Identification and functional study of Fib-L, a major silk fibroin gene component in rice leaf folders.

The rice leaf folder, Cnaphalocrocis medinalis (Lepidoptera: Pyralidae), is a major migratory pest in rice agriculture. This pest is characterised by its larvae's ability to fold rice leaves using silk, a behaviour that culminates in the formation of a silken cocoon during the pupal stage. The fibroin light chain (CmFib-L) gene is crucial for silk production, yet its specific function in C. medinalis has reminded elusive. This study presents a comprehensive analysis of the CmFib-L gene, revealing its complete open reading frame (ORF) and expression patterns. Notably, the gene is highly expressed in the fifth-instar larvae and the silk gland, which are critical stages for silk production. Our experiments demonstrate that silencing the CmFib-L gene leads to a reduction in pupal weight, an extension of the pupal stage and a disorganised silk cocoon. Furthermore, the larval behaviour of leaf folding and spinning is significantly impaired when the expression of CmFib-L is downregulated. These findings not only show the importance of fibroin light chain in silk production but also reveal a new target gene to regulate and control the behaviour and development of C. medinalis.

Effect of Roasting on the Chemical Composition and Oxidative Stability of Tomato (Solanum lycopersicum L.) Seed Oil.

In this study, tomato seed (TS) samples were subjected to different roasting conditions (90-170 °C and 10-30 min) to compare their effects on the chemical composition and oxidative stability of tomato seed oil (TSO). Unroasted TS was considered as a control sample. Our results revealed that moderate roasting (130 °C/20 min) can significantly increase the content of linoleic acid (54.01-54.89%), linolenic acid (2.17-2.41%), phytosterols (2789.56-3037.31 mg/kg), squalene (5.06-13.10 mg/kg), total phenols (22.37-22.67 mg GAE/100 g), and other functional components (p < 0.05) in TSO, while the antioxidant activity (via DPPH, ABTS, and FRAP assays) also increased. In addition, the tocopherol content decreased significantly (758.53-729.50 mg/kg). Accelerated oxidation experiments showed that roasting (170 °C/30 min) increased the oxidative stability index (OSI) of TSO from 5.35 to 7.07 h (p < 0.05). Furthermore, roasting gradually increased the content of 5-hydroxymethylfurfural (HMF) (0-1.74 mg/kg), which indicates that the oxidative stability and the degree of the Maillard reaction increased upon roasting. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) showed that moderate roasting (130 °C/20 min) improved the chemical composition, antioxidant activity, and oxidative stability of TSO. Furthermore, this work provides a useful theoretical basis for the processing and wide application of TSO in the pharmaceutical and food industries.

CELSR3 is a prognostic marker in HNSCC and correlates with immune cell infiltration in the tumor microenvironment.

PURPOSE: To look at the diagnostic value of the CELSR receptor 3 (CELSR3) gene in head and neck squamous cell carcinoma (HNSCC) and its effect on tumor immune invasion, which is important for enhancing HNSCC treatment. METHODS: Several bioinformatics tools were employed to investigate CELSR3's putative oncogenic pathway in HNSCC, and datasets from The Tumor Genome Atlas (TCGA), Tumor Immune Estimation Resource (TIMER), Gene Expression Profile Interaction Analysis (GEPIA) and LinkedOmics were extracted and evaluated. CELSR3 has been linked to tumor immune cell infiltration, immunological checkpoints, and immune-related genes. CELSR3's putative roles were investigated using Gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and pathway enrichment analysis. The expression level of CELSR3 in HNSCC tissues and cells was detected by RT-qPCR. The effects of CELSR3 on proliferation of HNSCC cells were detected by CCK-8 assay. RESULTS: CELSR3 was shown to be expressed differently in different types of cancer and normal tissues. CELSR3 gene expression was linked to pN-stage and pM-stage. Patients with high CELSR3 expression also have a well prognosis. CELSR3 expression was found to be an independent predictive factor for HNSCC in both univariate and multivariate Cox regression analyses. We discovered the functional network of CELSR3 in HNSCC using GO and KEGG analysis. CELSR3 expression levels were found to be favorably associated with immune cell infiltration levels. Furthermore, CELSR3 expression levels were significantly correlated with the expression levels of many immune molecules, such as MHC genes, immune activation genes, chemokine receptors, and chemokines. CELSR3 is highly expressed in HNSCC tissues and cells. CELSR3 overexpression significantly inhibited the proliferation of HNSCC cells. CELSR3 expression may affect the immune microenvironment and, as a result, the prognosis of HNSCC. CONCLUSION: CELSR3 expression is elevated in HNSCC tumor tissues, and high CELSR3 expression is associated with well prognosis, which inhibited the proliferation of NHSCC cells. CELSR3 has the potential to influence tumor formation by controlling tumor-infiltrating cells in the tumor microenvironment (TME). As a result, CELSR3 may have diagnostic significance in HNSCC.

