Integrated Transcriptome Analysis Research Articles

Integrative transcriptome and metabolome analysis reveals the mechanism of fulvic acid alleviating drought stress in oat

Drought stress inhibits oat growth and yield. The application of fulvic acid (FA) can improve the drought resistance of oats, but the corresponding molecular mechanism of FA-mediated drought resistance remains unclear. Here, we studied the effects of FA on the drought tolerance of oat leaves through physiological, transcriptomic, and metabolomics analyses, and identified FA-induced genes and metabolites related to drought tolerance. Physiological analysis showed that under drought stress, FA increased the relative water and chlorophyll contents of oat leaves, enhanced the activity of antioxidant enzymes (SOD, POD, PAL, CAT and 4CL), inhibited the accumulation of malondialdehyde (MDA), hydrogen peroxide (H2O2) and dehydroascorbic acid (DHA), reduced the degree of oxidative damage in oat leaves, improved the drought resistance of oats, and promoted the growth of oat plants. Transcriptome and metabolite analyses revealed 652 differentially expressed genes (DEGs) and 571 differentially expressed metabolites (DEMs) in FA-treated oat leaves under drought stress. These DEGs and DEMs are involved in a variety of biological processes, such as phenylspropanoid biosynthesis and glutathione metabolism pathways. Additionally, FA may be involved in regulating the role of DEGs and DEMs in phenylpropanoid biosynthesis and glutathione metabolism under drought stress. In conclusion, our results suggest that FA promotes oat growth under drought stress by attenuating membrane lipid peroxidation and regulating the antioxidant system, phenylpropanoid biosynthesis, and glutathione metabolism pathways in oat leaves. This study provides new insights into the complex mechanisms by which FA improves drought tolerance in crops.

Methyl jasmonate induced tolerance effect of Pinus koraiensis to Bursaphelenchus xylophilus.

Methyl jasmonate (MeJA) can affect the balance of hormones and regulate the disease resistance of plants. Exploring the application and mechanism of MeJA in inducing the tolerance of Pinus koraiensis to pine wood nematode (PWN) infection is of great significance for developing new strategies for pine wilt disease control. Different concentrations (0.1, 1, 5 and 10 mm) of MeJA treatment groups showed differences in relative tolerance index and relative anti-nematode index of P. koraiensis seedlings to PWN infection. The treatment of 5 mm MeJA solution induced the best tolerance effect, followed by the 1 mm MeJA solution. Transcriptome analysis indicated that many plant defense-related genes upregulated after treatment with 1, 5 and 10 mm MeJA solutions. Among them, genes such as jasmonate ZIM domain-containing protein, phenylalanine ammonia-lyase and peroxidase also continuously upregulated after PWN infection. Metabolome analysis indicated that jasmonic acid (JA) was significantly increased at 7 days postinoculation with PWN, and after treatment with both 1 and 5 mm MeJA solutions. Integrated analysis of transcriptome and metabolome indicated that differences in JA accumulation might lead to ubiquitin-mediated proteolysis, and expression changes in trans-caffeic acid and trans-cinnamic acid-related genes, leading to the abundance differences of these two metabolisms and the formation of multiple lignin and glucosides. MeJA treatment could activate the expression of defense-related genes that correlated with JA, regulate the abundance of defense-related secondary metabolites, and improve the tolerance of P. koraiensis seedlings to PWN infection. © 2024 Society of Chemical Industry.

Integrative analysis of transcriptome and metabolome reveals how ethylene increases natural rubber yield in Hevea brasiliensis.

