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Background: Circular RNAs (circRNAs) have been identified as playing an integral role in the development of bladder cancer (BC). However, the mechanism by which circRNAs operate in the chemical carcinogenesis of BC remains unclear. Methods: To explore this mechanism, we used RNA high-throughput sequencing to identify differentially expressed circRNA in bladder epithelial cells and chemically induced malignant transformed BC cells. Subsequently, in vitro experiments were conducted to investigate the biological function and molecular mechanism of circLMBR1 in BC. Finally, animal experiments were conducted to examine the clinical relevance of circLMBR1 in vivo. Results: Our profiling of circular RNA expression during cellular malignant transformation induced by chemical carcinogens identified a subset of circRNAs associated with cell transformation. We verified that the expression of circLMBR1 in bladder epithelial malignant transformed cells was decreased compared with control cells, as well as in BC tissues and bladder cell lines. Furthermore, circLMBR1 was seen to inhibit the proliferation, invasion, and migration of BC cells both in vitro and in vivo. Mechanistically, circLMBR1 was found to exert its antitumor effect by binding to the protein ALDH1A3. Conclusions: Our findings have revealed that circLMBR1 inhibits the progression of BC cells by binding to ALDH1A3 and upregulating its expression. As such, circLMBR1 serves as a promising predictor of BC and may provide a novel therapeutic target for the treatment of BC.
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Emerging evidence indicates that transfer RNA (tRNA)-derived small RNAs (tsRNAs), originated from tRNA with high abundance RNA modifications, play an important role in many complex physiological and pathological processes. However, the biological functions and regulatory mechanisms of modified tsRNAs in cancer remain poorly understood. Here, it is screened for and confirmed the presence of a novel m7G-modified tsRNA, m7G-3'-tiRNA LysTTT (mtiRL), in a variety of chemical carcinogenesis models by combining small RNA sequencing with an m7G small RNA-modified chip. Moreover, it is found that mtiRL, catalyzed by the tRNA m7G-modifying enzyme mettl1, promotes bladder cancer (BC) malignancy in vitro and in vivo. Mechanistically, mtiRL is found to specifically bind the oncoprotein Annexin A2 (ANXA2) to promote its Tyr24 phosphorylation by enhancing the interactions between ANXA2 and Yes proto-oncogene 1 (Yes1), leading to ANXA2 activation and increased p-ANXA2-Y24 nuclear localization in BC cells. Together, these findings define a critical role for mtiRL and suggest that targeting this novel m7G-modified tsRNA can be an efficient way for to treat BC.
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Objective: Bladder cancer is one of the most prominent malignancies affecting the urinary tract, characterized by a poor prognosis. Our previous research has underscored the pivotal role of m6A methylation in the progression of bladder cancer. Nevertheless, the precise relationship between N6-methyladenosine (m6A) regulation of long non-coding RNA (lncRNA) and bladder cancer remains elusive. Methods: This study harnessed sequencing data and clinical records from 408 bladder cancer patients in the TCGA database. Employing R software, we conducted bioinformatics analysis to establish an m6A-lncRNA co-expression network. Analyzing the differences between high and low-risk groups, particularly at the immunological level, and subsequently investigating the primary regulatory factors of these lncRNA, validating the findings through experiments, and exploring their specific cellular functions. Results: We identified 50 m6A-related lncRNA with prognostic significance through univariate Cox regression analysis. In parallel, we employed a LASSO-Cox regression model to pinpoint 11 lncRNA and calculate risk scores for bladder cancer patients. Based on the median risk score, patients were categorized into low-risk and high-risk groups. The high-risk cohort exhibited notably lower survival rates than their low-risk counterparts. Further analysis pointed to RBM15 and METTL3 as potential master regulators of these m6A-lncRNA. Experimental findings also shed light on the upregulated expression of METTlL3 and RBM15 in bladder cancer, where they contributed to the malignant progression of tumors. The experimental findings demonstrated a significant upregulation of METTL3 and RBM15 in bladder cancer specimens, implicating their contributory role in the oncogenic progression. Knockdown of METTL3 and RBM15 resulted in a marked attenuation of tumor cell proliferation, invasion, and migration, which was concomitant with a downregulation in the cellular m6A methylation status. Moreover, these results revealed that RBM15 and METTL3 function in a synergistic capacity, positing their involvement in cancer promotion via the upregulation of m6A modifications in long non-coding RNAs. Additionally, this study successfully developed an N-methyl-N-nitrosourea (MNU)-induced rat model of in situ bladder carcinoma, confirming the elevated expression of RBM15 and METTL3, which paralleled the overexpression of m6A-related- lncRNAs observed in bladder cancer cell lines. This congruence underscores the potential utility of these molecular markers in in vivo models that mirror human malignancies. Conclusion: This study not only offers novel molecular targets,but also enriches the research on m6A modification in bladder cancer, thereby facilitating its clinical translation.
