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Prostate cancer (PCa) currently ranks second in male tumor mortality. Targeting immune checkpoint in tumor as immunotherapy is a new direction for tumor treatment. However, targeting PD-1/PD-L1 and CTLA4 to treat PCa has poor immunotherapeutic efficacy because PCa is known as a cold tumor. Understanding the mechanism of immunosuppression in PCa can promote the use of immunotherapy to treat PCa. ELAVL1 is highly expressed in many tumors, participates in almost all tumor biological activities and is an oncogene. ELAVL1 is also involved in the development and differentiation of T and B lymphocytes. However, the relationship between ELAVL1 and tumor immunity has not yet been reported. In recent years, ELAVL1 has been shown to regulate downstream targets in an m6A -dependent manner. PD-L1 has been shown to have m6A sites in multiple tumors that are regulated by m6A. In this study, ELAVL1 was highly expressed in PCa, and PCa with high ELAVL1 expression is immunosuppressive. Knocking down ELAVL1 reduced PD-L1 expression in PCa. Moreover, PD-L1 was shown to have an m6A site, and its m6A level was upregulated in PCa. ELAVL1 interacts with PD-L1 mRNA and promotes PD-L1 RNA stability via m6A, ultimately inhibiting the infiltration of CD4-positive T cells. In addition, androgen receptor (AR) was shown to be regulated with ELAVL1, and knocking down AR could also affect the expression of PD-L1. Therefore, ELAVL1 can directly or indirectly regulate the expression of PD-L1, thereby affecting the infiltration of CD4-positive T cells in PCa and ultimately leading to immune suppression.
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OBJECTIVE: To find the relationship between N6-methyladenosine (m6A) genes and Major Depressive Disorder (MDD). METHODS: Differential expression of m6A associated genes between normal and MDD samples was initially identified. Subsequent analysis was conducted on the functions of these genes and the pathways they may affect. A diagnostic model was constructed using the expression matrix of these differential genes, and visualized using a nomogram. Simultaneously, an unsupervised classification method was employed to classify all patients based on the expression of these m6A associated genes. Following this, common differential genes among different clusters were computed. By analyzing the functions of the common differential expressed genes among clusters, the role of m6A-related genes in the pathogenesis of MDD patients was elucidated. RESULTS: Differential expression was observed in ELAVL1 and YTHDC2 between the MDD group and the control group. ELAVL1 was associated with comorbid anxiety in MDD patients. A linear regression model based on these two genes could accurately predict whether patients in the GSE98793 dataset had MDD and could provide a net benefit for clinical decision-making. Based on the expression matrix of ELAVL1 and YTHDC2, MDD patients were classified into three clusters. Among these clusters, there were 937 common differential genes. Enrichment analysis was also performed on these genes. The ssGSEA method was applied to predict the content of 23 immune cells in the GSE98793 dataset samples. The relationship between these immune cells and ELAVL1, YTHDC2, and different clusters was analyzed. CONCLUSION: Among all the m6A genes, ELAVL1 and YTHDC2 are closely associated with MDD, ELAVL1 is related to comorbid anxiety in MDD. ELAVL1 and YTHDC2 have opposite associations with immune cells in MDD.
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Adenosina , Trastorno Depresivo Mayor , Humanos , Trastorno Depresivo Mayor/genética , Adenosina/análogos & derivados , Adenosina/genética , Femenino , Masculino , Metilación , Proteínas de Unión al ARN/genética , Adulto , Nomogramas , ARN HelicasasRESUMEN
Circular RNAs (circRNAs) have been discovered to serve as vital regulators in atherosclerosis (AS). However, the role and mechanism of circ_0002331 in AS process are still unclear. Human umbilical vein endothelial cells (HUVECs) were treated with ox-LDL to establish an in vitro model for AS. The expression levels of circ_0002331, Cyclin D2 (CCND2) and ELAVL1 were analyzed by quantitative real-time PCR. Cell proliferation, apoptosis, migration, invasion and angiogenesis were assessed by EdU assay, flow cytometry, transwell assay and tube formation assay. The protein levels of CCND2, ELAVL1, and autophagy-related markers were detected using western blot analysis. IL-8 level was analyzed by ELISA. The relationship between ELAVL1 and circ_0002331 or CCND2 was analyzed by RIP assay and RNA pull-down assay. Moreover, FISH assay was used to analyze the co-localization of ELAVL1 and CCND2 in HUVECs. Our data showed that circ_0002331 was obviously downregulated in AS patients and ox-LDL-induced HUVECs. Overexpression of circ_0002331 could promote proliferation, migration, invasion and angiogenesis, while inhibit apoptosis, autophagy and inflammation in ox-LDL-induced HUVECs. Furthermore, CCND2 was positively regulated by circ_0002331, and circ_0002331 could bind with ELAVL1 to promote CCND2 mRNA stability. Besides, CCND2 overexpression suppressed ox-LDL-induced HUVECs dysfunction, and its knockdown also reversed the regulation of circ_0002331 on ox-LDL-induced HUVECs dysfunction. In conclusion, circ_0002331 might be a potential target for AS treatment, which could improve ox-LDL-induced dysfunction of HUVECs via regulating CCND2 by binding with ELAVL1.