Hevea brasiliensis is an important cash crop with the product named natural rubber (NR) for markets. Ethylene (ET) is the most effective yield stimulant in NR production but the molecular mechanism remains incomplete. Here, latex properties analysis, transcriptome analysis, and metabolic profiling were performed to investigate the mechanism of NR yield increase in four consecutive tappings after ET stimulation. The results revealed that sucrose and inorganic phosphate content correlated positively with dry-rubber yield and were induced upon ET stimulation. Stimulation with ET also led to significant changes in gene expression and metabolite content. Genes involved in phytohormone biosynthesis and general signal transduction as well as 51 transcription factors potentially involved in the ET response were also identified. Additionally, KEGG annotation of differentially accumulated metabolites suggested that metabolites involved in secondary metabolites, amino-acid biosynthesis, ABC transporters, and galactose metabolism were accumulated in response to ET. Integrative analysis of the data collected by transcriptomics and metabolomics identified those differentially expressed genes and differentially accumulated metabolites are mainly involved in amino-acid biosynthesis and carbohydrate metabolism. Correlation analysis of genes and metabolites showed a strong correlation between amino-acid biosynthesis during ET stimulation. These findings provide new insights into the molecular mechanism underlying the ET-induced increase in rubber yield and further our understanding of the regulatory mechanism of ethylene signaling in rubber biosynthesis.

Validation studies and multi-omics analysis of Zhx2 as a candidate quantitative trait gene underlying brain oxycodone metabolite (oxymorphone) levels and behavior.

Sensitivity to the subjective reinforcing properties of opioids has a genetic component and can predict addiction liability of opioid compounds. We previously identified Zhx2 as a candidate gene underlying increased brain concentration of the oxycodone ( OXY ) metabolite oxymorphone ( OMOR ) in BALB/cJ ( J ) versus BALB/cByJ ( By ) females that could increase OXY state-dependent reward. A large structural intronic variant is associated with a robust reduction of Zhx2 expression in J mice, which we hypothesized enhances OMOR levels and OXY addiction-like behaviors. We tested this hypothesis by restoring the Zhx2 loss-of-function in Js ( MVKO ) and modeling the loss-of-function variant through knocking out the Zhx2 coding exon ( E3KO ) in Bys and assessing brain OXY metabolite levels and behavior. Consistent with our hypothesis, Zhx2 E3KO females showed an increase in brain OMOR levels and OXY-induced locomotor activity. However, contrary to our hypothesis, state-dependent expression of OXY-CPP was decreased in E3KO females and increased in E3KO males. We also overexpressed Zhx2 in the livers and brains of Js and observed Zhx2 overexpression in select brain regions that was associated with reduced OXY state-dependent learning. Integrative transcriptomic and proteomic analysis of E3KO mice identified astrocyte function, cell adhesion, extracellular matrix properties, and endothelial cell functions as pathways influencing brain OXY metabolite concentration and behavior. These results support Zhx2 as a quantitative trait gene underlying brain OMOR concentration that is associated with changes in OXY behavior and implicate potential quantitative trait mechanisms that together inform our overall understanding of Zhx2 in brain function.

Quercetin relieves compression-induced cell death and lumbar disc degeneration by stabilizing HIF1A protein

BackgroundLumbar disc degeneration (LDD) is a prevalent condition characterized by the decreased viability and functional impairment of nucleus pulposus mesenchymal stem cells (NPMSCs). Shaoyao-Gancao decoction (SGD), a traditional Chinese medicine formula, has been used to treat LDD, but its active components and mechanisms are unclear. MethodsAn integrative network pharmacology and transcriptome analysis were conducted to identify bioactive compounds in SGD that could target LDD. NPMSCs were cultured under mechanical compression as a cellular model of LDD. A rat model of annulus fibrosus needle-puncture was established to induce intervertebral disc degeneration. The effects of quercetin, a predicted active component, on alleviating compression-induced NPMSC death and LDD were evaluated in vitro and in vivo. ResultsThe analysis identified hypoxia-inducible factor 1-alpha (HIF1A) as a potential target of quercetin in LDD. HIF1A was upregulated in degenerated human disc samples and compression-treated NPMSCs. Quercetin treatment alleviated compression-induced oxidative stress, apoptosis, and loss of viability in NPMSCs by stabilizing HIF1A. The protective effects of quercetin were abrogated by HIF1A inhibition. In the rat model, quercetin ameliorated intervertebral disc degeneration. ConclusionOur study identified HIF1A as a protective factor against compression-induced cell death in NPMSCs. Quercetin, a bioactive compound found in the traditional Chinese medicine formula SGD, improved the survival of NPMSCs and alleviated LDD progression by stabilizing HIF1A. Targeting the HIF1A pathway through natural compounds like quercetin could represent a promising strategy for the clinical management of LDD and potentially other degenerative disc diseases.

