RESUMO
Pescadillo ribosomal biogenesis factor 1 (PES1), a nucleolar protein initially identified in zebrafish, plays an important role in embryonic development and ribosomal biogenesis. Notably, PES1 has been found to be overexpressed in a number of cancer types, where it contributes to tumorigenesis and cancer progression by promoting cell proliferation, suppressing cellular senescence, modulating the tumor microenvironment (TME) and promoting drug resistance in cancer cells. Moreover, recent emerging evidence suggests that PES1 expression is significantly elevated in the livers of Type 2 diabetes mellitus (T2DM) and obese patients, indicating its involvement in the pathogenesis of metabolic diseases through lipid metabolism regulation. In this review, we present the structural characteristics and biological functions of PES1, as well as complexes in which PES1 participates. Furthermore, we comprehensively summarize the multifaceted role of PES1 in various diseases and the latest insights into its underlying molecular mechanisms. Finally, we discuss the potential clinical translational perspectives of targeting PES1, highlighting its promising as a therapeutic intervention and treatment target.
Assuntos
Neoplasias , Proteínas de Ligação a RNA , Humanos , Animais , Neoplasias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/tratamento farmacológico , Microambiente Tumoral , Metabolismo dos Lipídeos , Terapia de Alvo Molecular/métodos , Obesidade/metabolismo , Obesidade/genéticaRESUMO
Previously, we showed that the germ cell-specific nuclear protein RBMXL2 represses cryptic splicing patterns during meiosis and is required for male fertility (Ehrmann et al., 2019). Here, we show that in somatic cells the similar yet ubiquitously expressed RBMX protein has similar functions. RBMX regulates a distinct class of exons that exceed the median human exon size. RBMX protein-RNA interactions are enriched within ultra-long exons, particularly within genes involved in genome stability, and repress the selection of cryptic splice sites that would compromise gene function. The RBMX gene is silenced during male meiosis due to sex chromosome inactivation. To test whether RBMXL2 might replace the function of RBMX during meiosis we induced expression of RBMXL2 and the more distantly related RBMY protein in somatic cells, finding each could rescue aberrant patterns of RNA processing caused by RBMX depletion. The C-terminal disordered domain of RBMXL2 is sufficient to rescue proper splicing control after RBMX depletion. Our data indicate that RBMX and RBMXL2 have parallel roles in somatic tissues and the germline that must have been conserved for at least 200 million years of mammalian evolution. We propose RBMX family proteins are particularly important for the splicing inclusion of some ultra-long exons with increased intrinsic susceptibility to cryptic splice site selection.
Assuntos
Éxons , Sítios de Splice de RNA , Splicing de RNA , Proteínas de Ligação a RNA , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Humanos , Éxons/genética , Sítios de Splice de RNA/genética , Masculino , Meiose/genética , Animais , Ribonucleoproteínas Nucleares HeterogêneasRESUMO
BACKGROUND: Neuroinflammation plays a critical role in various neurological disorders. Oxycodone has anti-inflammatory properties. The purpose of this work was to look into the effect of oxycodone in controlling lipopolysaccharide (LPS)-induced neuroinflammation in microglia. METHODS: LPS-induced HMC3 cells were subjected to oxycodone (2.5, 5, 10 and 20 µg/mL). The mRNA and protein expressions were examined by qRT-PCR and western blotting. TNF-α, IL-1ß, IL-6, and IL-8 levels were assessed by ELISA. MTT assay was adopted to measure cell viability. The interactions between CREB, miR-181c and PDCD4 were analyzed by dual-luciferase reporter assay, ChIP and/or RIP assays. RESULTS: Oxycodone treatment alleviated LPS-induced inflammation in HMC3 cells and increased p-CREB level, but reduced PDCD4 and iNOS levels in LPS-treated cells. Mechanistically, oxycodone mitigated LPS-induced neuroinflammation by upregulating miR-181c. In addition, CREB promoted miR-181c expression by directly binding to the MIR181C promoter, and miR-181c inhibited PDCD4 expression by directly binding to PDCD4 3'UTR. As expected, oxycodone alleviated LPS-induced neuroinflammation by regulating the CREB/miR-181c/PDCD4 axis. CONCLUSION: Oxycodone attenuated LPS-induced neuroinflammation in microglia by regulating the CREB/miR-181c/PDCD4 axis. These findings proved that oxycodone is a potential drug for treating neuroinflammation and elucidate the mechanisms involved.
