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Sci Rep ; 7(1): 3508, 2017 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-28615644


microRNAs (miRNAs) are tightly regulated during T lymphocyte activation to enable the establishment of precise immune responses. Here, we analyzed the changes of the miRNA profiles of T cells in response to activation by cognate interaction with dendritic cells. We also studied mRNA targets common to miRNAs regulated in T cell activation. pik3r1 gene, which encodes the regulatory subunits of PI3K p50, p55 and p85, was identified as target of miRNAs upregulated after T cell activation. Using 3'UTR luciferase reporter-based and biochemical assays, we showed the inhibitory relationship between miR-132-3p upregulation and expression of the pik3r1 gene. Our results indicate that specific miRNAs whose expression is modulated during T cell activation might regulate PI3K signaling in T cells.

Linfócitos T CD4-Positivos/metabolismo , Classe Ia de Fosfatidilinositol 3-Quinase/metabolismo , MicroRNAs/metabolismo , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Animais , Células Cultivadas , Células HEK293 , Humanos , Camundongos Endogâmicos C57BL , Regulação para Cima
Mol Cell ; 63(3): 397-407, 2016 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-27477908


Long noncoding RNAs (lncRNAs) are involved in diverse cellular processes through multiple mechanisms. Here, we describe a previously uncharacterized human lncRNA, CONCR (cohesion regulator noncoding RNA), that is transcriptionally activated by MYC and is upregulated in multiple cancer types. The expression of CONCR is cell cycle regulated, and it is required for cell-cycle progression and DNA replication. Moreover, cells depleted of CONCR show severe defects in sister chromatid cohesion, suggesting an essential role for CONCR in cohesion establishment during cell division. CONCR interacts with and regulates the activity of DDX11, a DNA-dependent ATPase and helicase involved in DNA replication and sister chromatid cohesion. These findings unveil a direct role for an lncRNA in the establishment of sister chromatid cohesion by modulating DDX11 enzymatic activity.

Cromátides/metabolismo , Replicação do DNA , DNA de Neoplasias/biossíntese , Neoplasias/metabolismo , RNA Longo não Codificante/metabolismo , Células A549 , Animais , Apoptose , Proliferação de Células , Cromátides/genética , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , DNA Helicases/genética , DNA Helicases/metabolismo , DNA de Neoplasias/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Células HeLa , Humanos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Neoplasias/genética , Neoplasias/patologia , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Interferência de RNA , RNA Longo não Codificante/genética , Fatores de Tempo , Transcrição Genética , Ativação Transcricional , Transfecção , Carga Tumoral , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
Sci Rep ; 6: 20223, 2016 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-26838552


Evidence links aryl hydrocarbon receptor (AHR) activation to rheumatoid arthritis (RA) pathogenesis, although results are inconsistent. AHR agonists inhibit pro-inflammatory cytokine expression in macrophages, pivotal cells in RA aetiopathogenesis, which hints at specific circuits that regulate the AHR pathway in RA macrophages. We compared microRNA (miR) expression in CD14(+) cells from patients with active RA or with osteoarthritis (OA). Seven miR were downregulated and one (miR-223) upregulated in RA compared to OA cells. miR-223 upregulation correlated with reduced Notch3 and Notch effector expression in RA patients. Overexpression of the Notch-induced repressor HEY-1 and co-culture of healthy donor monocytes with Notch ligand-expressing cells showed direct Notch-mediated downregulation of miR-223. Bioinformatics predicted the AHR regulator ARNT (AHR nuclear translocator) as a miR-223 target. Pre-miR-223 overexpression silenced ARNT 3'UTR-driven reporter expression, reduced ARNT (but not AHR) protein levels and prevented AHR/ARNT-mediated inhibition of pro-inflammatory cytokine expression. miR-223 counteracted AHR/ARNT-induced Notch3 upregulation in monocytes. Levels of ARNT and of CYP1B1, an AHR/ARNT signalling effector, were reduced in RA compared to OA synovial tissue, which correlated with miR-223 levels. Our results associate Notch signalling to miR-223 downregulation in RA macrophages, and identify miR-223 as a negative regulator of the AHR/ARNT pathway through ARNT targeting.