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Apoptosis , Aterosclerosis , Movimiento Celular , Proliferación Celular , Ciclina D2 , Proteína 1 Similar a ELAV , Células Endoteliales de la Vena Umbilical Humana , Lipoproteínas LDL , Estabilidad del ARN , ARN Circular , ARN Mensajero , Transducción de Señal , Humanos , ARN Circular/metabolismo , ARN Circular/genética , Lipoproteínas LDL/toxicidad , Lipoproteínas LDL/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/patología , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Ciclina D2/metabolismo , Ciclina D2/genética , Proteína 1 Similar a ELAV/metabolismo , Proteína 1 Similar a ELAV/genética , Células Cultivadas , Aterosclerosis/metabolismo , Aterosclerosis/patología , Aterosclerosis/genética , ARN Mensajero/metabolismo , ARN Mensajero/genética , Regulación de la Expresión Génica , Estudios de Casos y Controles , Autofagia , Masculino , Persona de Mediana Edad , FemeninoRESUMEN
RNA-binding proteins (RBPs) are essential for RNA metabolism and profoundly impact health and disease. The subcellular organization of RBP interaction networks with target RNAs remains largely unexplored. Here, we develop colocalization CLIP (coCLIP), a method that combines cross-linking and immunoprecipitation (CLIP) with proximity labeling, to explore in-depth the subcellular RNA interactions of the RBP human antigen R (HuR). Using this method, we uncover HuR's dynamic and location-specific interactions with RNA, revealing alterations in sequence preferences and interactions in the nucleus, cytosol, or stress granule (SG) compartments. We uncover HuR's unique binding preferences within SGs during arsenite stress, illuminating intricate interactions that conventional methodologies cannot capture. Overall, coCLIP provides a powerful method for revealing RBP-RNA interactions based on localization and lays the foundation for an advanced understanding of RBP models that incorporate subcellular location as a critical determinant of their functions.
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Unión Proteica , Proteínas de Unión al ARN , ARN , Humanos , ARN/metabolismo , ARN/genética , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Inmunoprecipitación/métodos , Proteína 1 Similar a ELAV/metabolismo , Proteína 1 Similar a ELAV/genética , Núcleo Celular/metabolismo , Núcleo Celular/genética , Gránulos Citoplasmáticos/metabolismo , Arsenitos , Células HeLa , Citosol/metabolismo , Células HEK293RESUMEN
The posttranslational modification of proteins critically influences many biological processes and is a key mechanism that regulates the function of the RNA-binding protein Hu antigen R (HuR), a hub in liver cancer. Here, we show that HuR is SUMOylated in the tumor sections of patients with hepatocellular carcinoma in contrast to the surrounding tissue, as well as in human cell line and mouse models of the disease. SUMOylation of HuR promotes major cancer hallmarks, namely proliferation and invasion, whereas the absence of HuR SUMOylation results in a senescent phenotype with dysfunctional mitochondria and endoplasmic reticulum. Mechanistically, SUMOylation induces a structural rearrangement of the RNA recognition motifs that modulates HuR binding affinity to its target RNAs, further modifying the transcriptomic profile toward hepatic tumor progression. Overall, SUMOylation constitutes a mechanism of HuR regulation that could be potentially exploited as a therapeutic strategy for liver cancer.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Animales , Humanos , Ratones , Carcinoma Hepatocelular/metabolismo , Modelos Animales de Enfermedad , Proteína 1 Similar a ELAV/metabolismo , Neoplasias Hepáticas/patología , ARN/metabolismo , SumoilaciónRESUMEN
Macroautophagy/autophagy-mediated anoikis resistance is crucial for tumor metastasis. As a key autophagy-related protein, ATG4B has been demonstrated to be a prospective anti-tumor target. However, the existing ATG4B inhibitors are still far from clinical application, especially for tumor metastasis. In this study, we identified a novel circRNA, circSPECC1, that interacted with ATG4B. CircSPECC1 facilitated liquid-liquid phase separation of ATG4B, which boosted the ubiquitination and degradation of ATG4B in gastric cancer (GC) cells. Thus, pharmacological addition of circSPECC1 may serve as an innovative approach to suppress autophagy by targeting ATG4B. Specifically, the circSPECC1 underwent significant m6A modification in GC cells and was subsequently recognized and suppressed by the m6A reader protein ELAVL1/HuR. The activation of the ELAVL1-circSPECC1-ATG4B pathway was demonstrated to mediate anoikis resistance in GC cells. Moreover, we also verified that the above pathway was closely related to metastasis in tissues from GC patients. Furthermore, we determined that the FDA-approved compound lopinavir efficiently enhanced anoikis and prevented metastasis by eliminating repression of ELAVL1 on circSPECC1. In summary, this study provides novel insights into ATG4B-mediated autophagy and introduces a viable clinical inhibitor of autophagy, which may be beneficial for the treatment of GC with metastasis.