Assuntos
Proteínas Reguladoras de Apoptose , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico , Lipopolissacarídeos , MicroRNAs , Microglia , Doenças Neuroinflamatórias , Oxicodona , Proteínas de Ligação a RNA , MicroRNAs/genética , MicroRNAs/metabolismo , Oxicodona/farmacologia , Oxicodona/efeitos adversos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Humanos , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas Reguladoras de Apoptose/genética , Microglia/efeitos dos fármacos , Microglia/metabolismo , Doenças Neuroinflamatórias/induzido quimicamente , Doenças Neuroinflamatórias/tratamento farmacológico , Doenças Neuroinflamatórias/genética , Anti-Inflamatórios/farmacologia , Linhagem Celular , Inflamação/induzido quimicamente , Inflamação/genética , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND: Somatic copy number alterations (SCNAs) are pivotal in cancer progression and patient prognosis. Dysregulated long non-coding RNAs (lncRNAs), modulated by SCNAs, significantly impact tumorigenesis, including colorectal cancer (CRC). Nonetheless, the functional significance of lncRNAs induced by SCNAs in CRC remains largely unexplored. METHODS: The dysregulated lncRNA LOC101927668, induced by copy number amplification, was identified through comprehensive bioinformatic analyses utilizing multidimensional data. Subsequent in situ hybridization was employed to ascertain the subcellular localization of LOC101927668, and gain- and loss-of-function experiments were conducted to elucidate its role in CRC progression. The downstream targets and signaling pathway influenced by LOC101927668 were identified and validated through a comprehensive approach, encompassing RNA sequencing, RT-qPCR, Western blot analysis, dual-luciferase reporter assay, evaluation of mRNA and protein degradation, and rescue experiments. Analysis of AU-rich elements (AREs) within the mRNA 3' untranslated region (UTR) of the downstream target, along with exploration of putative ARE-binding proteins, was conducted. RNA pull-down, mass spectrometry, RNA immunoprecipitation, and dual-luciferase reporter assays were employed to elucidate potential interacting proteins of LOC101927668 and further delineate the regulatory mechanism between LOC101927668 and its downstream target. Moreover, subcutaneous xenograft and orthotopic liver xenograft tumor models were utilized to evaluate the in vivo impact of LOC101927668 on CRC cells and investigate its correlation with downstream targets. RESULTS: Significantly overexpressed LOC101927668, driven by chr7p22.3-p14.3 amplification, was markedly correlated with unfavorable clinical outcomes in our CRC patient cohort, as well as in TCGA and GEO datasets. Moreover, we demonstrated that enforced expression of LOC101927668 significantly enhanced cell proliferation, migration, and invasion, while its depletion impeded these processes in a p53-dependent manner. Mechanistically, nucleus-localized LOC101927668 recruited hnRNPD and translocated to the cytoplasm, accelerating the destabilization of RBM47 mRNA, a transcription factor of p53. As a nucleocytoplasmic shuttling protein, hnRNPD mediated RBM47 destabilization by binding to the ARE motif within RBM47 3'UTR, thereby suppressing the p53 signaling pathway and facilitating CRC progression. CONCLUSIONS: The overexpression of LOC101927668, driven by SCNAs, facilitates CRC proliferation and metastasis by recruiting hnRNPD, thus perturbing the RBM47/p53/p21 signaling pathway. These findings underscore the pivotal roles of LOC101927668 and highlight its therapeutic potential in anti-CRC interventions.
Assuntos
Neoplasias Colorretais , Progressão da Doença , RNA Longo não Codificante , Transdução de Sinais , Proteína Supressora de Tumor p53 , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Camundongos , Animais , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Proliferação de Células , Feminino , Linhagem Celular Tumoral , Variações do Número de Cópias de DNA , Masculino , Regulação Neoplásica da Expressão Gênica , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/genética , Camundongos NusRESUMO
TRIM25 is an RNA-binding ubiquitin E3 ligase with central but poorly understood roles in the innate immune response to RNA viruses. The link between TRIM25's RNA binding and its role in innate immunity has not been established. Thus, we utilized a multitude of biophysical techniques to identify key RNA-binding residues of TRIM25 and developed an RNA-binding deficient mutant (TRIM25-m9). Using iCLIP2 in virus-infected and uninfected cells, we identified TRIM25's RNA sequence and structure specificity, that it binds specifically to viral RNA, and that the interaction with RNA is critical for its antiviral activity.