SLA2 is a prognostic marker in HNSCC and correlates with immune cell infiltration in the tumor microenvironment.

PURPOSE: To investigate Src-like adaptor 2 gene (SLA2) expression in head and neck squamous cell carcinoma (HNSCC), its potential prognostic value, and its effect on immune cell infiltration. METHODS: Through a variety of bioinformatics analyses, we extracted and analyzed data sets from the Cancer Genome Atlas (TCGA), Tumor Immune Estimation Resource (TIMER), and Gene Expression Profile Interaction Analysis (GEPIA) to analyze the correlation between SLA2 and the prognosis, immune checkpoint, tumor microenvironment (TME) and immune cell infiltration of HNSCC, and to explore its potential oncogenic mechanism. To further explore the potential role of SLA2 in HNSCC by Gene ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. RESULTS: SLA2 messenger ribonucleic acid (mRNA) levels were increased in HNSCC tumor tissues compared with normal tissues. In addition, we found that SLA2 may be an independent prognostic factor for HNSCC, and high SLA2 expression is associated with favorable prognosis in HNSCC. SLA2 expression was positively correlated with B cells, cluster of differentiation 8-positive T cells (CD8 + T cells), cluster of differentiation 4-positive T cells (CD4 + T cells), macrophages, neutrophil and dendritic cells infiltration. SLA2 has also been shown to co-express immune-related genes and immune checkpoints. Significant GO term analysis by Gene Set Enrichment Analysis (GSEA) indicated that genes correlated with SLA2 were located mainly in the side of membrane, receptor complex, secretory granule membrane, endocytic vesicle, membrane region, and endosome membrane, where they were involved in leukocyte cell-cell adhesion, response to interferon-gamma, and regulation of immune effector process. These related genes also served as antigen binding, cytokine receptor activity, phosphatidylinositol 3-kinase activity, peptide receptor activity, Src homology domain 3 (SH3) domain binding, and cytokine receptor binding. KEGG pathway analysis demonstrated that these genes related to SLA2 were mainly enriched in signal pathways, such as hematopoietic cell lineage, cell adhesion molecules (CAMs), natural killer cell mediated cytotoxicity, measles, and chemokine signaling pathway. CONCLUSIONS: SLA2 is increased in HNSCC, and high SLA2 expression is associated with favorable prognosis. SLA2 may affect tumor development by regulating tumor infiltrating cells in TME. SLA2 may be a potential target for immunotherapy.

Prediction of network pharmacology, molecular docking-based strategy, and vitro assays to determine potential pharmacological mechanism of Dioscoreae bulbiferae and Bruceae fructus against laryngocarcinoma.