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Anoicis , Autofagia , Cisteína Endopeptidasas , Lopinavir , ARN Circular , Anoicis/efectos de los fármacos , Autofagia/efectos de los fármacos , Humanos , ARN Circular/metabolismo , ARN Circular/genética , Línea Celular Tumoral , Cisteína Endopeptidasas/metabolismo , Lopinavir/farmacología , Neoplasias Gástricas/patología , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/tratamiento farmacológico , Neoplasias Gástricas/genética , Proteínas Relacionadas con la Autofagia/metabolismo , Animales , Ratones , Ubiquitinación/efectos de los fármacosRESUMEN
RNA-binding proteins (RBPs) play a crucial role in the regulation of posttranscriptional RNA networks, which can undergo dysregulation in many pathological conditions. Human antigen R (HuR) is a highly researched RBP that plays a crucial role as a posttranscriptional regulator. HuR plays a crucial role in the amplification of inflammatory signals by stabilizing the messenger RNA of diverse inflammatory mediators and key molecular players. The noteworthy correlations between HuR and its target molecules, coupled with the remarkable impacts reported on the pathogenesis and advancement of multiple diseases, position HuR as a promising candidate for therapeutic intervention in diverse inflammatory conditions. This review article examines the significance of HuR as a member of the RBP family, its regulatory mechanisms, and its implications in the pathophysiology of inflammation and cardiometabolic illnesses. Our objective is to illuminate potential directions for future research and drug development by conducting a comprehensive analysis of the existing body of research on HuR.
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Enfermedades Cardiovasculares , Proteína 1 Similar a ELAV , Inflamación , Humanos , Proteína 1 Similar a ELAV/metabolismo , Proteína 1 Similar a ELAV/genética , Inflamación/genética , Inflamación/patología , Enfermedades Cardiovasculares/genética , Enfermedades Cardiovasculares/inmunología , Enfermedades Cardiovasculares/metabolismo , Animales , Regulación de la Expresión Génica , Enfermedades Metabólicas/genética , Enfermedades Metabólicas/inmunología , Enfermedades Metabólicas/metabolismo , Transducción de Señal , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismoRESUMEN
Plasma microRNA (miR)-9 has been identified as a promising diagnostic biomarker for traumatic brain injury (TBI). This study aims to investigate the possible role and mechanisms of miR-9a-5p affecting TBI. Microarray-based gene expression profiling of TBI was used for screening differentially expressed miRNAs and genes. TBI rat models were established. miR-9a-5p, ELAVL1 and VEGF expression in the brain tissue of TBI rats was detected. The relationship among miR-9a-5p, ELAVL1 and VEGF was tested. TBI modeled rats were injected with miR-9a-5p-, ELAVL1 or VEGF-related sequences to identify their effects on TBI. miR-9a-5p was poorly expressed in the brain tissue of rats with TBI. ELAVL1 was a downstream target gene of miR-9a-5p, which could negatively regulate its expression. Enforced miR-9a-5p expression prevented brain tissue damage in TBI rats by targeting ELAVL1. Meanwhile, ELAVL1 could increase the expression of VEGF, which was highly expressed in the brain tissue of rats with TBI. In addition, ectopically expressed miR-9a-5p alleviated brain tissue damage in TBI rats by downregulating the ELAVL1/VEGF axis. Overall, miR-9a-5p can potentially reduce brain tissue damage in TBI rats by targeting ELAVL1 and down-regulating VEGF expression.
