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2.
Cell ; 147(1): 132-46, 2011 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-21924763

RESUMO

Alternative splicing (AS) is a key process underlying the expansion of proteomic diversity and the regulation of gene expression. Here, we identify an evolutionarily conserved embryonic stem cell (ESC)-specific AS event that changes the DNA-binding preference of the forkhead family transcription factor FOXP1. We show that the ESC-specific isoform of FOXP1 stimulates the expression of transcription factor genes required for pluripotency, including OCT4, NANOG, NR5A2, and GDF3, while concomitantly repressing genes required for ESC differentiation. This isoform also promotes the maintenance of ESC pluripotency and contributes to efficient reprogramming of somatic cells into induced pluripotent stem cells. These results reveal a pivotal role for an AS event in the regulation of pluripotency through the control of critical ESC-specific transcriptional programs.


Assuntos
Processamento Alternativo , Reprogramação Celular , Células-Tronco Embrionárias/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Pluripotentes/metabolismo , Proteínas Repressoras/metabolismo , Animais , DNA/metabolismo , Células-Tronco Embrionárias/citologia , Genes Homeobox , Humanos , Camundongos , Células-Tronco Pluripotentes/citologia , Isoformas de Proteínas/metabolismo
3.
Mol Cell ; 65(3): 539-553.e7, 2017 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-28157508

RESUMO

Networks of coordinated alternative splicing (AS) events play critical roles in development and disease. However, a comprehensive knowledge of the factors that regulate these networks is lacking. We describe a high-throughput system for systematically linking trans-acting factors to endogenous RNA regulatory events. Using this system, we identify hundreds of factors associated with diverse regulatory layers that positively or negatively control AS events linked to cell fate. Remarkably, more than one-third of the regulators are transcription factors. Further analyses of the zinc finger protein Zfp871 and BTB/POZ domain transcription factor Nacc1, which regulate neural and stem cell AS programs, respectively, reveal roles in controlling the expression of specific splicing regulators. Surprisingly, these proteins also appear to regulate target AS programs via binding RNA. Our results thus uncover a large "missing cache" of splicing regulators among annotated transcription factors, some of which dually regulate AS through direct and indirect mechanisms.


Assuntos
Processamento Alternativo , Redes Reguladoras de Genes , Análise de Sequência de RNA/métodos , Fatores de Transcrição/metabolismo , Animais , Linhagem Celular , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Células HEK293 , Humanos , Camundongos , Neurônios/citologia , Neurônios/metabolismo , RNA Mensageiro/genética
4.
Microb Pathog ; 187: 106527, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38163490

RESUMO

Abnormal activation of macrophage and gut Bacteroides fragilis (BF) are the important induction factors in the occurrence of type 2 diabetes (T2D) and vascular complications. However, it remains unknown whether BF involves in macrophage polarization. In this study, we found that BF extracellular vesicles (EV) can be uptaken by macrophage. BF-EV promote macrophage M1/M2 polarization significantly, and increase Sting expression significantly. Bioinformatics analysis found that Sema7a is an important gene involving in macrophage polarization. The expression of Sema7a can be induced by BF-EV and can be inhibited after C-176 treated. The inhibition expression of Sema7a prevent BF-EV to induce macrophage polarization. Further analysis reveals that there is no direct interaction between Sting and Sema7a, but Sgpl1 can interact with Sting or Sema7a. BF-EV promote the expression of Sgpl1, which the phenomenon can be inhibited after C-176 treated. Importantly, overexpression of Sgpl1 reversed the effect of C-176 for Sema7a expression, while inhibit Sema7a expression has limitation influence for Sting and Sgpl1 expression. In conclusion, this study confirms that Sting-Sgpl1-Sema7a is a key mechanism by which BF-EV regulates macrophage polarization.


Assuntos
Diabetes Mellitus Tipo 2 , Vesículas Extracelulares , Humanos , Bacteroides fragilis , Diabetes Mellitus Tipo 2/metabolismo , Macrófagos/metabolismo , Vesículas Extracelulares/metabolismo , Ativação de Macrófagos
5.
Cell ; 138(5): 898-910, 2009 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-19737518

RESUMO

Alternative splicing is a key process underlying the evolution of increased proteomic and functional complexity and is especially prevalent in the mammalian nervous system. However, the factors and mechanisms governing nervous system-specific alternative splicing are not well understood. Through a genome-wide computational and expression profiling strategy, we have identified a tissue- and vertebrate-restricted Ser/Arg (SR) repeat splicing factor, the neural-specific SR-related protein of 100 kDa (nSR100). We show that nSR100 regulates an extensive network of brain-specific alternative exons enriched in genes that function in neural cell differentiation. nSR100 acts by increasing the levels of the neural/brain-enriched polypyrimidine tract binding protein and by interacting with its target transcripts. Disruption of nSR100 prevents neural cell differentiation in cell culture and in the developing zebrafish. Our results thus reveal a critical neural-specific alternative splicing regulator, the evolution of which has contributed to increased complexity in the vertebrate nervous system.