Integrating transcriptomics, metabolomics, and network pharmacology to investigate multi-target effects of Sporoderm-broken spores of Ganoderma lucidum on improving HFD-induced diabetic nephropathy rats

Diabetes mellitus (DM) is a major metabolic disease endangering global health, with diabetic nephropathy (DN) as a primary complication lacking curative therapy. Sporoderm-broken spores of Ganoderma lucidum (GLP), an herbal medicine, has been used for the treatment of metabolic disorders. In this study, DN was induced in Sprague-Dawley rats using streptozotocin (STZ) and a high-fat diet (HFD), and the protective mechanisms of GLP were investigated through transcriptomic, metabolomic, and network pharmacology analyses. Our results demonstrated that GLP intervention ameliorated renal damage and inflammation levels in DN rats. Integrative metabolomic and transcriptomic analysis revealed that GLP treatment modulated glucose and cellular energy metabolisms by regulating relevant genes. GLP significantly suppressed the inflammations by impacting glucose and energy metabolism-related gene expression (Igfbp1 and Angptl4) and enhanced metabolic biomarkers of 4-Aminocatechol. In addition, network pharmacology analysis further indicated that GLP may efficiently alleviate DN via immune-related pathways. In conclusion, this study provides supportive evidence of the anti-inflammatory effects of GLP supplements, highlighting their potential for promising clinical applications in treating diabetic nephropathy.

Integrative analysis of transcriptome and metabolome reveals the heterogeneity of ovarian follicles between high-altitude Yak (Bos grunniens) and Low-altitude cattle (Bos taurus)

Differences including reproductive efficiency have been proved existence between high-altitude yak (Bos grunniens) and low-altitude cattle (Bos taurus), however, the discrepancies of ovarian follicles across them are still elusive. In the present study, RNA-seq results exhibited an obvious separation of yak and cattle granulosa cells (GCs), and 2329 differently expressed genes (DEGs) was obtained, with 1297 upregulated and 1032 downregulated DEGs in yaks. Advanced analysis revealed that DEGs were mainly enriched in cellular structure, DNA packaging and remodeling (GO and GSEA), hypoxia adaptability, energy metabolism (KEGG and GSEA), and epigenetic modification of histones (Reactome and GSEA). Metabolite profiles of yak and cattle follicular fluid (FF) were depicted by LC-MS/MS non-target metabolomics, and 350 different metabolites (DMs) were acquired, with 242 upregulated and 108 downregulated DMs in yaks. The KEGG pathway enrichment results demonstrated that DMs were primarily enriched in metabolism including lipid, carbohydrate and nucleic acid metabolism. Finally, twenty-nine pathways were significant enriched via integrative analysis, most of which were associated with metabolism including carbohydrate metabolism (e.g. Glycolysis / Gluconeogenesis) and lipid metabolism (e.g. Sphingolipid metabolism). Of note, the key hypoxia adaptation pathway, HIF-1 signaling pathway, was predominant enriched. Overall, we ascertained that the heterogeneity in ovarian follicles of yaks and cattle was existence, which were mainly reflected on hypoxia adaptation and energy metabolism. These results implied that the above processes were evolved specifically for the normal development of yak follicles under plateau atmosphere.