Assuntos
Ligação Proteica , RNA Viral , Proteínas com Motivo Tripartido , Ubiquitina-Proteína Ligases , Humanos , Proteínas com Motivo Tripartido/metabolismo , Proteínas com Motivo Tripartido/genética , RNA Viral/metabolismo , RNA Viral/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Células HEK293 , Imunidade Inata , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Antivirais/metabolismo , Antivirais/farmacologia , Vírus de RNA/genética , Sítios de LigaçãoRESUMO
Fibrosis is an abnormal tissue healing process characterized by the excessive accumulation of ECM components, such as COL I and COL III, in response to tissue injury or chronic inflammation. Recent advances in epitranscriptomics have underscored the importance of m6A modification in fibrosis. m6A, the most prevalent modification in eukaryotic RNA, is catalyzed by methyltransferases (e.g., METTL3), removed by demethylases (e.g., FTO), and recognized by reader proteins (e.g., YTHDF1/2). These modifications are crucial in regulating collagen metabolism and associated diseases. Understanding the role of m6A modification in fibrosis and other collagen-related conditions holds promise for developing targeted therapies. This review highlights the latest progress in this area.
Assuntos
Adenosina , Fibrose , Metiltransferases , Humanos , Adenosina/análogos & derivados , Adenosina/genética , Adenosina/metabolismo , Fibrose/genética , Metiltransferases/genética , Epigênese Genética/genética , Doenças do Colágeno/genética , Animais , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Colágeno/genética , Colágeno/metabolismo , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/metabolismo , RNA/genéticaRESUMO
In our cells, a limited number of RNA binding proteins (RBPs) are responsible for all aspects of RNA metabolism across the entire transcriptome. To accomplish this, RBPs form regulatory units that act on specific target regulons. However, the landscape of RBP combinatorial interactions remains poorly explored. Here, we perform a systematic annotation of RBP combinatorial interactions via multimodal data integration. We build a large-scale map of RBP protein neighborhoods by generating in vivo proximity-dependent biotinylation datasets of 50 human RBPs. In parallel, we use CRISPR interference with single-cell readout to capture transcriptomic changes upon RBP knockdowns. By combining these physical and functional interaction readouts, along with the atlas of RBP mRNA targets from eCLIP assays, we generate an integrated map of functional RBP interactions. We then use this map to match RBPs to their context-specific functions and validate the predicted functions biochemically for four RBPs. This study provides a detailed map of RBP interactions and deconvolves them into distinct regulatory modules with annotated functions and target regulons. This multimodal and integrative framework provides a principled approach for studying post-transcriptional regulatory processes and enriches our understanding of their underlying mechanisms.
Assuntos
RNA Mensageiro , Proteínas de Ligação a RNA , Humanos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , Transcriptoma , Processamento Pós-Transcricional do RNA , Regulação da Expressão Gênica , Células HEK293 , Análise de Célula Única , Redes Reguladoras de Genes , Regulon/genéticaRESUMO
PURPOSE: The current study aims to investigate the significance of N6-methyladenosine (m6A) methylationrelated genes in the clinical prognosis of childhood relapsed B-cell acute lymphoblastic leukemia (B-ALLL) patient. METHODS: Transcriptome data and corresponding clinical data on m6A methylation-related genes (including 20 genes) were obtained from the Therapeutically Applicable Research To Generate Effective Treatments (TARGET) database. RESULTS: The bone marrow (BM) samples of 134 newly diagnosed (naive) and 116 relapsed B-ALL from TARGET were enrolled in the current study. Three genes (FTO, HNRNPC, RBM15B) showed significant up-regulation in relapsed B-ALL compared with that in naive B-ALL.The three genes had a significantly worse survival (P < 0.05). The LASSO Cox regression model was used to select the most predictive genes as prognostic indicators, and YTHDC1 and FTO were identified as prognostic factors for relapsed B-ALL. Finally, the results of multivariate regression analysis showed that the risk score of m6A methylation-related genes was an independent prognostic factor in relapsed B-ALL (P < 0.05). CONCLUSION: We found that the expression levels of m6A methylation-related genes were different in naive and relapsed patients with B-ALL and correlated with survival and prognosis.This implies that m6A methylation-related genes may be promising prognostic indicators or therapeutic targets for relapsed B-ALL.