Assuntos
Processamento Alternativo , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Encéfalo/citologia , Diferenciação Celular , Linhagem Celular , Humanos , Camundongos , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Proteínas Nucleares/química , Proteínas de Ligação a RNA/química , Fatores de Processamento de Serina-Arginina
6.
Artigo em Inglês | MEDLINE | ID: mdl-39393997

RESUMO

Gallbladder cancer (GBC) is the most common malignant tumor in the biliary system, characterized by high malignancy, aggressiveness, and poor prognosis. Early diagnosis holds paramount importance in ameliorating therapeutic outcomes. Presently, the clinical diagnosis of GBC primarily relies on clinical-radiological-pathological approach. However, there remains a potential for missed diagnosis and misdiagnose in the realm of clinical practice. We firstly analyzed the blood-based biomarkers, such as carcinoembryonic antigen and carbohydrate antigen 19-9. Subsequently, we evaluated the diagnostic performance of various imaging modalities, including ultrasound (US), endoscopic ultrasound (EUS), computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography/computed tomography (PET/CT) and pathological examination, emphasizing their strengths and limitations in detecting early-stage GBC. Furthermore, we explored the potential of emerging technologies, particularly artificial intelligence (AI) and liquid biopsy, to revolutionize GBC diagnosis. AI algorithms have demonstrated improved image analysis capabilities, while liquid biopsy offers the promise of non-invasive and real-time monitoring. However, the translation of these advancements into clinical practice necessitates further validation and standardization. The review highlighted the advantages and limitations of current diagnostic approaches and underscored the need for innovative strategies to enhance diagnostic accuracy of GBC. In addition, we emphasized the importance of multidisciplinary collaboration to improve early diagnosis of GBC and ultimately patient outcomes. This review endeavoured to impart fresh perspectives and insights into the early diagnosis of GBC.

7.
Genes Dev ; 29(8): 803-16, 2015 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-25877919

RESUMO

Embryonic stem cells are maintained in a self-renewing and pluripotent state by multiple regulatory pathways. Pluripotent-specific transcriptional networks are sequentially reactivated as somatic cells reprogram to achieve pluripotency. How epigenetic regulators modulate this process and contribute to somatic cell reprogramming is not clear. Here we performed a functional RNAi screen to identify the earliest epigenetic regulators required for reprogramming. We identified components of the SAGA histone acetyltransferase complex, in particular Gcn5, as critical regulators of reprogramming initiation. Furthermore, we showed in mouse pluripotent stem cells that Gcn5 strongly associates with Myc and that, upon initiation of somatic reprogramming, Gcn5 and Myc form a positive feed-forward loop that activates a distinct alternative splicing network and the early acquisition of pluripotency-associated splicing events. These studies expose a Myc-SAGA pathway that drives expression of an essential alternative splicing regulatory network during somatic cell reprogramming.


Assuntos
Processamento Alternativo , Reprogramação Celular/genética , Epigenômica , Histona Acetiltransferases/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Animais , Diferenciação Celular , Movimento Celular/genética , Células Cultivadas , Células-Tronco Embrionárias , Regulação da Expressão Gênica no Desenvolvimento , Histona Acetiltransferases/genética , Camundongos , Células-Tronco Pluripotentes , Interferência de RNA , Processamento Pós-Transcricional do RNA/genética
8.
PLoS Pathog ; 16(2): e1008307, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32069328

RESUMO

The ability of HIV-1 to evolve resistance to combined antiretroviral therapies (cARTs) has stimulated research into alternative means of controlling this infection. We assayed >60 modulators of RNA alternative splicing (AS) to identify new inhibitors of HIV-1 RNA processing-a segment of the viral lifecycle not targeted by current drugs-and discovered compound N-[4-chloro-3-(trifluoromethyl)phenyl]-7-nitro-2,1,3-benzoxadiazol-4-amine (5342191) as a potent inhibitor of both wild-type (Ba-L, NL4-3, LAI, IIIB, and N54) and drug-resistant strains of HIV-1 (IC50: ~700 nM) with no significant effect on cell viability at doses tested. 5342191 blocks expression of four essential HIV-1 structural and regulatory proteins (Gag, Env, Tat, and Rev) without affecting total protein synthesis of the cell. This response is associated with altered unspliced (US) and singly-spliced (SS) HIV-1 RNA accumulation (~60% reduction) and transport to the cytoplasm (loss of Rev) whereas parallel analysis of cellular RNAs revealed less than a 0.7% of host alternative splicing (AS) events (0.25-0.67% by ≥ 10-20%), gene expression (0.01-0.46% by ≥ 2-5 fold), and protein abundance (0.02-0.34% by ≥ 1.5-2 fold) being affected. Decreased expression of Tat, but not Gag/Env, upon 5342191 treatment was reversed by a proteasome inhibitor, suggesting that this compound alters the synthesis/degradation of this key viral factor. Consistent with an affect on HIV-1 RNA processing, 5342191 treatment of cells altered the abundance and phosphorylation of serine/arginine-rich splicing factor (SRSF) 1, 3, and 4. Despite the activation of several intracellular signaling pathways by 5342191 (Ras, MEK1/2-ERK1/2, and JNK1/2/3), inhibition of HIV-1 gene expression by this compound could be reversed by pre-treatment with either a G-protein α-subunit inhibitor or two different MEK1/2 inhibitors. These observations demonstrate enhanced sensitivity of HIV-1 gene expression to small changes in host RNA processing and highlights the potential of modulating host intracellular signaling as an alternative approach for controlling HIV-1 infection.