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3-Methylcholanthracene (3-MC) is one of the most carcinogenic polycyclic aromatic hydrocarbons (PAHs). Long-term exposure to PAHs has been thought of as an important factor in urothelial tumorigenesis. N6-methyladenosine (m6A) exists widely in eukaryotic organisms and regulates the expression level of specific genes by regulating mRNA stability, translation efficiency, and nuclear export efficiency. Currently, the potential molecular mechanisms that regulate m6A modification for 3-MC carcinogenesis remain unclear. Here, we profiled mRNA, m6A, translation and protein level using "-omics" methodologies, including transcriptomes, m6A profile, translatomes, and proteomics in 3-MC-transformed urothelial cells and control cells. The key molecules SLC3A2/SLC7A5 were screened and identified in 3-MC-induced uroepithelial transformation. Moreover, SLC7A5/SLC3A2 promoted uroepithelial cells malignant phenotype in vitro and in vivo. Mechanically, METTL3 and ALKBH5 mediated m6A modification of SLC3A2/SLC7A5 mRNA in 3-MC-induced uroepithelial transformation by upregulating the translation of SLC3A2/SLC7A5. Furthermore, programmable m6A modification of SLC3A2/SLC7A5 mRNA affected the expression of its proteins. Taken together, our results revealed that the m6A modification-mediated SLC3A2/SLC7A5 translation promoted 3-MC-induced uroepithelial transformation, suggesting that targeting m6A modification of SLC3A2/SLC7A5 may be a potential therapeutic strategy for bladder cancer related to PAHs.
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Transportador 1 de Aminoácidos Neutros Grandes , Hidrocarbonetos Policíclicos Aromáticos , Humanos , Metilcolantreno/toxicidade , Carcinogênese , Transformação Celular Neoplásica/induzido quimicamente , Transformação Celular Neoplásica/genética , RNA Mensageiro/genética , Metiltransferases/genética , Cadeia Pesada da Proteína-1 Reguladora de FusãoRESUMO
INTRODUCTION: N6-methyladenosine (m6A) modification contributes to the pathogenesis and development of various cancers, including bladder cancer (BCa). In particular, integrin α6 (ITGA6) promotes BCa progression by cooperatively regulating multisite m6A modification. However, the therapeutic effect of targeting ITGA6 multisite m6A modifications in BCa remains unknown. OBJECTIVES: We aim to develop a multisite dCasRx- m6A editor for assessing the effects of the multisite dCasRx-m6A editor targeted m6A demethylation of ITGA6 mRNA in BC growth and progression. METHODS: The multisite dCasRx- m6A editor was generated by cloning. m6A-methylated RNA immunoprecipitation (meRIP), luciferase reporter, a single-base T3 ligase-based qPCR-amplification, Polysome profiling and meRIP-seq experiments were performed to determine the targeting specificity of the multisite dCasRx-m6A editor. We performed cell phenotype analysis and used in vivo mouse xenograft models to assess the effects of the multisite dCasRx-m6A editor in BC growth and progression. RESULTS: We designed a targeted ITGA6 multi-locus guide (g)RNA and established a bidirectional deactivated RfxCas13d (dCasRx)-based m6A-editing platform, comprising a nucleus-localized dCasRx fused with the catalytic domains of methyltransferase-like 3 (METTL3-CD) or α-ketoglutarate-dependent dioxygenase alkB homolog 5 (ALKBH5-CD), to simultaneously manipulate the methylation of ITGA6 mRNA at four m6A sites. The results confirmed the dCasRx-m6A editor modified m6A at multiple sites in ITGA6 mRNA, with low off-target effects. Moreover, targeted m6A demethylation of ITGA6 mRNA by the multisite dCasRx-m6A editor significantly reduced BCa cell proliferation and migration in vitro and in vivo. Furthermore, the dCasRx-ALKBH5-CD and ITGA6 multi-site gRNA delivered to 5-week-old BALB/cJNju-Foxn1nu/Nju nude mice via adeno-associated viral vectors significantly inhibited BCa cell growth. CONCLUSION: Our study proposes a novel therapeutic tool for the treatment of BC by applying the multisite dCasRx-m6A editor while highlighting its potential efficacy for treating other diseases associated with abnormal m6A modifications.