Assuntos
Adenosina , Dioxigenase FTO Dependente de alfa-Cetoglutarato , Leucemia-Linfoma Linfoblástico de Células Precursoras B , Fatores de Processamento de RNA , Proteínas de Ligação a RNA , Humanos , Prognóstico , Adenosina/análogos & derivados , Adenosina/genética , Criança , Feminino , Masculino , Proteínas de Ligação a RNA/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/mortalidade , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Fatores de Processamento de RNA/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo C/genética , Metilação , Pré-Escolar , Transcriptoma , Regulação para Cima , Biomarcadores Tumorais/genética , Recidiva , Recidiva Local de Neoplasia/genética , Adolescente , Proteínas do Tecido NervosoRESUMO
Pathological cardiac hypertrophy (CH) may lead to heart failure and sudden death. MicroRNAs (miRNAs) have been documented to play crucial parts in CH. The objective of this research was to discuss the potential along with molecule mechanism of miR-495-3p in CH. In vivo CH model was induced by aortic banding (AB) in rats. Cellular hypertrophy in H9c2 rat cardiomyocytes was stimulated by angiotensin II (Ang II) treatment. Haematoxylin and eosin (HE), echocardiography and immunofluorescence staining were used to examine the alterations in cardiac function. The outcomes showed that miR-495-3p expression was high in rat model as well as in Ang II-stimulated cardiomyocytes. Besides, silenced miR-495-3p attenuated CH both in vitro and in vivo. Mechanically, miR-495-3p bound to pumilio RNA binding family member 2 (Pum2) 3'UTR and silenced its expression. Rescue assays further notarized that Pum2 silence abrogated the inhibitory impacts of miR-495-3p inhibitor on CH. In a word, the present research uncovered that miR-495-3p promoted CH by targeting Pum2. Therefore, miR-495-3p may be a novel therapeutic molecule for this disease.
Assuntos
Angiotensina II , Cardiomegalia , MicroRNAs , Miócitos Cardíacos , Proteínas de Ligação a RNA , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Cardiomegalia/genética , Cardiomegalia/patologia , Cardiomegalia/metabolismo , Ratos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Angiotensina II/farmacologia , Masculino , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Ratos Sprague-Dawley , Linhagem Celular , Regiões 3' não Traduzidas/genética , Modelos Animais de Doenças , Sequência de BasesRESUMO
T cell receptor (TCR) sensitivity to peptide-major histocompatibility complex (MHC) dictates T cell fate. Canonical models of TCR sensitivity cannot be fully explained by transcriptional regulation. In this work, we identify a posttranscriptional regulatory mechanism of TCR sensitivity that guides alternative splicing of TCR signaling transcripts through an evolutionarily ultraconserved poison exon (PE) in the RNA-binding protein (RBP) TRA2ß in mouse and human. TRA2ß-PE splicing, seen during cancer and infection, was required for TCR-induced effector T cell expansion and function. Tra2ß-PE skipping enhanced T cell response to antigen by increasing TCR sensitivity. As antigen levels decreased, Tra2ß-PE reinclusion allowed T cell survival. Finally, we found that TRA2ß-PE was first included in the genome of jawed vertebrates that were capable of TCR gene rearrangements. We propose that TRA2ß-PE splicing acts as a gatekeeper of TCR sensitivity to shape T cell fate.
Assuntos
Processamento Alternativo , Éxons , Receptores de Antígenos de Linfócitos T , Fatores de Processamento de Serina-Arginina , Animais , Humanos , Camundongos , Sobrevivência Celular , Sequência Conservada , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Fatores de Processamento de Serina-Arginina/genética , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
BACKGROUND: Vasculogenic mimicry (VM) is a potential cause of resistance to antiangiogenic therapy and is closely related to the malignant progression of tumors. It has been shown that noncoding RNAs play an important role in the formation of VM in malignant tumors. However, the role of circRNAs in VM of bladder cancer and the regulatory mechanisms are unclear. METHODS: Firstly, hsa_circ_0000520 was identified to have circular character by Sanger sequencing and Rnase R assays. Secondly, the potential clinical value of hsa_circ_0000520 was explored by quantitative real-time polymerase chain reaction (qRT-PCR) and fluorescence in situ hybridization (FISH) of clinical specimens. Thirdly, the role of hsa_circ_0000520 in bladder cancer invasion, migration, and VM formation was examined by in vivo and in vitro experiments. Finally, the regulatory mechanisms of hsa_circ_0000520 in the malignant progression of bladder cancer were elucidated by RNA binding protein immunoprecipitation (RIP), RNA pulldown, co-immunoprecipitation (co-IP), qRT-PCR, Western blot (WB), and fluorescence co-localization. RESULTS: Hsa_circ_0000520 was characterized as a circular RNA and was lowly expressed in bladder cancer compared with the paracancer. Bladder cancer patients with high expression of hsa_circ_0000520 had better survival prognosis. Functionally, hsa_circ_0000520 inhibited bladder cancer invasion, migration, and VM formation. Mechanistically, hsa_circ_0000520 acted as a scaffold to promote binding of UBE2V1/UBC13 to Lin28a, further promoting the ubiquitous degradation of Lin28a, improving PTEN mRNA stability, and inhibiting the phosphorylation of the PI3K/AKT pathway. The formation of hsa_circ_0000520 in bladder cancer was regulated by RNA binding protein QKI. CONCLUSIONS: Hsa_circ_0000520 inhibits metastasis and VM formation in bladder cancer and is a potential target for bladder cancer diagnosis and treatment.