CRISPR screen of venetoclax response-associated genes identifies transcription factor ZNF740 as a key functional regulator

BCL-2 inhibitors such as venetoclax offer therapeutic promise in acute myeloid leukemia (AML) and other cancers, but drug resistance poses a significant challenge. It is crucial to understand the mechanisms that regulate venetoclax response. While correlative studies have identified numerous genes linked to venetoclax sensitivity, their direct impact on the drug response remains unclear. In this study, we targeted around 1400 genes upregulated in venetoclax-sensitive primary AML samples and carried out a CRISPR knockout screen to evaluate their direct effects on venetoclax response. Our screen identified the transcription factor ZNF740 as a critical regulator, with its expression consistently predicting venetoclax sensitivity across subtypes of the FAB classification. ZNF740 depletion leads to increased resistance to ventoclax, while its overexpression enhances sensitivity to the drug. Mechanistically, our integrative transcriptomic and genomic analysis identifies NOXA as a direct target of ZNF740, which negatively regulates MCL-1 protein stability. Loss of ZNF740 downregulates NOXA and increases the steady state protein levels of MCL-1 in AML cells. Restoring NOXA expression in ZNF740-depleted cells re-sensitizes AML cells to venetoclax treatment. Furthermore, we demonstrated that dual targeting of MCL-1 and BCL-2 effectively treats ZNF740-deficient AML in vivo. Together, our work systematically elucidates the causal relationship between venetoclax response signature genes and establishes ZNF740 as a novel transcription factor regulating venetoclax sensitivity.

Gut dysbiosis of Rana zhenhaiensis tadpoles after lead (Pb) exposure based on integrated analysis of microbiota and gut transcriptome

Lead (Pb) is a ubiquitously detected heavy metal pollutant in aquatic ecosystems. Previous studies focused mainly on the response of gut microbiota to Pb stress, with less emphasis on gene expression in intestine, thereby limiting the information about impacts of Pb on intestinal homeostasis in amphibians. Here, microbial community and transcriptional response of intestines in Rana zhenhaiensis tadpoles to Pb exposure were evaluated. Our results showed that 10 μg/L Pb significantly decreased bacterial diversity compared to the controls by the Simpson index. Additionally, 1000 μg/L Pb exposure resulted in a significant reduction in the abundance of Fusobacteriota phylum and Cetobacterium genus but a significant expansion in Hafnia-Obesumbacterium genus. Moreover, transcriptome analysis revealed that about 90 % of the DEGs (8458 out of 9450 DEGs) were down-regulated in 1000 μg/L Pb group, mainly including genes annotated with biological functions in fatty acid degradation, and oxidative phosphorylation, while up-regulated DEGs involved in metabolism of xenobiotics by cytochrome P450. The expression of Gsto1, Gsta5, Gstt4, and Nadph showed strong correlation with the abundance of genera Serratia, Lactococcus, and Hafnia-Obesumbacterium. The findings of this study provide important insights into understanding the influence of Pb on intestinal homeostasis in amphibians.

Research Note: Integrative analysis of transcriptome and gut microbiome reveals foie gras capacity difference between cage and floor rearing systems

To explore the differences in foie gras performance between geese raised in cages and on the ground, we conducted an integrative analysis of liver transcriptome and gut microbial metagenomes. The results showed extremely significant differences in the liver weight (P < 0.01) and liver lipid accumulation of FRS and CRS groups. The levels of triglyceride (TG), high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) of CRS were significantly higher than those of FRS (P < 0.05). Transcriptome analysis showed that 3,917 upregulated and 1,395 downregulated genes were identified, and lipid metabolism pathway and fatty acid metabolism were significantly enriched. Analysis of cecum microbiota revealed that several inflammation-related bacteria (including Gallibacterium, Escherichia-Shigella, Desulfovibrio, Alistipes, and Fournierella) were enriched in CRS, while beneficial bacteria (including Lactobacillus, Limosilactobacillus, and Ligilactobacillus) were significantly enriched in FRS. In conclusion, CRS was better than FRS in foie gras production, which was more conducive to lipid deposition in the goose liver.

Exploring the Molecular Mechanism of Skeletal Muscle Development in Ningxiang Pig by Weighted Gene Co-Expression Network Analysis.