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RNA Guia de Sistemas CRISPR-Cas , Neoplasias da Bexiga Urinária , Humanos , Camundongos , Animais , Integrina alfa6/genética , Integrina alfa6/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Camundongos Nus , Linhagem Celular Tumoral , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Desmetilação , Metiltransferases/genética , Metiltransferases/metabolismoRESUMO
Iron deficiency (ID) is the biggest cause of anemia. This pilot study aimed to investigate the effects of food-derived oligopeptide iron chelates on ameliorating liver injury and restoring gut microbiota homeostasis in iron-deficiency anemia (IDA) female rats. Female Sprague-Dawley rats at 21 days old were selected and randomly divided into a control group (N = 4) and an ID model group (N = 16). The ID model group was fed an iron-deficient diet containing 4 mg kg-1 iron for 28 days to generate the IDA rat model and then randomly subdivided into four groups (N = 4 for each group): ID group, ferrous sulfate group, marine fish oligopeptide iron chelate (MCOP-Fe) group, and whey protein oligopeptide iron chelate (WPP-Fe) group. Iron supplements were given to rats in the three intervention groups once per day via intragastric administration for three weeks. After iron supplementation, the hemoglobin levels in the three intervention groups were significantly improved, with the MCOP-Fe and WPP-Fe groups returning to normal. The ALT and AST levels in the ID group increased significantly, while levels in all intervention groups decreased to normal levels. Liver glutathione in the WPP-Fe group was increased, while the activity of superoxide dismutase also tended to be higher. In addition, 16S rRNA gene sequencing showed that IDA resulted in changes to intestinal microbiota. After intervention, the WPP-Fe group showed increased alpha diversity of intestinal microbes. Therefore, MCOP-Fe and WPP-Fe may improve the iron status of IDA female rats as well as ameliorate liver damage, with WPP-Fe showing a greater potential in improving gut microbiota imbalance.
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Anemia Ferropriva , Microbioma Gastrointestinal , Deficiências de Ferro , Ratos , Feminino , Animais , Ferro/metabolismo , Anemia Ferropriva/tratamento farmacológico , Anemia Ferropriva/metabolismo , Projetos Piloto , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/metabolismo , Ratos Sprague-Dawley , Oligopeptídeos/metabolismo , Fígado/metabolismo , Quelantes de Ferro/metabolismoRESUMO
RNA modifications, including adenine methylation (m6A) of mRNA and guanine methylation (m7G) of tRNA, are crucial for the biological function of RNA. However, the mechanism underlying the translation of specific genes synergistically mediated by dual m6A/m7G RNA modifications in bladder cancer (BCa) remains unclear. We demonstrated that m6A methyltransferase METTL3-mediated programmable m6A modification of oncogene trophoblast cell surface protein 2 (TROP2) mRNA promoted its translation during malignant transformation of bladder epithelial cells. m7G methyltransferase METTL1 enhanced TROP2 translation by mediating m7G modification of certain tRNAs. TROP2 protein inhibition decreased the proliferation and invasion of BCa cells in vitro and in vivo. Moreover, synergistical knockout of METTL3/METTL1 inhibited BCa cell proliferation, migration, and invasion; however, TROP2 overexpression partially abrogated its effect. Furthermore, TROP2 expression was significantly positively correlated with the expression levels of METTL3 and METTL1 in BCa patients. Overall, our results revealed that METTL3/METTL1-mediated dual m6A/m7G RNA modifications enhanced TROP2 translation and promoted BCa development, indicating a novel RNA epigenetic mechanism in BCa.
