RESUMEN
ABSTRACT: Regulation of RNA polymerase II (RNAPII) activity is an essential process that governs gene expression; however, its contribution to the fundamental process of erythropoiesis remains unclear. hexamethylene bis-acetamide inducible 1 (HEXIM1) regulates RNAPII activity by controlling the location and activity of positive transcription factor ß. We identified a key role for HEXIM1 in controlling erythroid gene expression and function, with overexpression of HEXIM1 promoting erythroid proliferation and fetal globin expression. HEXIM1 regulated erythroid proliferation by enforcing RNAPII pausing at cell cycle check point genes and increasing RNAPII occupancy at genes that promote cycle progression. Genome-wide profiling of HEXIM1 revealed that it was increased at both repressed and activated genes. Surprisingly, there were also genome-wide changes in the distribution of GATA-binding factor 1 (GATA1) and RNAPII. The most dramatic changes occurred at the ß-globin loci, where there was loss of RNAPII and GATA1 at ß-globin and gain of these factors at γ-globin. This resulted in increased expression of fetal globin, and BGLT3, a long noncoding RNA in the ß-globin locus that regulates fetal globin expression. GATA1 was a key determinant of the ability of HEXIM1 to repress or activate gene expression. Genes that gained both HEXIM1 and GATA1 had increased RNAPII and increased gene expression, whereas genes that gained HEXIM1 but lost GATA1 had an increase in RNAPII pausing and decreased expression. Together, our findings reveal a central role for universal transcription machinery in regulating key aspects of erythropoiesis, including cell cycle progression and fetal gene expression, which could be exploited for therapeutic benefit.
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Eritropoyesis , Factores de Transcripción , Humanos , Eritropoyesis/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Regulación de la Expresión Génica , Transcripción Genética , Globinas beta/genética , Globinas beta/metabolismo , ARN Polimerasa II/genética , ARN Polimerasa II/metabolismo , Factor de Transcripción GATA1/genética , Factor de Transcripción GATA1/metabolismo , Proteínas de Unión al ARN/genéticaRESUMEN
This work studies the effect of Nb, Mo, Re dopant, and Se vacancy in WSe2 on the electronic and optical properties of the MoS2/WSe2 bilayer heterostructure based on first-principles calculations. Our research shows that the MoS2/WSe2 bilayer heterostructure exhibits a type-II band alignment with a valence band offset (VBO) of 1.07 eV and a conduction band offset (CBO) of 1.00 eV. It also shows that different dopants or defects can considerably modulate the energy band alignment and interlayer charge transfer of the heterostructure. Owing to the orbital hybridization of the dopant atoms with other atoms and the consequent enhancement of the coupling between the two structural layers, a transition of the band alignment from type-II to type-I is realized with the Re dopant. The effect of doping and defects on the electronic properties of heterojunctions contributes to applications in high-performance optoelectronic devices.
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Hirschsprung's disease (HSCR) is characterized by the lack of ganglion cells in the distal part of the digestive tract. It occurs due to migration disorders of enteric neural crest cells (ENCCs) from 5 to 12 weeks of embryonic development. More and more studies show that HSCR is a result of the interaction of multiple genes and the microenvironments, but its specific pathogenesis has not been fully elucidated. Studies have confirmed that many substances in the intestinal microenvironment, such as laminin and ß1-integrin, play a vital regulatory role in cell growth and disease progression. In addition to these high-molecular-weight proteins, research on endogenous polypeptides derived from these proteins has been increasing in recent years. However, it is unclear whether these endogenous peptides have effects on the migration of ENCCs and thus participate in the occurrence of HSCR. Previously, our research group found that compared with the normal intestinal tissue, the expression of AHNAK protein in the stenosed intestinal tissue of HSCR patients was significantly upregulated, and overexpression of AHNAK could inhibit cell migration and proliferation. In this study, endogenous peptides were extracted from the normal control intestinal tissue and the stenosed HSCR intestinal tissue. The endogenous polypeptide expression profile was analyzed by liquid chromatography-mass spectrometry, and multiple peptides derived from AHNAK protein were found. We selected one of them, "EGPEVDVNLPK", for research. Because there is no uniform naming system, this peptide is temporarily named PDAHNAK (peptide derived from AHNAK). This project aims to clarify the potential role of PDAHNAK in the development of HSCR and to further understand its relationship with its precursor protein AHNAK and how they contribute to the development of HSCR.