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Lesiones Traumáticas del Encéfalo , Lesiones Encefálicas , MicroARNs , Animales , Ratas , Lesiones Encefálicas/metabolismo , Lesiones Traumáticas del Encéfalo/genética , Perfilación de la Expresión Génica , MicroARNs/genética , MicroARNs/metabolismo , Factor A de Crecimiento Endotelial Vascular/genéticaRESUMEN
Breast cancer (BC) is the most prevalent malignant tumor in women worldwide with high morbidity and mortality. NSUN2, a crucial RNA methyltransferase, plays a pivotal role in regulating the proliferation and metastasis of tumor cells. Our study demonstrated that NSUN2 is upregulated in BC tissues and cell lines, and its high expression is associated with a poor prognosis in BC patients. Knockout of NSUN2 exerted inhibitory effects on the proliferation and migration of BC cells in vitro and in vivo. Mechanistic investigations revealed that the RNA-binding protein ELAVL1 can bind to NSUN2 mRNA and increase its stability. Additionally, we identified HOST2, a long non-coding RNA, as a key player in blocking the ubiquitin-dependent proteasomal degradation of ELAVL1, thereby influencing the stability of NSUN2 mRNA. In conclusion, this study revealed for the first time that HOST2 maintains NSUN2 mRNA stability by blocking ubiquitin-dependent degradation of ELAVL1, which in turn affects BC progression. HOST2/ELAVL1/NSUN2 oncogenic cascade has the potential to be a novel therapeutic target for BC.
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Neoplasias de la Mama , Proteína 1 Similar a ELAV , Metiltransferasas , ARN Largo no Codificante , Femenino , Humanos , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Proliferación Celular/genética , Proteína 1 Similar a ELAV/genética , Proteína 1 Similar a ELAV/metabolismo , Regulación Neoplásica de la Expresión Génica , Metiltransferasas/metabolismo , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , ARN Mensajero/genética , Ubiquitinas/metabolismoRESUMEN
Objective: As the most prevalent type of thyroid malignancy, papillary thyroid carcinoma (PTC) accounts for over 80% of all thyroid cancers. Circular RNAs (circRNAs) have been found to regulate multiple cancers, including PTC. Materials and methods: Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to analyse RNA and protein levels. Fluorescence in situ hybridization (FISH) was used to detect the distribution of the target genes. Functional experiments and animal experiments were implemented to analyse the biological functions of target genes in vitro and in vivo. Luciferase reporter, RNA pulldown, RNA binding protein immunoprecipitation (RIP) and mRNA stability assays were used to probe the underlying mechanisms. Results: CircSEMA6Awas found to be upregulated in PTC tissues and cells, and its circular structure was verified. CircSEMA6A promotes PTC cell migration and invasion. Moreover, circSEMA6A functions as a competing endogenous RNA (ceRNA) to upregulate proline rich and Gla domain 4 (PRRG4) expression by sponging microRNA-520h (miR-520h). CircSEMA6A recruits ELAV1 to stabilize PRRG4 mRNA and drives PTC progression via PRRG4. Conclusion: CircSEMA6A upregulates PRRG4 by targeting miR-520h and recruiting ELAVL1 to affect the invasion and migration of PTC cells, offering insight into the molecular mechanisms of PTC.
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MicroARNs , Neoplasias de la Tiroides , Animales , Cáncer Papilar Tiroideo/genética , Cáncer Papilar Tiroideo/patología , Hibridación Fluorescente in Situ , Proliferación Celular/genética , Línea Celular Tumoral , MicroARNs/genética , MicroARNs/metabolismo , Neoplasias de la Tiroides/patologíaRESUMEN
YPEL2 is a member of the evolutionarily conserved YPEL family involved in cellular proliferation, mobility, differentiation, senescence, and death. However, the mechanism by which YPEL2, or YPEL proteins, mediates its effects is largely unknown. Proteins perform their functions in a network of proteins whose identities, amounts, and compositions change spatiotemporally in a lineage-specific manner in response to internal and external stimuli. Here, we explored interaction partners of YPEL2 by using dynamic TurboID-coupled mass spectrometry analyses to infer a function for the protein. Our results using inducible transgene expressions in COS7 cells indicate that proximity interaction partners of YPEL2 are mainly involved in RNA and mRNA metabolic processes, ribonucleoprotein complex biogenesis, regulation of gene silencing by miRNA, and cellular responses to stress. We showed that YPEL2 interacts with the RNA-binding protein ELAVL1 and the selective autophagy receptor SQSTM1. We also found that YPEL2 localizes stress granules in response to sodium arsenite, an oxidative stress inducer, which suggests that YPEL2 participates in stress granule-related processes. Establishing a point of departure in the delineation of structural/functional features of YPEL2, our results suggest that YPEL2 may be involved in stress surveillance mechanisms.