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Antígenos de Neoplasias , Moléculas de Adesão Celular , Neoplasias da Bexiga Urinária , Humanos , Metilação , Metiltransferases/genética , Metiltransferases/metabolismo , RNA/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Neoplasias da Bexiga Urinária/patologia , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismoRESUMO
It has been reported that particulate matter with an aerodynamic diameter of <2.5 µm (PM2.5) could induce epithelial-mesenchymal transition (EMT)- and extracellular matrix (ECM)-related pulmonary fibrosis (PF). The transcription factor Nrf2 alleviated PM2.5-induced PF by antagonizing oxidative stress. The N6-methyladenosine (m6A) modification plays a significant role in the stress response. However, the effect of m6A modification on the mechanisms of Nrf2-mediated defense against PM2.5-induced PF remained unknown. Here, we explored the role and the underlying molecular mechanisms of m6A methylation of Nrf2 mRNA in PM2.5-induced PF. We established filtered air (FA), unfiltered air (UA), and concentrated PM2.5 air (CA) group mice model and 0, 50, and 100 µg/mL PM2.5-treated 16HBE cell models. The extent of lung fibrosis in mice and fibrosis indicators were detected by histopathological analysis, immunohistochemical staining and western blotting. The molecular mechanism of m6A-modified Nrf2 was demonstrated by m6A-methylated RNA immunoprecipitation (MeRIP), RNA immunoprecipitation (RIP), qRT-PCR and T3 ligase-based PCR. Our data showed that PM2.5 exposure for 16 weeks could induce pulmonary fibrosis and activate Nrf2 signaling pathway. m6A methyltransferase METTL3 was upregulated after PM2.5 treatment in vivo and in vitro. Moreover, METTL3 mediated m6A modification of Nrf2 mRNA and promoted Nrf2 translation in mice and 16HBE cells after PM2.5 exposure. Mechanistically, three m6A-modified sites (1317, 1376 and 935; numbered relative to the first nucleotide of 3'UTR) of Nrf2 mRNA were identified in PM2.5-treatment 16HBE cells. Furthermore, the m6A binding proteins YTHDF1/IGF2BP1 promoted Nrf2 translation by binding to m6A residues of Nrf2 mRNA. Our results revealed the mechanism of m6A mediated Nrf2 signaling pathway against oxidative stress, which affected the development of PM2.5-induced PF.
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Fibrose Pulmonar , Camundongos , Animais , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Material Particulado/toxicidade , RNA , RNA Mensageiro/genéticaRESUMO
Background: One of the most common malignant tumors of the urinary system is muscle-invasive bladder cancer (MIBC). With the increased use of immunotherapy, its importance in the field of cancer is becoming abundantly evident. This study classifies MIBC according to GSVA score from the perspective of the GSEA immune gene set. Methods: This study integrated the sequencing and clinical data of MIBC patients in TCGA and GEO databases, then scored the data using the GSVA algorithm, the CNMF algorithm was implemented to divide the subtypes of GEO and TCGA datasets, respectively, and finally screened and determined the key pathways in combination with clinical data. Simultaneously, LASSO Cox regression model was constructed based on key pathway genes to assess the model's predictive ability (ROC) and describe the immune landscape differences between high- and low-risk groups; key genes were further analyzed and verified in patient tissues. Results: 404 TCGA and 297 GEO datasets were divided into C1-3 groups (TCGA-C1:120/C2:152/C3:132; GEO- C1:112/C2:101/C3:84), of which TCGA-C2 (n = 152) subtype and GEO-C1 (n = 112) subtype had the worst prognosis. LASSO Cox regression model with ROC (train set = 0.718, test set = 0.667) could be constructed. When combined with the Cancer Immunome Atlas database, it was found that patients with high-risk scores were more sensitive to PD-1 inhibitor and PD-1 inhibitor combined with CTLA-4. NXPH4, as a key gene, plays a role in MIBC with tissue validation results show that nxph4 is highly expressed in tumor. Conclusion: The immune gene score of MIBC data in TCGA and GEO databases was successfully evaluated using GSVA in this research. The lasso Cox expression model was successfully constructed by screening immune genes, the high-risk group had a worse prognosis and higher sensitivity to immunotherapy, PD-1 inhibitors or PD-1 combined with CTLA-4 inhibitors can be preferentially used in high-risk patients who are sensitive to immunotherapy, and NXPH4 may be a molecular target to adjust the effect of immunotherapy.