Assuntos
Movimento Celular , PTEN Fosfo-Hidrolase , Fosfatidilinositol 3-Quinases , RNA Circular , Proteínas de Ligação a RNA , Transdução de Sinais , Neoplasias da Bexiga Urinária , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/metabolismo , Humanos , RNA Circular/genética , RNA Circular/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Transdução de Sinais/genética , PTEN Fosfo-Hidrolase/metabolismo , PTEN Fosfo-Hidrolase/genética , Linhagem Celular Tumoral , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositol 3-Quinases/genética , Movimento Celular/genética , Masculino , Animais , Regulação Neoplásica da Expressão Gênica , Metástase Neoplásica , Feminino , Neovascularização Patológica/genética , Camundongos Nus , Camundongos , Pessoa de Meia-Idade , Camundongos Endogâmicos BALB CRESUMO
Peritumoral hepatocytes are critical components of the liver cancer microenvironment, However, the role of peritumoral hepatocytes in the local tumor immune interface and the underlying molecular mechanisms have not been elucidated. YTHDF2, an RNA N6-methyladenosine (m6A) reader, is critical for liver tumor progression. The function and regulatory roles of YTHDF2 in peritumoral hepatocytes are unknown. This study demonstrated that oxaliplatin (OXA) upregulated m6A modification and YTHDF2 expression in hepatocytes. Studies using tumor-bearing liver-specific Ythdf2 knockout mice revealed that hepatocyte YTHDF2 suppresses liver tumor growth through CD8+ T cell recruitment and activation. Additionally, YTHDF2 mediated the response to immunotherapy. Mechanistically, OXA upregulated YTHDF2 expression by activating the cGAS-STING signaling pathway and consequently enhanced the therapeutic outcomes of immunotherapeutic interventions. Ythdf2 stabilized Cx3cl1 transcripts in an m6A-dependent manner, regulating the interplay between CD8+ T cells and the progression of liver malignancies. Thus, this study elucidated the novel role of hepatocyte YTHDF2, which promotes therapy-induced antitumor immune responses in the liver. The findings of this study provide valuable insights into the mechanism underlying the therapeutic benefits of targeting YTHDF2.
Assuntos
Linfócitos T CD8-Positivos , Quimiocina CX3CL1 , Hepatócitos , Neoplasias Hepáticas , Oxaliplatina , Proteínas de Ligação a RNA , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Animais , Quimiocina CX3CL1/metabolismo , Quimiocina CX3CL1/genética , Hepatócitos/metabolismo , Camundongos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Humanos , Oxaliplatina/farmacologia , Microambiente Tumoral/imunologia , Camundongos Knockout , Regulação Neoplásica da Expressão Gênica , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular Tumoral , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Adenosina/análogos & derivados , Adenosina/metabolismo , Antineoplásicos/farmacologiaRESUMO
Messenger RNA splicing and degradation are critical for gene expression regulation, the abnormality of which leads to diseases. Previous methods for estimating kinetic rates have limitations, assuming uniform rates across cells. DeepKINET is a deep generative model that estimates splicing and degradation rates at single-cell resolution from scRNA-seq data. DeepKINET outperforms existing methods on simulated and metabolic labeling datasets. Applied to forebrain and breast cancer data, it identifies RNA-binding proteins responsible for kinetic rate diversity. DeepKINET also analyzes the effects of splicing factor mutations on target genes in erythroid lineage cells. DeepKINET effectively reveals cellular heterogeneity in post-transcriptional regulation.
