY family genes, CpFIL (FILAMENTOUS FLOWER), CpYABBY2, CpYABBY5-1, and CpYABBY5-2 in C. praecox (accession no. MH718960-MH718964). Quantitative RT-PCR (qRT-PCR) results showed that the expression characteristics of these YABBY family genes were similar to those of 11 floral scent genes, namely, CpSAMT, CpDMAPP, CpIPP, CpGPPS1, CpGPPS2, CpGPP, CpLIS, CpMYR1, CpFPPS, CpTER3, and CpTER5. The expression levels of these genes were generally higher in the lower part of the petals than in the upper halves in different rounds of petals, the highest being in the innermost petals, but the lowest in the outer petals. Relative expression level of CpFIL, CpCRC, CpYABBY5-1, and CpLIS in the innermost and middle petals in OF stages is significant higher than that of in outer petals, respectively. SEM and qRT-PCR results in C. praecox showed that floral scent emission is related to the distribution of nectaries. [ABSTRACT FROM AUTHOR]

The Asian tiger mosquito, Aedes albopictus, is a competent vector for the majority of arboviruses. The mosquito innate immune response is a primary determinant for arthropod-borne virus transmission, and the midgut is the first barrier to pathogen transmission. Mosquito antiviral immunity is primarily mediated by the small interfering RNA pathway. However, the roles that the P-element induced wimpy testis (PIWI)-interacting RNA (piRNA) pathway play in antiviral immunity in Ae. albopictus and its midgut still need further exploration. This study aimed to explore the profiles of both viral-derived and host-originated piRNAs in the whole body and midgut infected with Dengue virus 2 (DENV-2) in Ae. albopictus, and to elucidate gene expression profile differences of the PIWI protein family between adult females and their midguts. A deep sequencing-based method was used to identify and analyze small non-coding RNAs, especially the piRNA profiles in DENV-2-infected Ae. albopictus and its midgut. The top-ranked, differentially-expressed piRNAs were further validated using Stem-loop qRT-PCR. Bioinformatics analyses and reverse-transcription PCR (RT-PCR) methods were used to detect PIWI protein family members, and their expression profiles. DENV-2 derived piRNAs (vpiRNA, 24–30 nts) were observed in both infected Ae. albopictus and its midgut; however, only vpiRNA in the whole-body library had a weak preference for adenine at position 10 (10A) in the sense molecules as a feature of secondary piRNA. These vpiRNAs were not equally distributed, instead they were derived from a few specific regions of the genome, especially several hot spots, and displayed an obvious positive strand bias. We refer to the differentially expressed host piRNAs after DENV infection as virus-induced host endogenous piRNAs (vepiRNAs). However, we found that vepiRNAs were abundant in mosquito whole-body tissue, but deficient in the midgut. A total of eleven PIWI family genes were identified in Ae. albopictus; however, only AalPiwi5–7 and AalAgo3(1–2) were readily detected in the midgut. The characteristics of piRNAs in DENV-2-infected Ae. albopictus adult females were similar to those previously described for flavivirus infections but were not observed in the midgut. The reduced levels of vepiRNAs and incomplete expression of PIWI pathway genes in midgut samples from DENV-2-infected Ae. albopictus suggests that viral regulation of host piRNAs may not be an important factor in the midgut. [ABSTRACT FROM AUTHOR]