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This study aimed to investigate anemia treatment and other potential effects of two food-derived bioactive oligopeptide iron complexes on pregnant rats with iron deficiency anemia (IDA) and their offspring. Rats with IDA were established with a low iron diet and then mated. There were one control group and seven randomly assigned groups of pregnant rats with IDA: Control group [Control, 40 ppm ferrous sulfate (FeSO4)]; IDA model group (ID, 4 ppm FeSO4), three high-iron groups (H-FeSO4, 400 ppm FeSO4; MCOP-Fe, 400 ppm marine fish oligopeptide iron complex; WCOP-Fe, 400 ppm whey protein oligopeptide iron complex) and three low-iron groups (L-FeSO4, 40 ppm FeSO4; MOP-Fe, 40 ppm marine fish oligopeptide iron complex; WOP-Fe, 40 ppm whey protein oligopeptide iron complex). Rats in each group were fed the corresponding special diet during pregnancy until the day of delivery. After different doses of iron supplement, serum hemoglobin, iron, and ferritin levels in rats with IDA were significantly increased to normal levels (P < 0.05). Serum iron levels were significantly lower in two food-derived bioactive oligopeptide low-iron complex groups than in the low FeSO4 group (P<0.05). Liver malondialdehyde levels were significantly increased in the three high-iron groups compared with the other five groups (P < 0.05), and hemosiderin deposition was observed in liver tissue, indicating that the iron dose was overloaded and aggravated the peroxidative damage in pregnant rats. Liver inflammation was reduced in the three low-iron groups. Tumor necrosis factor α secretion was significantly decreased in all groups with supplemented oligopeptide (P < 0.05), with the concentration of tumor necrosis factor α declining to normal levels in the two whey protein oligopeptide iron complex groups. In the marine fish oligopeptide iron complex groups, body length, tail length, and weight of offspring were significantly increased (P < 0.05) and reached normal levels. Therefore, food-derived bioactive oligopeptide (derived from marine fish skin and milk) iron complexes may be an effective type of iron supplement for pregnancy to improve anemia, as well as reduce the side effects of iron overload, and improve the growth and nutritional status of offspring.
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BACKGROUND: Iron deficiency (ID) impairs patient physical activity, recognition and life quality, which is difficult to perceive but should not be underestimated. Worldwide efforts have been made to lower ID burden, however, whether it decreased equally in different regions and sexes is unclear. This study is to examine regional and sex inequalities in global ID from 1990 to 2017. METHODS: We conducted a longitudinal, comparative burden-of-disease study. Disability-adjusted life-years (DALYs) of ID were obtained from Global Burden of Disease Report 2017. Human Development Index (HDI) data were obtained from Human Development Report 2017. Gini coefficient and the concentration index were calculated to assess the equities in global burden of ID. RESULTS: A downward trend of global ID burden (from 569.3 (95% Uncertainty Interval [UI]: 387.8-815.6) to 403.0 (95% UI: 272.4-586.6), p < 0.001), age-adjusted DALYs per 100,000 population) but an uptrend of its inequalities (from 0.366 to 0.431, p < 0.001, Gini coefficients) was observed between 1990 and 2017. ID burden was heavier in women than that in men ([age-adjusted DALYs per 100,000 population from 742.2 to 514.3] vs [from 398.5 to 291.9]), but its inequalities were higher in men since 1990. The between-sex gap of ID burden was narrowed with higher HDI (ß = - 364.11, p < 0.001). East Asia & Pacific and South Asia regions made a big stride for ID control in both sexes over decades [age-adjusted DALYs per 100,000 population from 378.7 (95% UI: 255.8-551.7) in 1990 to 138.9 (95%UI: 91.8-206.5) in 2017], while a heavy burden among Sub-Saharan African men was persistent[age-adjusted DALYs per 100,000 population, 572.5 (95% UI: 385.3-815) in 1990 and 562.6 (95% UI: 367.9-833.3) in 2017]. CONCLUSIONS: Redistributing attention and resources to help countries with low HDI, especially take care of women with low socioeconomic status (SES) and men under high ID burden may help hold back the expanding ID inequality.
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Pessoas com Deficiência , Deficiências de Ferro , Feminino , Carga Global da Doença , Saúde Global , Humanos , Masculino , Anos de Vida Ajustados por Qualidade de VidaRESUMO
BACKGROUND: Oncogenic metabolic reprogramming contributes to tumor growth and immune evasion. The intertumoral metabolic heterogeneity and interaction of distinct metabolic pathways may determine patient outcomes. In this study, we aim to determine the clinical and immunological significance of metabolic subtypes according to the expression levels of genes related to glycolysis and cholesterol-synthesis in bladder cancer (BCa). METHODS: Based on the median expression levels of glycolytic and cholesterogenic genes, patients were stratified into 4 subtypes (mixed, cholesterogenic, glycolytic, and quiescent) in an integrated cohort including TCGA, GSE13507, and IMvigor210. Clinical, genomic, transcriptomic, and tumor microenvironment characteristics were compared between the 4 subtypes. RESULTS: The 4 metabolic subtypes exhibited distinct clinical, molecular, and genomic patterns. Compared to quiescent subtype, mixed subtype was more likely to be basal tumors and was significantly associated with poorer prognosis even after controlling for age, gender, histological grade, clinical stage, and molecular phenotypes. Additionally, mixed tumors harbored a higher frequency of RB1 and LRP1B copy number deletion compared to quiescent tumors (25.7% vs. 12.7 and 27.9% vs. 10.2%, respectively, both adjusted P value< 0.05). Furthermore, aberrant PIK3CA expression level was significantly correlated with those of glycolytic and cholesterogenic genes. The quiescent subtype was associated with lower stemness indices and lower signature scores for gene sets involved in genomic instability, including DNA replication, DNA damage repair, mismatch repair, and homologous recombination genes. Moreover, quiescent tumors exhibited lower expression levels of pyruvate dehydrogenase kinases 1-3 (PDK1-3) than the other subtypes. In addition, distinct immune cell infiltration patterns were observed across the 4 metabolic subtypes, with greater infiltration of M0/M2 macrophages observed in glycolytic and mixed subtypes. However, no significant difference in immunotherapy response was observed across the 4 metabolic subtypes. CONCLUSION: This study proposed a new metabolic subtyping method for BCa based on genes involved in glycolysis and cholesterol synthesis pathways. Our findings may provide novel insight for the development of personalized subtype-specific treatment strategies targeting metabolic vulnerabilities.