Assuntos
Splicing de RNA , Análise de Célula Única , Humanos , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Estabilidade de RNA , Prosencéfalo/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Animais , FemininoRESUMO
Rationale: Epilepsy affects over 70 million people globally, with temporal lobe epilepsy with hippocampal sclerosis (TLE-HS) often progressing to a drug-resistant state. Recent research has highlighted the role of reactive astrocytes and glutamate dysregulation in epilepsy pathophysiology. This study aims to investigate the involvement of astrocytic xCT, a glutamate-cystine antiporter, and its regulation by the m6A reader protein YTHDC2 in TLE-HS. Methods: A pilocarpine-induced epilepsy model in mice was used to study the role of xCT in reactive astrocytes. The expression of xCT and its regulation by YTHDC2 were assessed through various molecular and cellular techniques. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to measure mRNA and protein levels of xCT and YTHDC2, respectively; immunofluorescence was utilized to visualize their localization and expression in astrocytes. In vivo glutamate measurements were conducted using microdialysis to monitor extracellular glutamate levels in the hippocampus. RNA immunoprecipitation-qPCR (RIP-qPCR) was performed to investigate the binding of YTHDC2 to SLC7A11 mRNA, while methylated RNA immunoprecipitation-qPCR (MeRIP-qPCR) was performed to quantify m6A modifications on SLC7A11 mRNA. A dual-luciferase reporter assay was conducted to assess the effect of m6A modifications on SLC7A11 mRNA translation, and polysome profiling was employed to evaluate the translational efficiency of SLC7A11 mRNA. Inhibition experiments involved shRNA-mediated knockdown of SLC7A11 (commonly known as xCT) and YTHDC2 expression in astrocytes. Video-electroencephalogram (EEG) recordings were used to monitor seizure activity in mice. Results: The xCT transporter in reactive astrocytes significantly contributes to elevated extracellular glutamate levels, enhancing neuronal excitability and seizure activity. Increased xCT expression is influenced by the m6A reader protein YTHDC2, which regulates its expression through m6A methylation. Inhibition of xCT or YTHDC2 in astrocytes reduces glutamate levels and effectively controls seizures in a mouse model. Specifically, mice with SLC7A11- or YTHDC2-knockdown astrocytes showed decreased glutamate concentration in the hippocampus and reduced frequency and duration of epileptic seizures. Conclusions: This study highlights the therapeutic potential of targeting YTHDC2 and xCT in reactive astrocytes to mitigate epilepsy. The findings provide a novel perspective on the mechanisms of glutamate dysregulation and their implications in seizure pathophysiology, suggesting that modulation of YTHDC2 and xCT could be a promising strategy for treating TLE.
Assuntos
Sistema y+ de Transporte de Aminoácidos , Astrócitos , Modelos Animais de Doenças , Epilepsia do Lobo Temporal , Ácido Glutâmico , Animais , Astrócitos/metabolismo , Epilepsia do Lobo Temporal/metabolismo , Epilepsia do Lobo Temporal/genética , Epilepsia do Lobo Temporal/fisiopatologia , Camundongos , Sistema y+ de Transporte de Aminoácidos/metabolismo , Sistema y+ de Transporte de Aminoácidos/genética , Ácido Glutâmico/metabolismo , Masculino , Hipocampo/metabolismo , Pilocarpina , Camundongos Endogâmicos C57BL , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , HumanosRESUMO
Viral infections pose significant threats to human health, leading to a diverse spectrum of infectious diseases. The innate immune system serves as the primary barrier against viruses and bacteria in the early stages of infection. A rapid and forceful antiviral innate immune response is triggered by distinguishing between self-nucleic acids and viral nucleic acids. RNA-binding proteins (RBPs) are a diverse group of proteins which contain specific structural motifs or domains for binding RNA molecules. In the last decade, numerous of studies have outlined that RBPs influence viral replication via diverse mechanisms, directly recognizing viral nucleic acids and modulating the activity of pattern recognition receptors (PRRs). In this review, we summarize the functions of RBPs in regulation of host-virus interplay by controlling the activation of PRRs, such as RIG-I, MDA5, cGAS and TLR3. RBPs are instrumental in facilitating the identification of viral RNA or DNA, as well as viral structural proteins within the cellular cytoplasm and nucleus, functioning as co-receptor elements. On the other hand, RBPs are capable of orchestrating the activation of PRRs and facilitating the transmission of antiviral signals to downstream adaptor proteins by post-translational modifications or aggregation. Gaining a deeper comprehension of the interaction between the host and viruses is crucial for the development of novel therapeutics targeting viral infections.
Assuntos
Imunidade Inata , Proteínas de Ligação a RNA , Receptores de Reconhecimento de Padrão , Transdução de Sinais , Receptores de Reconhecimento de Padrão/metabolismo , Receptores de Reconhecimento de Padrão/imunologia , Humanos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/imunologia , Proteínas de Ligação a RNA/genética , Animais , Viroses/imunologia , Viroses/virologia , Interações Hospedeiro-Patógeno/imunologia , RNA Viral/metabolismo , RNA Viral/imunologia , RNA Viral/genética , Vírus/imunologia , Replicação ViralRESUMO
In addressing the challenges of lung cancer, attention has increasingly turned to molecular diagnostics and targeted therapies, with nucleolin (NCL) assuming a pivotal role, especially in non-small cell lung cancer. The aberrant activity and cellular distribution of NCL act as crucial biomarkers for early detection and treatment monitoring, showing a strong correlation with disease progression and patient prognosis. Elevated NCL levels signal advanced disease and poorer outcomes, underscoring its significance in evaluating disease severity and therapeutic response. Strategies targeting the molecular interactions of NCL have spurred innovative approaches to inhibit tumour growth, overcome drug resistance and improve treatment efficacy. These advancements are paving the way for personalized therapies tailored to the unique molecular profiles of patients' tumours. Consequently, NCL stands at the forefront of lung cancer management, symbolizing the move towards more precise and individualized oncology care, and marking substantial progress in therapeutic development.