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Enfermedad de Hirschsprung , Movimiento Celular , Proliferación Celular , Enfermedad de Hirschsprung/genética , Humanos , Sistema de Señalización de MAP Quinasas , Proteínas de la Membrana , Proteínas de Neoplasias , PéptidosRESUMEN
Circular RNAs (circRNAs) play a vital role in cancers. Accumulated evidences showed that the physiological condition of cells can be reflected by the circRNAs in the exosomes they secrete, and these exosomal circRNAs can be captured by the receptor cells, thereby inducing a series of cellular responses. We performed qRT-PCR to detect the expression level of circ-0000284 in cholangiocarcinoma cell lines, tissues and plasma exosomes. Then the direct interaction between circ-0000284 and miR-637 was investigated through dual-luciferase reporter assay, RNA binding protein immunoprecipitation (RIP) assay and Fluorescent in situ hybridization (FISH) assay. Subsequently, EdU (5-ethynyl-2'-deoxyuridine), migration, invasion assay, flow cytometry and nude mouse tumorigenicity assay were adopted to evaluate the effect of circ-0000284 on migration, invasion, proliferation and apoptosis of cholangiocarcinoma cells. Additionally, TEM was conducted to investigate the shape and size of exosomes from cholangiocarcioma and 293T cell lines. Circ-0000284 was evidently elevated in cholangiocarcinoma cell lines, tumor tissues and plasma exosomes. Meanwhile, the high expression of circ-0000284 enhanced the migration, invasion and proliferation abilities of cholangiocarcinoma cells in vivo and in vitro Besides, the levels of circ-0000284 were increased in cholangiocarcinoma cells and exosomes from them. Moreover, exosomes from cholangiocarcinoma cells enhanced circ-0000284 expression and stimulated migration and proliferation of the surrounding normal cells. Our findings suggest that on the one hand circ-0000284 functions as a competitive endogenous RNA to promote cholangiocarcinoma progression, and on the other hand, circ-0000284 can be directly transferred from cholangiocarcinoma cells to surrounding normal cells via exosomes and in this way regulate the biological functions of surrounding normal cells.
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Comunicación Celular , Colangiocarcinoma/genética , Colangiocarcinoma/patología , ARN Circular/metabolismo , Apoptosis/genética , Secuencia de Bases , Comunicación Celular/genética , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Colangiocarcinoma/ultraestructura , Exosomas/ultraestructura , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , MicroARNs/genética , MicroARNs/metabolismo , Persona de Mediana Edad , Invasividad Neoplásica , Metástasis de la Neoplasia , Fenotipo , ARN Circular/genéticaRESUMEN
OBJECTIVES: This study aims to investigate the role of demethylase ALKBH5 mediated demethylation of TAGLN mRNA in the occurrence of Hirschsprung's disease (HSCR), and to clarify how ALKBH5 reduces the m6A level of TAGLN mRNA and inhibits its degradation, thereby inhibiting the proliferation and migration of neural crest cells, and potentially contributing to the occurrence of HSCR. MATERIAL AND METHODS: Quantitative real-time PCR (qRT-PCR) and Western-Blot (WB) were conducted to test the expression level of ALKBH5 and TAGLN genes. Cell function assays were adopted to detect cell phenotypes. The qRT-PCR and methylated RNA immunoprecipitation (MeRIP-qPCR) were used to test the regulation of TAGLN by ALKBH5. RESULTS: 1. Compared with control intestinal tissue, the expression level of TAGLN and ALKBH5 in the aganglionic intestinal tissue of HSCR is increased. 2. The MeRIP-PCR and dualluciferase report confirmed that ALKBH5 could bind to m6A sites of TAGLN mRNA and reduce the m6A level of TAGLN mRNA. 3. In vitro cell experiments confirmed that overexpression of ALKBH5 can inhibit the degradation of TAGLN mRNA, increase the expression of TAGLN, thereby inhibiting cell proliferation and migration. 4. A zebrafish model of ALKBH5 overexpression was constructed. Studies have shown that ALKBH5 could inhibit the proliferation and migration of zebrafish enteric neurons. CONCLUSIONS: ALKBH5 could demethylate TAGLN mRNA and up-regulate TAGLN expression, leading to the inhibition of proliferation and migration of enteric neural crest cells and contributing to the occurrence of HSCR.