Artrite Reumatoide/genética , Translocador Nuclear Receptor Aril Hidrocarboneto/metabolismo , Citocinas/metabolismo , Macrófagos/metabolismo , MicroRNAs/genética , Receptores Notch/genética , Idoso , Artrite Reumatoide/patologia , Translocador Nuclear Receptor Aril Hidrocarboneto/genética , Técnicas de Cocultura , Citocinas/genética , Feminino , Perfilação da Expressão Gênica/métodos , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Osteoartrite/genética , Osteoartrite/patologia , Transdução de Sinais
Nat Commun ; 5: 4617, 2014 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-25099865


The enormous amount of environmental arsenic was a major factor in determining the biochemistry of incipient life forms early in the Earth's history. The most abundant chemical form in the reducing atmosphere was arsenite, which forced organisms to evolve strategies to manage this chemical species. Following the great oxygenation event, arsenite oxidized to arsenate and the action of arsenate reductases became a central survival requirement. The identity of a biologically relevant arsenate reductase in plants nonetheless continues to be debated. Here we identify a quantitative trait locus that encodes a novel arsenate reductase critical for arsenic tolerance in plants. Functional analyses indicate that several non-additive polymorphisms affect protein structure and account for the natural variation in arsenate reductase activity in Arabidopsis thaliana accessions. This study shows that arsenate reductases are an essential component for natural plant variation in As(V) tolerance.

Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Arseniato Redutases/metabolismo , Arsênico/química , Regulação da Expressão Gênica de Plantas , Alelos , Sequência de Aminoácidos , Arabidopsis/efeitos dos fármacos , Arsenitos/química , Mapeamento Cromossômico , Escherichia coli/metabolismo , Teste de Complementação Genética , Modelos Moleculares , Conformação Molecular , Dados de Sequência Molecular , Mutação , Oxigênio/química , Fenótipo , Polimorfismo Genético , Locos de Características Quantitativas , Homologia de Sequência de Aminoácidos , Tiossulfato Sulfurtransferase/química
BMC Genomics ; 15 Suppl 10: S2, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25559987


BACKGROUND: MicroRNAs are short RNA molecules that post-transcriptionally regulate gene expression. Today, microRNA target prediction remains challenging since very few have been experimentally validated and sequence-based predictions have large numbers of false positives. Furthermore, due to the different measuring rules used in each database of predicted interactions, the selection of the most reliable ones requires extensive knowledge about each algorithm. RESULTS: Here we propose two methods to measure the confidence of predicted interactions based on experimentally validated information. The output of the methods is a combined database where new scores and statistical confidences are re-assigned to each predicted interaction. The new scores allow the robust combination of several databases without the effect of low-performing algorithms dragging down good-performing ones. The combined databases obtained using both algorithms described in this paper outperform each of the existing predictive algorithms that were considered for the combination. CONCLUSIONS: Our approaches are a useful way to integrate predicted interactions from different databases. They reduce the selection of interactions to a unique database based on an intuitive score and allow comparing databases between them.

Biologia Computacional/métodos , MicroRNAs/metabolismo , RNA Mensageiro/metabolismo , Algoritmos , Animais , Bases de Dados de Ácidos Nucleicos , Humanos , MicroRNAs/genética , Modelos Estatísticos , RNA Mensageiro/genética , Reprodutibilidade dos Testes
Nat Commun ; 4: 2980, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24356509


Exosomes are released by most cells to the extracellular environment and are involved in cell-to-cell communication. Exosomes contain specific repertoires of mRNAs, microRNAs (miRNAs) and other non-coding RNAs that can be functionally transferred to recipient cells. However, the mechanisms that control the specific loading of RNA species into exosomes remain unknown. Here we describe sequence motifs present in miRNAs that control their localization into exosomes. The protein heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1) specifically binds exosomal miRNAs through the recognition of these motifs and controls their loading into exosomes. Moreover, hnRNPA2B1 in exosomes is sumoylated, and sumoylation controls the binding of hnRNPA2B1 to miRNAs. The loading of miRNAs into exosomes can be modulated by mutagenesis of the identified motifs or changes in hnRNPA2B1 expression levels. These findings identify hnRNPA2B1 as a key player in miRNA sorting into exosomes and provide potential tools for the packaging of selected regulatory RNAs into exosomes and their use in biomedical applications.

Exossomos/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/metabolismo , MicroRNAs/metabolismo , Motivos de Aminoácidos , Comunicação Celular , Inativação Gênica , Humanos , Células Jurkat , Leucócitos Mononucleares/metabolismo , Mutagênese , Mutagênese Sítio-Dirigida , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Ligação Proteica , Sumoilação