With the continuous improvement in living standards, people's demand for high-quality meat is increasing. Ningxiang pig has delicious meat of high nutritional value, and is loved by consumers. However, its slow growth and low meat yield seriously restrict its efficient utilization. Gene expression is the internal driving force of life activities, so in order to fundamentally improve its growth rate, it is key to explore the molecular mechanism of skeletal muscle development in Ningxiang pigs. In this paper, Ningxiang boars were selected in four growth stages (30 days: weaning period, 90 days: nursing period, 150 days: early fattening period, and 210 days: late fattening period), and the longissimus dorsi (LD) muscle was taken from three boars in each stage. The fatty acid content, amino acid content, muscle fiber diameter density and type of LD were detected by gas chromatography, acidolysis, hematoxylin eosin (HE) staining and immunofluorescence (IF) staining. After transcription sequencing, weighted gene co-expression network analysis (WGCNA) combined with the phenotype of the LD was used to explore the key genes and signaling pathways affecting muscle development. The results showed that 10 modules were identified by WGCNA, including 5 modules related to muscle development stage, module characteristics of muscle fiber density, 5 modules characteristic of muscle fiber diameter, and a module characteristic of palmitoleic acid (C16:1) and linoleic acid (C18:2n6C). Gene ontology (GO) enrichment analysis found that 52 transcripts relating to muscle development were enriched in these modules, including 44 known genes and 8 novel genes. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that these genes were enriched in the auxin, estrogen and cyclic guanosine monophosphate-protein kinase G (cGMP-PKG) pathways. Twelve of these genes were transcription factors, there were interactions among 20 genes, and the interactions among 11 proteins in human, pig and mouse were stable. To sum up, through the integrated analysis of phenotype and transcriptome, this paper analyzed the key genes and possible regulatory networks of skeletal muscle development in Ningxiang pigs at various stages, to provide a reference for the in-depth study of skeletal muscle development.

Inhibition of ALOX12-12-HETE Alleviates Lung Ischemia-Reperfusion Injury by Reducing Endothelial Ferroptosis-Mediated Neutrophil Extracellular Trap Formation.

Lung ischemia-reperfusion injury (IRI) stands as the primary culprit behind primary graft dysfunction (PGD) after lung transplantation, yet viable therapeutic options are lacking. In the present study, we used a murine hilar clamp (1 h) and reperfusion (3 h) model to study IRI. The left lung tissues were harvested for metabolomics, transcriptomics, and single-cell RNA sequencing. Metabolomics of plasma from human lung transplantation recipients was also performed. Lung histology, pulmonary function, pulmonary edema, and survival analysis were measured in mice. Integrative analysis of metabolomics and transcriptomics revealed a marked up-regulation of arachidonate 12-lipoxygenase (ALOX12) and its metabolite 12-hydroxyeicosatetraenoic acid (12-HETE), which played a pivotal role in promoting ferroptosis and neutrophil extracellular trap (NET) formation during lung IRI. Additionally, single-cell RNA sequencing revealed that ferroptosis predominantly occurred in pulmonary endothelial cells. Importantly, Alox12-knockout (KO) mice exhibited a notable decrease in ferroptosis, NET formation, and tissue injury. To investigate the interplay between endothelial ferroptosis and NET formation, a hypoxia/reoxygenation (HR) cell model using 2 human endothelial cell lines was established. By incubating conditioned medium from HR cell model with neutrophils, we found that the liberation of high mobility group box1 (HMGB1) from endothelial cells undergoing ferroptosis facilitated the formation of NETs by activating the TLR4/MYD88 pathway. Last, the administration of ML355, a targeted inhibitor of Alox12, mitigated lung IRI in both murine hilar clamp/reperfusion and rat left lung transplant models. Collectively, our study indicates ALOX12 as a promising therapeutic strategy for lung IRI.