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Estrés Oxidativo , Proteínas de Unión al ARN , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismoRESUMEN
ABSTRACT Objective: As the most prevalent type of thyroid malignancy, papillary thyroid carcinoma (PTC) accounts for over 80% of all thyroid cancers. Circular RNAs (circRNAs) have been found to regulate multiple cancers, including PTC. Materials and methods: Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to analyse RNA and protein levels. Fluorescence in situ hybridization (FISH) was used to detect the distribution of the target genes. Functional experiments and animal experiments were implemented to analyse the biological functions of target genes in vitro and in vivo. Luciferase reporter, RNA pulldown, RNA binding protein immunoprecipitation (RIP) and mRNA stability assays were used to probe the underlying mechanisms. Results: CircSEMA6Awas found to be upregulated in PTC tissues and cells, and its circular structure was verified. CircSEMA6A promotes PTC cell migration and invasion. Moreover, circSEMA6A functions as a competing endogenous RNA (ceRNA) to upregulate proline rich and Gla domain 4 (PRRG4) expression by sponging microRNA-520h (miR-520h). CircSEMA6A recruits ELAV1 to stabilize PRRG4 mRNA and drives PTC progression via PRRG4. Conclusion: CircSEMA6A upregulates PRRG4 by targeting miR-520h and recruiting ELAVL1 to affect the invasion and migration of PTC cells, offering insight into the molecular mechanisms of PTC.
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The aryl hydrocarbon receptor (AHR) is a transcription factor that is commonly upregulated in pancreatic ductal adenocarcinoma (PDAC). AHR hinders the shuttling of human antigen R (ELAVL1) from the nucleus to the cytoplasm, where it stabilises its target messenger RNAs (mRNAs) and enhances protein expression. Among these target mRNAs are those induced by gemcitabine. Increased AHR expression leads to the sequestration of ELAVL1 in the nucleus, resulting in chemoresistance. This study aimed to investigate the interaction between AHR and ELAVL1 in the pathogenesis of PDAC in vitro. AHR and ELAVL1 genes were silenced by siRNA transfection. The RNA and protein were extracted for quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot (WB) analysis. Direct binding between the ELAVL1 protein and AHR mRNA was examined through immunoprecipitation (IP) assay. Cell viability, clonogenicity, and migration assays were performed. Our study revealed that both AHR and ELAVL1 inter-regulate each other, while also having a role in cell proliferation, migration, and chemoresistance in PDAC cell lines. Notably, both proteins function through distinct mechanisms. The silencing of ELAVL1 disrupts the stability of its target mRNAs, resulting in the decreased expression of numerous cytoprotective proteins. In contrast, the silencing of AHR diminishes cell migration and proliferation and enhances cell sensitivity to gemcitabine through the AHR-ELAVL1-deoxycytidine kinase (DCK) molecular pathway. In conclusion, AHR and ELAVL1 interaction can form a negative feedback loop. By inhibiting AHR expression, PDAC cells become more susceptible to gemcitabine through the ELAVL1-DCK pathway.