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Colesterol/biossíntese , Glicólise/genética , Sistema Imunitário/citologia , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/imunologia , Classe I de Fosfatidilinositol 3-Quinases/genética , Variações do Número de Cópias de DNA , Reparo do DNA/genética , Bases de Dados Genéticas , Instabilidade Genômica/genética , Glicólise/imunologia , Humanos , Linfócitos do Interstício Tumoral/imunologia , Oncogenes/genética , Oncogenes/imunologia , Polimorfismo de Nucleotídeo Único , Prognóstico , Receptores de LDL/genética , Proteínas de Ligação a Retinoblastoma/genética , Transdução de Sinais , Transcriptoma , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia , Macrófagos Associados a Tumor/imunologia , Ubiquitina-Proteína Ligases/genética , Neoplasias da Bexiga Urinária/mortalidadeRESUMO
BACKGROUND: The posttranscriptional modifications of transfer RNA (tRNA) are critical for all aspects of the tRNA function and have been implicated in the tumourigenesis and progression of many human cancers. By contrast, the biological functions of methyltransferase-like 1 (METTL1)-regulated m7 G tRNA modification in bladder cancer (BC) remain obscure. RESULTS: In this research, we show that METTL1 was highly expressed in BC, and its level was correlated with poor patient prognosis. Silencing METTL1 suppresses the proliferation, migration and invasion of BC cells in vitro and in vivo. Multi-omics analysis reveals that METTL1-mediated m7 G tRNA modification altered expression of certain target genes, including EGFR/EFEMP1. Mechanistically, METTL1 regulates the translation of EGFR/EFEMP1 via modifying certain tRNAs. Furthermore, forced expression of EGFR/EFEMP1 partially rescues the effect of METTL1 deletion on BC cells. CONCLUSIONS: Our findings demonstrate the oncogenic role of METTL1 and the pathological significance of the METTL1-m7 G-EGFR/EFEMP1 axis in the BC development, thus providing potential therapeutic targets for the BC treatment.
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Proteínas da Matriz Extracelular/efeitos adversos , Metiltransferases/efeitos adversos , Neoplasias da Bexiga Urinária/genética , Carcinogênese , Receptores ErbB/efeitos adversos , Receptores ErbB/genética , Proteínas da Matriz Extracelular/genética , Humanos , Metiltransferases/genética , Neoplasias da Bexiga Urinária/etiologiaRESUMO
Both lncRNAs and the N6-methyladenosine (m6A) modification are key regulators of tumorigenesis and innate immunity. However, little is known about the m6A modification of lncRNAs and their clinical and immune relevance in bladder cancer. In this study, we identified m6A-related lncRNAs using Pearson correlation analysis in The Cancer Genome Atlas (TCGA) and the IMvigor210 datasets. Next, univariate Cox regression was performed using the TCGA dataset to filter prognostic m6A-related lncRNAs, which were further subjected to the least absolute shrinkage and selection operator (LASSO) Cox regression to establish a 12 m6A-related lncRNA prognostic score (m6A-LRS). The m6A-LRS was validated in the IMvigor210 dataset. In addition, high m6A-LRS tumors, characterized by decreased tumor mutation load and neoantigen load, showed poorer response to immunotherapy than those with low m6A-LRS in the IMvigor210 dataset. Further, we constructed an m6A-LRS-based nomogram that demonstrated a strong ability to predict overall survival in patients with bladder cancer. Moreover, enrichment analysis revealed that tumor-associated biological processes, oncogenic signaling, and tumor hallmarks were commonly associated with a high m6A-LRS. Gene set variation analysis also indicated that high m6A-LRS was associated with activation of canonical oncogenic signatures, such as the epithelial-to-mesenchymal transition, cell cycle regulators, and DNA replication, as well as activation of immunosuppressive signatures, such as the T-cell exhaustion and pan-fibroblast-TGF-ß response signatures. Furthermore, we observed distinct tumor microenvironment cell infiltration characteristics between high- and low-risk tumors. High m6A-LRS tumors showed reduced infiltration of CD8+ T-cells and enhanced infiltration of macrophages and fibroblasts. Additionally, we established a competing endogenous RNA network based on the12 m6A-related lncRNAs. Finally, three lncRNAs (SNHG16, SBF2-AS1, and BDNF-AS) were selected for further validation. The qualitative PCR assay on 10 pairs of bladder cancer and adjacent normal control samples validated the differential expression, and methylated RNA immunoprecipitation (MeRIP) analysis demonstrated a robust m6A enrichment in T24 bladder cancer cells compared with normal uroepithelial cells (SVHUC-1). In conclusion, this study introduced an m6A-related lncRNA signature that identified a subgroup of patients with poor prognoses and suboptimal immune responses, thus providing novel approaches for treatment response prediction and patient stratification in bladder cancer.