Integrated analysis of transcriptome, proteome and non-targeted metabolome for exploring the mechanism of selenium biotransformation in Pichia kudriavzevii

Pichia kudriavzevii could resist high concentrations of sodium selenite and efficiently biotransform over 80% of the absorbed selenium. However, the specific molecular mechanisms underlying this biotransformation remain unclear. This study aimed to investigate the mechanism of selenium biotransformation focusing on selenium transport, detoxification, reduction, and the formation of organic selenium compounds through a comprehensive approach combining transcriptomic, proteomic, and non-targeted metabolomic analyze. Results showed that the JEN1 gene was responsible for selenium transport, while the upregulation of cysJ, CYSK, TRXB1 and sat genes as well as the involvement of glutathione, are crucial for the accumulation of organic selenium. Furthermore, GSTY2, TRXB1 and GPX3 were pivotal genes in Se(IV) reduction and their corresponding proteins were key proteins. Additionally, the enhanced expression of genes related to antioxidant enzymes (GSTY2, TRXB1, and GPX3), ABC transports, heat shock proteins and antioxidant enzymes proteins (GSTY2, TRXB1, and GPX3) collectively contributed to selenium tolerance, detoxification and protection against oxidative damage. GO and KEGG analysis highlighted alterations in amino acid metabolism, carbohydrate metabolism, and energy metabolism in the selenium biotransformation process. These findings provide novel insight into the mechanisms of selenium biotransformation in P. kudriavzevii and its application in the fields of the food industry.

Multi-omics strategy to reveal the numbing taste of Zanthoxylum armatum DC. at different developmental stages

Green Sichuan pepper (Zanthoxylum armatum DC.) holds important development and utilization values in the food and pharmaceutical industries due to the special pungent and numbing taste of their fruits. Interactions among genes, metabolites and proteins are activated at the molecular level during the development of Z. armatum fruits. In this study, five key stages of Z. armatum development were characterized using the transcriptome, metabolome and proteome, and a comprehensive analysis of the numbing taste was performed. Transcriptome enrichment analyses showed that the differentially expressed genes were mainly associated with flavonoid biosynthesis and tyrosine metabolism. The results also revealed that there were significant differences in the accumulation of metabolites such as fatty acids, tyrosine metabolism, terpenoids, and alkaloids in Z. armatum at different developmental stages. Proteome enrichment analyses indicated that these proteins were mainly involved in flavonoid biosynthesis, and terpenoid backbone biosynthesis. Integrated analysis of transcriptome, metabolome and proteome reveals that different developmental stages of Z. armatum were associated with flavonoid biosynthesis, fatty acids and terpenoid biosynthesis pathways, contributing to its numbing taste and medicinal value. These findings provide novel insights into the numbing taste of Z. armatum, and are expected to inform optimal harvesting times for enhanced economic benefits.

Integrated analysis of spatial transcriptomics and CT phenotypes for unveiling the novel molecular characteristics of recurrent and non-recurrent high-grade serous ovarian cancer

BackgroundHigh-grade serous ovarian cancer (HGSOC), which is known for its heterogeneity, high recurrence rate, and metastasis, is often diagnosed after being dispersed in several sites, with about 80% of patients experiencing recurrence. Despite a better understanding of its metastatic nature, the survival rates of patients with HGSOC remain poor.MethodsOur study utilized spatial transcriptomics (ST) to interpret the tumor microenvironment and computed tomography (CT) to examine spatial characteristics in eight patients with HGSOC divided into recurrent (R) and challenging-to-collect non-recurrent (NR) groups.ResultsBy integrating ST data with public single-cell RNA sequencing data, bulk RNA sequencing data, and CT data, we identified specific cell population enrichments and differentially expressed genes that correlate with CT phenotypes. Importantly, we elucidated that tumor necrosis factor-α signaling via NF-κB, oxidative phosphorylation, G2/M checkpoint, E2F targets, and MYC targets served as an indicator of recurrence (poor prognostic markers), and these pathways were significantly enriched in both the R group and certain CT phenotypes. In addition, we identified numerous prognostic markers indicative of nonrecurrence (good prognostic markers). Downregulated expression of PTGDS was linked to a higher number of seeding sites (≥ 3) in both internal HGSOC samples and public HGSOC TCIA and TCGA samples. Additionally, lower PTGDS expression in the tumor and stromal regions was observed in the R group than in the NR group based on our ST data. Chemotaxis-related markers (CXCL14 and NTN4) and markers associated with immune modulation (DAPL1 and RNASE1) were also found to be good prognostic markers in our ST and radiogenomics analyses.ConclusionsThis study demonstrates the potential of radiogenomics, combining CT and ST, for identifying diagnostic and therapeutic targets for HGSOC, marking a step towards personalized medicine.