BACKGROUND: Based on network pharmacology, molecular docking, and vitro assays, investigate the probable pharmacological mechanism of Dioscoreae bulbiferae and Bruceae fructus in the treatment of laryngocarcinoma. METHODS: The active components and targets of Dioscoreae bulbiferae and Bruceae fructus were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform database. Targets linked with laryngocarcinoma were gathered from the GeneCards, DisGeNET, and DrugBank databases. The String database was utilized to build a protein-protein interaction network of common medication and illness targets, after which the core targets were filtered out. The Metascape database served for gene ontology enrichment and Kyoto encyclopedia of genes and genomes pathway analysis of common targets. AutoDock then performed molecular docking between the essential component and the vital target. To investigate the biological effects of diosbulbin B, we assessed the viability of laryngocarcinoma cells after diosbulbin B therapy using the Mahalanobis Taguchi system technique. Following that, we looked at how diosbulbin B affected colony formation after 14 days of culture of treated cells. Flow cytometry was utilized to detect apoptosis in order to examine the influence of diosbulbin B on laryngocarcinoma cell apoptosis. RESULTS: According to a study of the literature, the fundamental components of Dioscoreae bulbiferae and Bruceae fructus in the treatment of laryngocarcinoma include brusatol and diosbulbin B, which may operate on core targets such as cyclin D1, Cyclin Dependent Kinase Inhibitor 1A, and E2F Transcription Factor 1. The significant pathways discovered using Kyoto encyclopedia of genes and genomes enrichment analysis were the phosphoinositide 3-kinase-protein kinase B signaling route, the tumor necrosis factor signaling pathway, and so on. These pathways primarily influence the development and prognosis of laryngeal cancer by controlling cell growth, cell proliferation, angiogenesis, tumorigenesis, and metastasis. The molecular docking studies revealed that the affinity between the heart and crucial targets was robust. The results of vitro assays indicate that diosbulbin B suppressed Hep-2 cell activity in a concentration-dependent manner. Besides, diosbulbin B has powerful antiproliferative properties in Hep-2 cells. Flow cytometry results showed that diosbulbin B promoted laryngocarcinoma cell apoptosis in a concentration-dependent manner. CONCLUSION: The article delivered a preliminary discussion of the probable mechanism of Dioscoreae bulbiferae and Bruceae fructus in the treatment of laryngocarcinoma, which can serve as a theoretical basis and evidence for subsequent experimental investigation.

Integrating GEO, network pharmacology, and in vitro assays to explore the pharmacological mechanism of Bruceae Fructus against laryngeal cancer.

The goal of this study is to look into the pharmacological mechanism of Bruceae Fructus in conjunction with GEO, network pharmacology, and in vitro assays for the treatment of laryngeal cancer to provide theoretical support for its therapeutic use. The active components and matching targets of Bruceae Fructus were retrieved from the TCMSP database, while genes linked with laryngeal cancer were obtained from the GEO, GeneCards, DisGeNET, and DrugBank databases. Besides, the components and targets were supplemented by literatures in PubMed database. Cytoscape software was used to create the active ingredients-target network diagram. The String database was used to build the PPI network. Following that, the core targets were subjected to GO enrichment and KEGG pathway analysis using the DAVID database. Finally, AutoDock was used to perform molecular docking between the core components and the core targets. To investigate the biological effects of beta-sitosterol, the viability of laryngeal cancer cells was assessed after beta-sitosterol therapy using the MTS technique. Following that, how beta-sitosterol affected colony formation after 14 days of culture of treated cells was researched. Flow cytometry was utilized to detect apoptosis to examine the influence of beta-sitosterol on laryngeal cancer cell apoptosis, and then detected mRNA and protein expression levels of 10 key genes by RT-qPCR and Western Blot assay. There were 1258 laryngeal cancer-related genes and 15 Bruceae Fructus components, with beta-sitosterol and luteolin serving as key components. Bruceae Fructus' primary targets against laryngeal cancer were IL6, JUN, TNF, IL2, IL4, IFNG, RELA, TP53, CDKN1A, and AKT1. GO enrichment yielded 41 CC, 78 MF, and 383 BP. Platinum drug resistance, the PI3K-Akt signaling pathway, the p53 signaling pathway, apoptosis, the HIF-1 signaling pathway, and 147 additional pathways have been added to KEGG. The results of molecular docking revealed that the core components had a high affinity for the core target. The results of the cell experiment indicate that beta-sitosterol suppressed Hep-2 cell activity in a concentration-dependent manner. Besides, beta-sitosterol has powerful antiproliferative properties in Hep-2 cells. Flow cytometry results showed that beta-sitosterol promoted laryngeal cancer cell apoptosis in a concentration-dependent manner. The results of RT-qPCR and Western Blot assay showed that the mRNA and protein expression levels of TP53, JUN, TNF-α, CDKN1A, and IL-2 were significantly up-regulated after beta-sitosterol treatment, while the mRNA and protein expression levels of RELA, AKT1, IL-6, IFNG, and IL-4 were significantly down-regulated. This study integrating GEO, network pharmacology, and in vitro assays investigated the probable mechanism of Bruceae Fructus' anti-laryngeal cancer activity, which can give a theoretical foundation for additional future animal experiments.