Assuntos
Biomarcadores Tumorais , Neoplasias Pulmonares , Nucleolina , Fosfoproteínas , Proteínas de Ligação a RNA , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/terapia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/tratamento farmacológico , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Fosfoproteínas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/terapia , Carcinoma Pulmonar de Células não Pequenas/patologia , Prognóstico , Animais , Terapia de Alvo MolecularRESUMO
N6-methyladenosine (m6A) RNA methylation is a prevalent RNA modification that significantly impacts RNA metabolism and cancer development. Maintaining the global m6A levels in cancer cells relies on RNA accessibility to methyltransferases and the availability of the methyl donor S-adenosylmethionine (SAM). Here, we reveal that death associated protein 3 (DAP3) plays a crucial role in preserving m6A levels through two distinct mechanisms. First, although DAP3 is not a component of the m6A writer complex, it directly binds to m6A target regions, thereby facilitating METTL3 binding. Second, DAP3 promotes MAT2A's last intron splicing, increasing MAT2A protein, cellular SAM, and m6A levels. Silencing DAP3 hinders tumorigenesis, which can be rescued by MAT2A overexpression. This evidence suggests DAP3's role in tumorigenesis, partly through m6A regulation. Our findings unveil DAP3's complex role as an RNA-binding protein and tumor promoter, impacting RNA processing, splicing, and m6A modification in cancer transcriptomes.
Assuntos
Adenosina , Metionina Adenosiltransferase , Metiltransferases , Neoplasias , Humanos , Adenosina/análogos & derivados , Adenosina/metabolismo , Metiltransferases/metabolismo , Metiltransferases/genética , Metionina Adenosiltransferase/metabolismo , Metionina Adenosiltransferase/genética , Neoplasias/genética , Neoplasias/metabolismo , Metilação , Linhagem Celular Tumoral , S-Adenosilmetionina/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas Reguladoras de Apoptose/genética , Regulação Neoplásica da Expressão Gênica , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Splicing de RNA/genética , Animais , Camundongos , RNA/metabolismo , RNA/genética , Processamento Pós-Transcricional do RNA , Metilação de RNARESUMO
Mitochondrial dysfunction is a pivotal event contributing to the development of ageing-related kidney disorders. Lon protease 1 (LONP1) has been reported to be responsible for ageing-related renal fibrosis; however, the underlying mechanism(s) of LONP1-driven kidney ageing with respect to mitochondrial disturbances remains to be further explored. The level of LONP1 was tested in the kidneys of aged humans and mice. Renal fibrosis and mitochondrial quality control were confirmed in the kidneys of aged mice. Effects of LONP1 silencing or overexpression on renal fibrosis and mitochondrial quality control were explored. In addition, N6-methyladenosine (m6A) modification and methyltransferase like 3 (METTL3) levels, the relationship between LONP1 and METTL3, and the impacts of METTL3 overexpression on mitochondrial functions were confirmed. Furthermore, the expression of insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) and the regulatory effects of IGF2BP2 on LONP1 were confirmed in vitro. LONP1 expression was reduced in the kidneys of aged humans and mice, accompanied by renal fibrosis and mitochondrial dysregulation. Overexpression of LONP1 alleviated renal fibrosis and maintained mitochondrial homeostasis, while silencing of LONP1 had the opposite effect. Impaired METTL3-m6A signalling contributed at least in part to ageing-induced LONP1 modification, reducing subsequent degradation in an IGF2BP2-dependent manner. Moreover, METTL3 overexpression alleviated proximal tubule cell injury, preserved mitochondrial stability, inhibited LONP1 degradation, and protected mitochondrial functions. LONP1 mediates mitochondrial function in kidney ageing and that targeting LONP1 may be a potential therapeutic strategy for improving ageing-related renal fibrosis.