Exploration of Response Mechanisms in the Gills of Pacific Oyster (Crassostrea gigas) to Cadmium Exposure through Integrative Metabolomic and Transcriptomic Analyses.

Marine mollusks, including oysters, are highly tolerant to high levels of cadmium (Cd), but the molecular mechanisms underlying their molecular response to acute Cd exposure remain unclear. In this study, the Pacific oyster Crassostrea gigas was used as a biological model, exposed to acute Cd stress for 96 h. Transcriptomic analyses of their gills were performed, and metabolomic analyses further validated these results. In our study, a total of 111 differentially expressed metabolites (DEMs) and 2108 differentially expressed genes (DEGs) were identified under acute Cd exposure. Further analyses revealed alterations in key genes and metabolic pathways associated with heavy metal stress response. Cd exposure triggered physiological and metabolic responses in oysters, including enhanced oxidative stress and disturbances in energy metabolism, and these changes revealed the biological response of oysters to acute Cd stress. Moreover, oysters could effectively enhance the tolerance and detoxification ability to acute Cd exposure through activating ABC transporters, enhancing glutathione metabolism and sulfur relay system in gill cells, and regulating energy metabolism. This study reveals the molecular mechanism of acute Cd stress in oysters and explores the molecular mechanism of high tolerance to Cd in oysters by using combined metabolomics and transcriptome analysis.

The underlying mechanisms by which boron mitigates copper toxicity in Citrus sinensis leaves revealed by integrated analysis of transcriptome, metabolome and physiology.

Both copper (Cu) excess and boron (B) deficiency are often observed in some citrus orchard soils. The molecular mechanisms by which B alleviates excessive Cu in citrus are poorly understood. Seedlings of sweet orange (Citrus sinensis (L.) Osbeck cv. Xuegan) were treated with 0.5 (Cu0.5) or 350 (Cu350 or Cu excess) μM CuCl2 and 2.5 (B2.5) or 25 (B25) μM HBO3 for 24wk. Thereafter, this study examined the effects of Cu and B treatments on gene expression levels revealed by RNA-Seq, metabolite profiles revealed by a widely targeted metabolome, and related physiological parameters in leaves. Cu350 upregulated 564 genes and 170 metabolites, and downregulated 598 genes and 58 metabolites in leaves of 2.5μM B-treated seedlings (LB2.5), but it only upregulated 281 genes and 100 metabolites, and downregulated 136 genes and 40 metabolites in leaves of 25μM B-treated seedlings (LB25). Cu350 decreased the concentrations of sucrose and total soluble sugars and increased the concentrations of starch, glucose, fructose and total nonstructural carbohydrates in LB2.5, but it only increased the glucose concentration in LB25. Further analysis demonstrated that B addition reduced the oxidative damage and alterations in primary and secondary metabolisms caused by Cu350, and alleviated the impairment of Cu350 to photosynthesis and cell wall metabolism, thus improving leaf growth. LB2.5 exhibited some adaptive responses to Cu350 to meet the increasing need for the dissipation of excessive excitation energy (EEE) and the detoxification of reactive oxygen species (reactive aldehydes) and Cu. Cu350 increased photorespiration, xanthophyll cycle-dependent thermal dissipation, nonstructural carbohydrate accumulation, and secondary metabolite biosynthesis and abundances; and upregulated tryptophan metabolism and related metabolite abundances, some antioxidant-related gene expression, and some antioxidant abundances. Additionally, this study identified some metabolic pathways, metabolites and genes that might lead to Cu tolerance in leaves.