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Objective: This study aimed to investigate the mechanisms underlying Cd-induced urothelial transformation, using multi-omics analyses (transcriptome, epitranscriptome, and proteome).Methods: Transcriptomics analysis was performed to estimate the expression of genes, methylated RNA immunoprecipitation sequencing analysis was used to detect m6A modification, while proteomics analysis was used to identify differentially expressed proteins. Differentially expressed genes (DEGs) were subjected to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis.Results: A total of 9491 DEGs, 711 differentially expressed proteins, and 633 differentially m6A modified genes between Cd-transformed cells and control cells were identified. The regulation of most genes varied at different omics layers. The three omics data shared 57 genes, and these genes were enriched in response to DNA damage stimulus and cell proliferation. Interestingly, 13 genes, most of which are related to the onset or progression of cancer, were shared by the m6A and proteomics data, but not the transcriptome data. This suggested that m6A modification is crucial for post-transcriptional regulation related to Cd2+-induced malignant transformation.Conclusion: Our multi-omics analysis provided a comprehensive reference map of gene activity and revealed m6A signalling pathways crucial for Cd2+ carcinogenesis.
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Cádmio/toxicidade , Transformação Celular Neoplásica/efeitos dos fármacos , Perfilação da Expressão Gênica , Metiltransferases/metabolismo , RNA Mensageiro/metabolismo , Neoplasias da Bexiga Urinária/induzido quimicamente , Urotélio/efeitos dos fármacos , Linhagem Celular Transformada , Humanos , Proteômica/métodos , Análise de Sequência de RNA/métodos , Urotélio/patologiaRESUMO
Accumulating evidence has revealed significant roles for N6-methyladenosine (m 6 A) modification in the development of various cancers. We previously demonstrated an oncogenic role of m 6 A-modified CUB domain containing protein 1 (CDCP1) in bladder cancer (BC) progression. However, the biological functions and underlying molecular mechanisms of engineered programmable m 6 A modification of CDCP1 mRNA in BC remain obscure. Here, we established a targeted m 6 A RNA methylation system by fusing the catalytic domain of methyltransferase like 3 (METTL3CD) to RCas9 as the RNA-targeting module. The constructed RCas9- METTL3 retained methylation activity and mediated efficient site-specific m 6 A installation in the presence of a cognate single guide RNA and short protospacer adjacent motif-containing ssDNA molecule . Subsequently, targeting m 6 A installation onto the 3' untranslated region of CDCP1 promoted CDCP1 mRNA translation and facilitated BC development in vitro and in vivo. Our findings demonstrate that the RCas9-METTL3 system mediates efficient sitespecific m 6 A installation on CDCP1 mRNA and promotes BC development. Thus, the RCas9-METTL3 system provides a new tool for studying m 6 A function and a potential strategy for BC epitranscriptome-modulating therapies.