Assuntos
Adenosina , Envelhecimento , Fibrose , Homeostase , Nefropatias , Rim , Metiltransferases , Mitocôndrias , Proteínas Mitocondriais , Proteínas de Ligação a RNA , Mitocôndrias/metabolismo , Animais , Metiltransferases/metabolismo , Metiltransferases/genética , Humanos , Envelhecimento/metabolismo , Camundongos , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Rim/patologia , Rim/metabolismo , Masculino , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Adenosina/análogos & derivados , Adenosina/metabolismo , Nefropatias/metabolismo , Nefropatias/patologia , Nefropatias/etiologia , Nefropatias/genética , Proteases Dependentes de ATP/metabolismo , Proteases Dependentes de ATP/genética , Transdução de Sinais , Camundongos Endogâmicos C57BLRESUMO
Biological systems encompass intricate networks governed by RNA-protein interactions that play pivotal roles in cellular functions. RNA and proteins constituting 1.1% and 18% of the mammalian cell weight, respectively, orchestrate vital processes from genome organization to translation. To date, disentangling the functional fraction of the human genome has presented a major challenge, particularly for noncoding regions, yet recent discoveries have started to unveil a host of regulatory functions for noncoding RNAs (ncRNAs). While ncRNAs exist at different sizes, structures, degrees of evolutionary conservation and abundances within the cell, they partake in diverse roles either alone or in combination. However, certain ncRNA subtypes, including those that have been described or remain to be discovered, are poorly characterized given their heterogeneous nature. RNA activity is in most cases coordinated through interactions with RNA-binding proteins (RBPs). Extensive efforts are being made to accurately reconstruct RNA-RBP regulatory networks, which have provided unprecedented insight into cellular physiology and human disease. In this review, we provide a comprehensive view of RNAs and RBPs, focusing on how their interactions generate functional signals in living cells, particularly in the context of post-transcriptional regulatory processes and cancer.
Assuntos
Homeostase , Neoplasias , RNA não Traduzido , Proteínas de Ligação a RNA , Humanos , Neoplasias/genética , Neoplasias/patologia , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Homeostase/genética , RNA não Traduzido/genética , Redes Reguladoras de Genes , AnimaisRESUMO
BACKGROUND: N6-methyladenosine (m6A) modification of messenger RNA (mRNA) is crucial for liquid-liquid phase separation in mammals. Increasing evidence indicates that liquid-liquid phase separation in proteins and RNAs affects diabetic cardiomyopathy. However, the molecular mechanism by which m6A-mediated phase separation regulates diabetic cardiac fibrosis remains elusive. METHODS: Leptin receptor-deficient mice (db/db), cardiac fibroblast-specific Notch1 conditional knockout (POSTN-Cre × Notch1flox/flox) mice, and Cre mice were used to induce diabetic cardiac fibrosis. Adeno-associated virus 9 carrying cardiac fibroblast-specific periostin (Postn) promoter-driven small hairpin RNA targeting Alkbh5, Ythdf2, or Notch1, and the phase separation inhibitor 1,6-hexanediol were administered to investigate their roles in diabetic cardiac fibrosis. Histological and biochemical analyses were performed to determine how Alkbh5 and Ythdf2 regulate Notch1 expression in diabetic cardiac fibrosis. NOTCH1 was reconstituted in ALKBH5- and YTHDF2-deficient cardiac fibroblasts and mouse hearts to study its effects on mitochondrial fission and diabetic cardiac fibrosis. Heart tissue samples from patients with diabetic cardiomyopathy were used to validate our findings. RESULTS: In mice with diabetic cardiac fibrosis, decreased Notch1 expression was accompanied by high m6A mRNA levels and mitochondrial fission. Fibroblast-specific deletion of Notch1 enhanced mitochondrial fission and cardiac fibroblast proliferation and induced diabetic cardiac fibrosis in mice. Notch1 downregulation was associated with Alkbh5-mediated m6A demethylation in the 3'UTR of Notch1 mRNA and elevated m6A mRNA levels. These elevated m6A levels in Notch1 mRNA markedly enhanced YTHDF2 phase separation, increased the recognition of m6A residues in Notch1 mRNA by YTHDF2, and induced Notch1 degradation. Conversely, epitranscriptomic downregulation rescues Notch1 expression, resulting in the opposite effects. Human heart tissues from patients with diabetic cardiomyopathy were used to validate the findings in mice with diabetic cardiac fibrosis. CONCLUSIONS: We identified a novel epitranscriptomic mechanism by which m6A-mediated phase separation suppresses Notch1 expression, thereby promoting mitochondrial fission in diabetic cardiac fibrosis. Our findings provide new insights for the development of novel treatment approaches for patients with diabetic cardiac fibrosis.