Integrative transcriptome analysis reveals the molecular events underlying impaired T-cell responses in EGFR-mutant lung cancer

EGFR mutations are critical oncogenic drivers in lung adenocarcinoma (LUAD). However, the mechanisms by which they impact the tumor microenvironment (TME) and tumor immunity are unclear. Furthermore, the reasons underlying the poor response of EGFR-mutant (EGFR-MU) LUADs to immunotherapy with PD-1/PD-L1 inhibitors are unknown. Utilizing single-cell RNA (sc-RNA) and bulk RNA sequencing datasets, we conducted high-dimensional weighted gene coexpression network analysis to identify key genes and immune-related pathways contributing to the immunosuppressive TME. EGFR-MU cancer cells downregulated MHC class I genes to evade CD8+ cytotoxic T cells, expressed substantial levels of MHC class II molecules, and engaged with CD4+ regulatory T cells (Tregs). EGFR-MU tumors may recruit Tregs primarily through the CCL17/CCL22/CCR4 axis, leading to a Treg-enriched TME. High levels of MHC class II-positive cancer-associated fibroblasts and tumor endothelial cells were found within EGFR-MU tumors. Owing to the absence of costimulatory factors, they may inhibit rather than activate the tumor antigen-specific CD4+ T-cell response, contributing further to immune suppression. Multiplex immunohistochemistry analyses in a LUAD cohort confirmed increased expression of MHC class II molecules in cancer cells and fibroblasts in EGFR-MU tumors. Our research elucidates the highly immunosuppressive TME in EGFR-MU LUAD and suggests potential targets for effective immunotherapy.

Integrative transcriptomic and metabolomic analysis to elucidate the effect of gossypol on Enterobacter sp. GD5.

Gossypol, a yellow polyphenolic compound found in the Gossypium genus, is toxic to animals that ingest cotton-derived feed materials. However, ruminants display a notable tolerance to gossypol, attributed to the pivotal role of ruminal microorganisms in its degradation. The mechanisms of how rumen microorganisms degrade and tolerate gossypol remain unclear. Therefore, in this study, Enterobacter sp. GD5 was isolated from rumen fluid, and the effects of gossypol on its metabolism and gene expression were investigated using liquid chromatography-mass spectrometry (LC-MS) and RNA analyses. The LC-MS results revealed that gossypol significantly altered the metabolic profiles of 15 metabolites (eight upregulated and seven downregulated). The Kyoto Encyclopedia of Genes and Genomes analysis results showed that significantly different metabolites were associated with glutathione metabolism in both positive and negative ion modes, where gossypol significantly affected the biosynthesis of amino acids in the negative ion mode. Transcriptomic analysis indicated that gossypol significantly affected 132 genes (104 upregulated and 28 downregulated), with significant changes observed in the expression of catalase peroxidase, glutaredoxin-1, glutathione reductase, thioredoxin 2, thioredoxin reductase, and alkyl hydroperoxide reductase subunit F, which are related to antioxidative stress. Furthermore, Gene Ontology analysis revealed significant changes in homeostatic processes following gossypol supplementation. Overall, these results indicate that gossypol induces oxidative stress, resulting in the increased expression of antioxidative stress-related genes in Enterobacter sp. GD5, which may partially explain its tolerance to gossypol.

Establishment and transcriptome analysis of single blastomere-derived cell lines from zebrafish

Maintaining chromosome euploidy in zebrafish embryonic cells is challenging because of the degradation of genomic integrity during cell passaging. In this study, we report the derivation of zebrafish cell lines from single blastomeres. These cell lines have a stable chromosome status attributed to BMP4 and exhibit continuous proliferation in vitro. Twenty zebrafish cell lines are successfully established from single blastomeres. Single-cell transcriptome sequencing analysis confirms the fidelity of gene expression profiles throughout long-term culturing of at least 45 passages. The long-term cultured cells are specialized into epithelial cells, exhibiting similar expression patterns validated by integrative transcriptomic analysis. Overall, this work provides a protocol for establishing zebrafish cell lines from single blastomeres, which can serve as valuable tools for in vitro investigations of epithelial cell dynamics in terms of life–death balance and cell fate determination during normal homeostasis.