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Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Carcinoma Ductal Pancreático/tratamiento farmacológico , Carcinoma Ductal Pancreático/genética , Proteína 1 Similar a ELAV/genética , Gemcitabina , Páncreas , Hormonas Pancreáticas , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Receptores de Hidrocarburo de Aril/genética , ARN Mensajero/genética , Desoxicitidina Quinasa/efectos de los fármacos , Desoxicitidina Quinasa/metabolismo , Neoplasias PancreáticasRESUMEN
Long noncoding RNAs (lncRNAs) modulate many aspects of biological and pathological processes. Recent studies have shown that host lncRNAs participate in the antiviral immune response, but functional lncRNAs in coxsackievirus B5 (CVB5) infection remain unknown. Here, we identified a novel cytoplasmic lncRNA, LINC1392, which was highly inducible in CVB5 infected RD cells in a time- and dose-dependent manner, and also can be induced by the viral RNA and IFN-ß. Further investigation showed that LINC1392 promoted several important interferon-stimulated genes (ISGs) expression, including IFIT1, IFIT2, and IFITM3 by activating MDA5, thereby inhibiting the replication of CVB5 in vitro. Mechanistically, LINC1392 bound to ELAV like RNA binding protein 1 (ELAVL1) and blocked ELAVL1 interaction with MDA5. Functional study revealed that the 245-835 ânt locus of LINC1392 exerted the antiviral effect and was also an important site for ELAVL1 binding. In mice, LINC1392 could inhibit CVB5 replication and alleviated the histopathological lesions of intestinal and brain tissues induced by viral infection. Our findings collectively reveal that the novel LINC1392 acts as a positive regulator in the IFN-I signaling pathway against CVB5 infection. Elucidating the underlying mechanisms on how lncRNA regulats the host innate immunity response towards CVB5 infection will lay the foundation for antiviral drug research.
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Interferón Tipo I , ARN Largo no Codificante , Animales , Ratones , Enterovirus Humano B/genética , Interacciones Huésped-Patógeno/genética , Inmunidad Innata , Interferón Tipo I/genética , ARN Largo no Codificante/genética , Transducción de Señal/genéticaRESUMEN
An abnormal immune response induces melanoma development. IL-17 and the classical downstream signal STAT1 are associated with melanoma development. TRAF2 also mediates the downstream signaling of IL-17; however, its role in IL-17-stimulated melanoma remains unclear. Bioinformatic analysis revealed that TRAF2 can bind to PIAS2 (a SUMO E3 ligase), ELAVL1 (an RNA-binding protein), and EPHA5 (an ephrin receptor of the tyrosine kinase family). To elucidate the IL-17 downstream signal, the IL-17 receptor (R), STAT1, TRAF2, PIAS2, ELAVL1, and EPHA5 were knocked down before melanoma cells were treated with recombinant IL-17A protein. Co-immunoprecipitation and RNA immunoprecipitation were conducted to determine the interaction of TRAF2 with PIAS2, ELAVL1, and EPHA5 proteins, as well as the interaction of ELAVL1 protein with EPHA5 mRNA. STAT1 knockdown suppressed the proliferation and invasion triggered by IL-17A, but the suppressive effects were much weaker than those caused by IL-17R knockdown. This implies that another nonclassical signal mediates IL-17 effects. IL-17A induces TRAF2 recruitment of ELAVL1, PIAS2, and EPHA5 proteins. We speculated that ELAVL1 bound to the AU-rich elements in the 3' untranslated region of the EPHA5 mRNA, thereby enhancing mRNA stability. Furthermore, PIAS2 induced EPHA5 SUMOylation, which suppressed EPHA5 ubiquitination and degradation. Through pre- and post-translational regulation, IL-17A induced EPHA5 expression in melanoma, and EPHA5 knockdown markedly suppressed IL-17A-induced proliferation and invasion. This study revealed a non-classical signaling mechanism responsible for the effects of IL-17 in melanoma.
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OBJECTIVE: To investigate the regulatory role of the long non-coding RNA LINC00926 in pyroptosis of hypoxia-induced human umbilical vein vascular endothelial cells (HUVECs) and explore the molecular mechanism. METHODS: HUVECs were transfected with a LINC00926-overexpressing plasmid (OE-LINC00926), a siRNA targeting ELAVL1, or both, followed by exposure to hypoxia (5% O2) or normoxia. The expression of LINC00926 and ELAVL1 in hypoxia-treated HUVECs was detected using real-time quantitative PCR (RT-qPCR) and Western blotting. Cell proliferation was detected using Cell Counting Kit-8 (CCK-8), and the levels of IL-1ß in the cell cultures was determined with ELISA. The protein expression levels of pyroptosis-related proteins (caspase-1, cleaved caspase-1 and NLRP3) in the treated cells were analyzed using Western blotting, and the binding between LINC00926 and ELAVL1 was verified with RNA immunoprecipitation (RIP) assay. RESULTS: Exposure to hypoxia obviously up-regulated the mRNA expression of LINC00926 and the protein expression of ELAVL1 in HUVECs, but did not affect the mRNA expression of ELAVL1. LINC00926 overexpression in the cells significantly inhibited cell proliferation, increased IL-1ß level and enhanced the expressions of pyroptosis-related proteins (all P < 0.05). LINC00926 overexpression further up-regulated the protein expression of ELAVL1 in hypoxia-exposed HUVECs. The results of RIP assay confirmed the binding between LINC00926 and ELAVL1. ELAVL1 knockdown significantly decreased IL-1ß level and the expressions of pyroptosis-related proteins in hypoxia-exposed HUVECs (P < 0.05), while LINC00926 overexpression partially reversed the effects of ELAVL1 knockdown. CONCLUSION: LINC00926 promotes pyroptosis of hypoxia-induced HUVECs by recruiting ELAVL1.