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Desmetilasa de ARN, Homólogo 5 de AlkB/metabolismo , Enfermedad de Hirschsprung/metabolismo , Intestinos/patología , Proteínas de Microfilamentos/metabolismo , Proteínas Musculares/metabolismo , Cresta Neural/metabolismo , Pez Cebra/genética , Animales , Sitios de Unión , Estudios de Casos y Controles , Movimiento Celular , Proliferación Celular , Femenino , Perfilación de la Expresión Génica , Células HEK293 , Enfermedad de Hirschsprung/genética , Humanos , Lactante , Masculino , Metiltransferasas/metabolismoRESUMEN
Aim: To discover the potential roles of plasma exosomal miRNAs in Hirschsprung's disease (HSCR) and identify potential noninvasive biomarkers for early diagnosis of HSCR. Materials & methods: Plasma samples were collected from HSCR patients and matched controls. Exosomes were isolated before high-throughput Illumina sequencing was utilized to gain a profile of dysregulated exosomal miRNAs, followed with further verification in two separate cohorts. Bioinformatics analyses were also adopted to explore the molecular functions of dysregulated miRNAs in Hirschsprung's disease. Results & conclusion: 31 dysregulated miRNAs were identified with five considered as promising HSCR signatures. Gene enrichment analysis disclosed that the upregulated miRNAs were most likely to participate in 'extracellular matrix-receptor interaction' and contribute to HSCR through interfering in cell junctions.
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Biomarcadores , MicroARN Circulante , Exosomas , Enfermedad de Hirschsprung/diagnóstico , Enfermedad de Hirschsprung/genética , MicroARNs/genética , Biología Computacional/métodos , Exosomas/genética , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Enfermedad de Hirschsprung/sangre , Humanos , Biopsia Líquida/métodos , MicroARNs/sangre , Pronóstico , Interferencia de ARNRESUMEN
Intestinal malrotation in newborns often requires urgent surgical treatment, especially in the presence of volvulus. Therefore, earlystage diagnosis is critical. In the present study, differentially expressed plasma microRNAs (miRNAs) were screened for in patients with intestinal malrotation using highthroughput Illumina sequencing, and validated using reverse transcriptionquantitative PCR. Receiver operating characteristic curve (ROC) analysis was conducted to evaluate their specificity, sensitivity and assess their diagnostic value for intestinal malrotation. Bioinformatics analysis was performed to investigate the functions associated with the dysregulated miRNAs. A profile consisting of 28 differentially expressed plasma miRNAs was obtained, of which nine were verified to exhibit significantly altered expression. According to a ROC analysis, four of these could represent novel earlystage, noninvasive biomarkers for intestinal malrotation. Bioinformatics analysis demonstrated that the differentially expressed miRNAs were predominantly involved in 'metal ion transmembrane transporter activity' and 'calciumdependent protein binding', which may be related to the 'endocytosis' pathway. In conclusion, significantly differentially expressed plasma miRNAs were identified in congenital intestinal malrotation and their potential roles were described. These differentially expressed miRNAs may serve as biomarkers of intestinal malrotation and improve early diagnosis for this condition.