Generating waxy rice starch with target type of amylopectin fine structure and gelatinization temperature by waxy gene editing.

Waxy rice, which lacks amylose, is an important variant in rice, and its starches have been widely used. New waxy rice varieties generated via the CRISPR/Cas9 gene-editing system is described. Herein, four waxy rice starches with different physicochemical properties were successfully obtained by the CRISPR/Cas9 editing Waxy (Wx) gene. The results showed that the amylose content (AC) of wx mutant starches ranged from 0.26 to 1.78 %, and CZBwx1 starches had the best gel consistency and highest water solubility among all wx mutants. Mutations of Wxb allele produced more short-chains (degree of chain polymerization (DP) 6-11), and less medium- and long-chains (DP12-70) than that of Wxa, while the AC of Wxa allele mutants was higher than the mutations of Wxb allele. The gelatinization temperature (GT) of wxa mutant starches was higher than that of wxb mutant starches. Moreover, we found that the GT and amylopectin fine structure type of waxy rice starch did not change after Wx gene editing. Based on these findings, it is possible to produce waxy rice starch with different physicochemical properties, containing target GT and chain length distributions of amylopectin.

Prediction of network pharmacology and molecular docking-based strategy to determine potential pharmacological mechanism of Liuwei Dihuang pill against tinnitus.

BACKGROUND: Liuwei Dihuang Pill is widely used to treat tinnitus in China. However, the underlying mechanism of Liuwei Dihuang Pill in treating tinnitus still remains unclear. OBJECTIVE: To explore the potential pharmacological mechanism of Liuwei Dihuang Pill in the treatment of tinnitus based on network pharmacology and molecular docking. METHODS: The active components of the Liuwei Dihuang Pill were obtained from the traditional Chinese medicine systems pharmacology database and analysis platform (TCMSP) database. Cytoscape software was used to draw the active component-target network diagram of Liuwei Dihuang Pill, and obtain the core components. Then the corresponding targets were also obtained from the TCMSP database. Targets related to tinnitus were obtained from the GeneCards, DisGeNET, TTD and DrugBank databases. The String database was used to construct protein-protein interaction (PPI) network of common targets of drugs and diseases, then the core targets were screened out. The Annotation, Visualization and Integrated Discovery (DAVID) database was used for gene ontology (GO) enrichment and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis of common targets. Finally, the molecular docking between the core component and the core target was carried out by AutoDock. RESULTS: The core components of Liuwei Dihuang Pill in the treatment of tinnitus including quercetin, stigmasterol, kaempferol, ß-sitosterol, tetrahydroalstonine, which may act on core targets such as STAT3, transcription factor AP-1 (JUN), tumor necrosis factor (TNF), interleukin-6 and MAPK3. HIF-1 signaling pathway, Influenza A, P53 signaling pathway, and Toll-like receptor signaling pathway play a role in anti-inflammatory, improving microcirculation in the blood-labyrinth barrier, increasing cochlear blood flow, and preventing hair cell damage. The molecular docking results showed that the affinity between core components and core targets was good. CONCLUSION: The potential mechanism of Liuwei Dihuang Pill in the treatment of tinnitus was preliminarily discussed in this study, which may provide a theoretical basis and evidence for further experimental research.