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Hipoxia de la Célula , Proteína 1 Similar a ELAV , Piroptosis , ARN Largo no Codificante , Humanos , Caspasa 1 , Células Endoteliales de la Vena Umbilical Humana , ARN Mensajero , ARN Largo no Codificante/genéticaRESUMEN
Sepsis is a life-threatening systemic organ dysfunction caused by the host's unregulated response to a widespread bacterial infection. Endothelial injury is a major pathophysiologic symptom of sepsis and is considered a critical factor in promoting the progression of disease severity. ELAV like RNA binding protein 1(ELAVL1) is a ubiquitously expressed RNA-binding protein that may play an important role during sepsis. Nonetheless, the molecular mechanisms of ELAVL1 on endothelial cell damage in sepsis have not been well defined. Here, we aimed to confirm the role of ELAVL1 in sepsis-induced endothelial cell damage using lipopolysaccharide (LPS)-induced zebrafish and endothelial cells (ECs) models. We found that zebrafish larvae treated with LPS exhibited systemic endothelial cell damage, mostly manifested as pericardial edema, curved tail, and impaired angiogenesis. LPS treatments also significantly induced the expression levels of inflammatory cytokines (interleukin-6 (IL-6), IL-8, and tumor necrosis factor (TNF)-α) in vivo. In vitro, we observed the increase of ELAVL1 cytoplasmic translocation with LPS treatment. Mechanistically, targeted disruption of the ELAVL1 gene decreased the expression of TNF-α, IL-6, and IL-8 during induction of sepsis and alleviated LPS-induced blood vessel injury in zebrafish. Taken together, our study indicates that ELAVL1 knockdown may alleviate sepsis-induced endothelial cells injury by suppressing cytokine storm. Our research suggests that inhibition of ELAVL1 could reduce the level of inflammatory cytokine production induced by LPS and protect against endothelial cell injury. ELAVL1 might be a potential therapeutic target to block endothelial cells injury associated with sepsis.
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Interleucina-6 , Sepsis , Animales , Interleucina-6/metabolismo , Lipopolisacáridos/farmacología , Células Endoteliales/metabolismo , Pez Cebra/metabolismo , Síndrome de Liberación de Citoquinas/metabolismo , Interleucina-8/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Sepsis/tratamiento farmacológicoRESUMEN
Triple-negative breast cancer is high heterogeneous, aggressive, and metastatic with poor prognosis. Despite of advances in targeted therapies, TNBC has been reported to cause high morbidity and mortality. A rare subpopulation within the tumor microenvironment organized into a hierarchy of cancer stem cells is responsible for therapy resistance and tumor recurrence. Repurposing of antiviral drugs for cancer treatment is gaining momentum due to reduced cost, labour, and research time, but limited due to lack of prognostic, and predictive markers. The present study investigates proteomic profiling and ROC analysis to identify CD151 and ELAVL1 as potential therapy response markers for the antiviral drug 2-thio-6-azauridine (TAU) in resistant TNBC. The stemness of MDA-MB 231 and MDA-MD 468 adherent cells was enriched by culturing them under non-adherent and non-differentiation conditions. Then, CD151+ subpopulation was isolated and characterized for the enrichment of stemness. This study found that CD151 has overexpressed in stemness enriched subpopulations, and also showed CD44 high and CD24 low expression along with stem cell-related transcription factors octamer-binding transcription factor 4 (OCT4) and Sex determining Y-box 2 (SOX2). This study also found that TAU induced significant cytotoxicity and genotoxicity in the CD151+TNBC subpopulation and inhibited their proliferation by inducing DNA damage, cell cycle arrest at the G2M phase, and apoptosis. Further, a proteomic profiling study showed that the expression of CD151 along with ELAVL1, an RNA-binding protein, was significantly reduced with TAU treatment. KM plotter showed correlation of CD151 and ELAVL1 gene expression with a poor prognosis of TNBC. ROC analysis predicted and validated CD151 and ELAVL1 as best therapy response marker for TAU in TNBC. These findings provide new insight into repurposing antiviral drug TAU for treatment of metastatic and drug resistant TNBC.