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Biomarcadores/sangre , Vólvulo Intestinal/congénito , Vólvulo Intestinal/diagnóstico , MicroARNs/sangre , Biología Computacional , Diagnóstico Precoz , Femenino , Perfilación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Recién Nacido , Vólvulo Intestinal/sangre , Vólvulo Intestinal/genética , Masculino , Análisis de Secuencia de ARNRESUMEN
Hirschsprung disease (HSCR) is a congenital disorder attributed to the failure of the neural crest derivatives migrating and/or differentiating along the hindgut. The most frequent complication in Hirschsprung disease patients is Hirschsprung-associated enterocolitis (HAEC). However, its pathogenesis has not been fully understood. This study investigated miRNAs influenced by Lipopolysaccharide (LPS) in postoperative HAEC patients, their effect on enterocolitis and the underlying mechanism. MiR-132 and miR-212 were up-regulated in HAEC dilated tissues and LPS-treated mice enteritis samples. LPS-stimulated HT29 cells showed a high expression of miR-132 and miR-212. QRT-PCR analysis, western blotting, luciferase reporter assay, and flow cytometric analysis were carried out in vitro, showing that miR-132 and miR-212 could directly inhibit Sirtuin 1 (SIRT1) expression. Consequently, SIRT1 deficiency in LPS-stimulated HT29 cell line and LPS-treated mice activated NLRP3 inflammasome and Caspase-1-mediated pyroptosis. Furthermore, the above inflammation activation was reversed by miR-132/212 inhibitor or SIRT1 overexpression plasmid transfection.In conclusion, LPS upregulated miR-132 and miR-212 expression in HAEC, suppressing SIRT1 and facilitating NLRP3 inflammasome activation, which induced pyroptosis. Our findings illustrated the role of LPS/miR-132/-212/SIRT1/NLRP3 regulatory network in the occurrence and progression of HAEC and proposed a new molecular pathway for LPS-mediated cell pyroptosis.
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Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide. Recent years, circular RNA (circRNA) have been shown to exert vital functions in the pathological progressions of many diseases. A growing number of evidences have identified the representative function of exosomal circRNAs in the physiological state of donor cells, which further induces cellular responses after captured by recipient cells. However, the contributions of circRNAs to HCC remain largely unknown. In vitro and in vivo regulatory roles of circRNA Cdr1as in proliferative and migratory abilities of HCC were evaluated by CCK8, EdU, Transwell and tumourigenicity assays, respectively. Results showed circRNA Cdr1as was highly expressed in HCC cell lines and tissues. Overexpression of circRNA Cdr1as greatly accelerated HCC cells to proliferate and migrate. Mechanistically, we found that Cdr1as could promote the expression of AFP, a well-known biomarker for HCC, by sponging miR-1270. Further studies showed exosomes extracted from HCC cells overexpressing circRNA Cdr1as accelerated the proliferative and migratory abilities of surrounding normal cells. In all, circRNA Cdr1as serves as a ceRNA to promote the progression of HCC. Meanwhile, it is directly transferred from HCC cells to surrounding normal cells via exosomes to further mediate the biological functions of surrounding cells.
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Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , ARN Circular/metabolismo , ARN Largo no Codificante/metabolismo , Animales , Línea Celular , Movimiento Celular , Proliferación Celular , Humanos , Hígado/citología , Ratones , Ratones Desnudos , MicroARNs/genética , MicroARNs/metabolismo , ARN Circular/genética , ARN Largo no Codificante/genética , Regulación hacia ArribaRESUMEN
Long noncoding RNAs (lncRNAs) are characterized as a group of endogenous RNAs that are more than 200 nucleotides in length and have no protein-encoding function. More and more evidence indicates that lncRNAs play vital roles in various human diseases, especially in tumorigenesis. Focally amplified lncRNA on chromosome 1 (FAL1), a novel lncRNA with enhancer-like activity, has been identified as an oncogene in multiple cancers and high expression level of FAL1 is usually associated with poor prognosis. Dysregulation of FAL1 has been shown to promote the proliferation and metastasis of cancer cells. In the present review, we summarized and illustrated the functions and underlying molecular mechanisms of FAL1 in the occurrence and development of different cancers and other diseases. FAL1 has the potential to appear as a feasible diagnostic and prognostic tool and new therapeutic target for cancer patients though further investigation is needed so as to accelerate clinical application.