Damage of brown planthopper (BPH) Nilaparvata lugens and rice leaf folder (LF) Cnaphalocrocis medinalis in parent plants lead to distinct resistance in ratoon rice.

Herbivore-induced defense responses are often specific, whereas plants could induce distinct defense responses corresponding to infestation by different herbivorous insects. Brown plant hopper (BPH) Nilaparvata lugens, a phloem-feeding insect, and rice leaf folder (LF) Cnaphalocrocis medinalis, a chewing insect, are both specialist herbivores on rice. To characterize the distinct resistance primed by prior damage to these two specialist herbivores, we challenged rice plants with two herbivores during vegetative growth of parent plants and assessed plant resistance in subsequent ratoons. Here, we show that LF and BPH induce different suites of defense responses in parent rice plants, LF induced higher level of JA accumulation and OsAOS, OsCOI1 transcripts, while BPH induced higher accumulation of SA and OsPAL1 transcripts. Moreover, an apparent loss of LF resistance was observed in OsAOS, OsCOI1 RNAi lines. Ratoon plants generated from parents receiving prior LF infestation exhibited higher jasmonic acid (JA) levels and elevated levels of transcripts of defense-related genes associated with JA signaling, while ratoon generated from parents receiving prior BPH infestation exhibited higher salicylic acid (SA) levels and elevated levels of transcripts of defense-related genes associated with SA signaling. Moreover, previous LF infestation obviously elevated ratoons resistance to LF, while previous infestation by BPH led to enhanced resistance in ratoons to BPH. Pre-priming of ratoons defense to LF was significantly reduced in OsAOS and OsCOI1 RNAi plant, but silencing OsAOS and OsCOI1 did not attenuate ratoons resistance to BPH. These results suggest that infestation of two specialist herbivores with different feeding styles in parent crop led to distinct defense responses in subsequent rations, and the acquired resistance to LF in ratoons is associated with priming of jasmonic acid-dependent defense responses.

Seed priming with calcium chloride enhances wheat resistance against wheat aphid Schizaphis graminum Rondani.

BACKGROUND: Calcium is an essential macronutrient for plant growth. Although it has been shown that exogenous Ca application can increase plant resistance to abiotic stress, little is known about its potential to enhance plant tolerance to biotic stress. Here, we investigated whether pretreatment of wheat (Triticum aestivum L.) seeds with calcium chloride (CaCl2 ) improves plant resistance against wheat aphid (Schizaphis graminum Rondani). The developmental time, population size, feeding behavior of aphids on plants grown from CaCl2 - and water-pretreated seeds, and plant defense responses to aphid attack were investigated. RESULTS: Seed pretreatment with CaCl2 extended aphid development time and reduced aphid population size and feeding efficiency. In addition, the pretreatment significantly increased the concentration of Ca2+ in wheat leaves, and upregulated expression levels of TaCaM genes and callose synthase genes (TaGSL2, TaGSL8, TaGSL10, TaGSL12, TaGSL19, TaGSL22 and TaGSL23). Callose concentration in the leaves of plants grown from CaCl2 -pretreated seeds increased significantly upon aphid attack. Further, callose deposition was observed mainly in the phloem. CONCLUSION: These results suggest that seed pretreatment with CaCl2 primes the plant response against wheat aphid attack, leading to modulation of callose deposition in the phloem in response to aphid attack. © 2021 Society of Chemical Industry.