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Neoplasias de la Mama Triple Negativas , Humanos , Línea Celular Tumoral , Neoplasias de la Mama Triple Negativas/patología , Curva ROC , Proteómica , Recurrencia Local de Neoplasia , Proliferación Celular , Microambiente Tumoral , Tetraspanina 24/metabolismo , Proteína 1 Similar a ELAVRESUMEN
Lipopolysaccharide (LPS) exposure to macrophages induces an inflammatory response, which is regulated at the transcriptional and post-transcriptional levels. HuR (ELAVL1) is an RNA-binding protein that regulates cytokines and chemokines transcripts containing AU/U-rich elements (AREs) and mediates the LPS-induced response. Here, we show that small-molecule tanshinone mimics (TMs) inhibiting HuR-RNA interaction counteract LPS stimulus in macrophages. TMs exist in solution in keto-enolic tautomerism, and molecular dynamic calculations showed the ortho-quinone form inhibiting binding of HuR to mRNA targets. TM activity was lost in vitro by blocking the diphenolic reduced form as a diacetate, but resulted in prodrug-like activity in vivo. RNA and ribonucleoprotein immunoprecipitation sequencing revealed that LPS induces a strong coupling between differentially expressed genes and HuR-bound genes, and TMs reduced such interactions. TMs decreased the association of HuR with genes involved in chemotaxis and immune response, including Cxcl10, Il1b and Cd40, reducing their expression and protein secretion in primary murine bone marrow-derived macrophages and in an LPS-induced peritonitis model. Overall, TMs show anti-inflammatory properties in vivo and suggest HuR as a potential therapeutic target for inflammation-related diseases.
Asunto(s)
Proteína 1 Similar a ELAV , Lipopolisacáridos , Ratones , Animales , Lipopolisacáridos/farmacología , Lipopolisacáridos/metabolismo , Proteína 1 Similar a ELAV/genética , Proteína 1 Similar a ELAV/metabolismo , Macrófagos/metabolismo , ARN/metabolismo , ARN Mensajero/genéticaRESUMEN
Introduction: ELAVL1/HuR is a keystone regulator of gene expression at the posttranscriptional level, including stress response and homeostasis maintenance. The aim of this study was to evaluate the impact of hur silencing on the age-related degeneration of retinal ganglion cells (RGC), which potentially describes the efficiency of endogenous neuroprotection mechanisms, as well as to assess the exogenous neuroprotection capacity of hur-silenced RGC in the rat glaucoma model. Methods: The study consisted of in vitro and in vivo approaches. In vitro, we used rat B-35 cells to investigate, whether AAV-shRNA-HuR delivery affects survival and oxidative stress markers under temperature and excitotoxic insults. In vivo approach consisted of two different settings. In first one, 35 eight-week-old rats received intravitreal injection of AAV-shRNA-HuR or AAV-shRNA scramble control. Animals underwent electroretinography tests and were sacrificed 2, 4 or 6 months after injection. Retinas and optic nerves were collected and processed for immunostainings, electron microscopy and stereology. For the second approach, animals received similar gene constructs. To induce chronic glaucoma, 8 weeks after AAV injection, unilateral episcleral vein cauterization was performed. Animals from each group received intravitreal injection of metallothionein II. Animals underwent electroretinography tests and were sacrificed 8 weeks later. Retinas and optic nerves were collected and processed for immunostainings, electron microscopy and stereology. Results: Silencing of hur induced apoptosis and increased oxidative stress markers in B-35 cells. Additionally, shRNA treatment impaired the cellular stress response to temperature and excitotoxic insults. In vivo, RGC count was decreased by 39% in shRNA-HuR group 6 months after injection, when compared to shRNA scramble control group. In neuroprotection study, the average loss of RGCs was 35% in animals with glaucoma treated with metallothionein and shRNA-HuR and 11.4% in animals with glaucoma treated with metallothionein and the scramble control shRNA. An alteration in HuR cellular content resulted in diminished photopic negative responses in the electroretinogram. Conclusions: Based on our findings, we conclude that HuR is essential for the survival and efficient neuroprotection of RGC and that the induced alteration in HuR content accelerates both the age-related and glaucoma-induced decline in RGC number and function, further confirming HuR's key role in maintaining cell homeostasis and its possible involvement in the pathogenesis of glaucoma.