Assuntos
Adenosina/análogos & derivados , Antígenos de Neoplasias/genética , Carcinogênese/patologia , Moléculas de Adesão Celular/genética , Metiltransferases/metabolismo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Adenosina/metabolismo , Antígenos de Neoplasias/metabolismo , Sistemas CRISPR-Cas/genética , Carcinogênese/genética , Moléculas de Adesão Celular/metabolismo , Linhagem Celular Tumoral , Humanos , Biossíntese de Proteínas , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
Yersinia pestis, a Gram-negative bacterium, is the etiologic agent of plague. A hallmark of Y. pestis infection is the organism's ability to rapidly disseminate through an animal host. Y. pestis expresses the outer membrane protein, Ail (Attachment invasion locus), which is associated with host invasion and serum resistance. However, whether Ail plays a role in host dissemination remains unclear. In this study, C57BL/6J mice were challenged with a defined Y. pestis strain, KimD27, or an isogenic ail-deleted mutant derived from KimD27 via metacarpal paw pad inoculation, nasal drops, orogastric infection, or tail vein injection to mimic bubonic, pneumonic, oral, or septicemic plague, respectively. Our results showed that ail-deleted Y. pestis KimD27 lost the ability to invade host cells, leading to failed host dissemination in the pneumonic and oral plague models but not in the bubonic or septicemic plague models, which do not require invasiveness. Therefore, this study demonstrated that whether Ail plays a role in Y. pestis pathogenesis depends on the infection route.
Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Peste/microbiologia , Fatores de Virulência/metabolismo , Virulência , Yersinia pestis , Animais , Proteínas de Bactérias/metabolismo , Modelos Animais de Doenças , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Boca/microbiologia , Yersinia pestis/metabolismo , Yersinia pestis/patogenicidadeRESUMO
BACKGROUND: Accumulating evidence has revealed the critical roles of N6-methyladenosine (m6A) modification of mRNA in various cancers. However, the biological function and regulation of m6A in bladder cancer (BC) are not yet fully understood. METHODS: We performed cell phenotype analysis and established in vivo mouse xenograft models to assess the effects of m6A-modified ITGA6 on BC growth and progression. Methylated RNA immunoprecipitation (MeRIP), RNA immunoprecipitation and luciferase reporter and mutagenesis assays were used to define the mechanism of m6A-modified ITGA6. Immunohistochemical analysis was performed to assess the correlation between METTL3 and ITGA6 expression in bladder cancer patients. FINDINGS: We show that the m6A writer METTL3 and eraser ALKBH5 altered cell adhesion by regulating ITGA6 expression in bladder cancer cells. Moreover, upregulation of ITGA6 is correlated with the increase in METTL3 expression in human BC tissues, and higher expression of ITGA6 in patients indicates a lower survival rate. Mechanistically, m6A is highly enriched within the ITGA6 transcripts, and increased m6A methylations of the ITGA6 mRNA 3'UTR promotes the translation of ITGA6 mRNA via binding of the m6A readers YTHDF1 and YTHDF3. Inhibition of ITGA6 results in decreased growth and progression of bladder cancer cells in vitro and in vivo. Furthermore, overexpression of ITGA6 in METTL3-depleted cells partially restores the BC adhesion, migration and invasion phenotypes. INTERPRETATION: Our results demonstrate an oncogenic role of m6A-modified ITGA6 and show its regulatory mechanisms in BC development and progression, thus identifying a potential therapeutic target for BC. FUND: This work was supported by National Natural Science Foundation of China (81772699, 81472999).
Assuntos
Adenosina/análogos & derivados , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Integrina alfa6/genética , RNA Mensageiro/genética , Neoplasias da Bexiga Urinária/genética , Adenosina/farmacologia , Adulto , Idoso , Homólogo AlkB 5 da RNA Desmetilase/genética , Animais , Adesão Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Modelos Animais de Doenças , Progressão da Doença , Feminino , Humanos , Imuno-Histoquímica , Integrina alfa6/metabolismo , Masculino , Metiltransferases/genética , Camundongos , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologiaRESUMO
Salmonella enterica serovar Typhimurium, a Gram-negative bacterium, can cause infectious diseases ranging from gastroenteritis to systemic dissemination and infection. However, the molecular mechanisms underlying this bacterial dissemination have yet to be elucidated. A study indicated that using the lipopolysaccharide (LPS) core as a ligand, S Typhimurium was able to bind human dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (hCD209a), an HIV receptor that promotes viral dissemination by hijacking antigen-presenting cells (APCs). In this study, we showed that S Typhimurium interacted with CD209s, leading to the invasion of APCs and potentially the dissemination to regional lymph nodes, spleen, and liver in mice. Shielding of the exposed LPS core through the expression of O-antigen reduces dissemination and infection. Thus, we propose that similar to HIV, S Typhimurium may also utilize APCs via interactions with CD209s as a way to disseminate to the lymph nodes, spleen, and liver to initiate host infection.