Assuntos
Processamento Alternativo/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Processamento Alternativo/fisiologia , Expressão Gênica/genética , Regulação Viral da Expressão Gênica/genética , Infecções por HIV , Soropositividade para HIV , HIV-1/fisiologia , Células HeLa , Humanos , MAP Quinase Quinase 1/metabolismo , MAP Quinase Quinase Quinase 2/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Processamento Pós-Transcricional do RNA/fisiologia , Splicing de RNA/genética , RNA Viral/genética , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/fisiologia , Bibliotecas de Moléculas Pequenas , Replicação Viral/fisiologia , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética
9.
Mol Cell ; 49(2): 262-72, 2013 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-23246434

RESUMO

At least half of the human genome is derived from repetitive elements, which are often lineage specific and silenced by a variety of genetic and epigenetic mechanisms. Using a transchromosomic mouse strain that transmits an almost complete single copy of human chromosome 21 via the female germline, we show that a heterologous regulatory environment can transcriptionally activate transposon-derived human regulatory regions. In the mouse nucleus, hundreds of locations on human chromosome 21 newly associate with activating histone modifications in both somatic and germline tissues, and influence the gene expression of nearby transcripts. These regions are enriched with primate and human lineage-specific transposable elements, and their activation corresponds to changes in DNA methylation at CpG dinucleotides. This study reveals the latent regulatory potential of the repetitive human genome and illustrates the species specificity of mechanisms that control it.


Assuntos
Cromossomos Humanos Par 21/genética , Elementos de DNA Transponíveis , Inativação Gênica , Ativação Transcricional , Animais , Cromossomos Humanos Par 21/metabolismo , Metilação de DNA , Feminino , Histonas/metabolismo , Humanos , Rim/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Especificidade de Órgãos , Ligação Proteica , Especificidade da Espécie , Testículo/metabolismo , Fatores de Transcrição/metabolismo , Iniciação da Transcrição Genética
10.
Parasitol Res ; 120(6): 1935-1942, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34002262

RESUMO

Type 2 diabetes mellitus (T2D) is a prevalent inflammation-related disease characterized by insulin resistance and elevated blood glucose levels. The high incidence rate of T2D in Western societies may be due to environmental conditions, including reduced worm exposure. In human and animal models, some helminths, such as Schistosoma, Nippostrongylus, Strongyloides, and Heligmosomoides, and their products reportedly ameliorate or prevent T2D progression. T2D induces adaptive immune pathways involved in the inhibition of type 1 immune responses, promotion of type 2 immune responses, and expansion of regulatory T cells and innate immune cells, such as macrophages, eosinophils, and group 2 innate lymphoid cells. Among immune cells expanded in T2DM, type 2 immune cells and macrophages are the most important and may have synergistic effects. The stimulation of host immunity by helminth infections also promotes interactions between the innate and adaptive immune systems. In this paper, we provide a comprehensive review of intestinal helminths' protective effects against T2D.


Assuntos
Imunidade Adaptativa , Diabetes Mellitus Tipo 2/complicações , Helmintíase/complicações , Helmintos/fisiologia , Imunidade Inata , Animais , Helmintíase/imunologia , Humanos
11.
Parasitol Res ; 120(11): 3851-3856, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34626234

RESUMO

This study aimed to explore the effect and mechanism underlying the role of the Schistosoma japonicum antigen of fatty acid-binding protein (SjFABP) on the growth of the schistosomula. SjFABP levels were evaluated by quantitative real-time polymerase chain reaction of samples of mice infected with S. japonicum; SjFABP was expressed and its levels gradually increased during all stages of S. japonicum schistosomula, including on 3, 10, 14, and 21 days of the growth process. Immunohistochemistry results demonstrated that SjFABP was distributed in the parenchyma, especially in the digestive tract of the S. japonicum schistosomula. RNA interference resulted in more than 60% knockdown of SjFABP leading to a reduction in length, volume, width, and area of the schistosomula as compared to control samples, as determined by light microscopy. Terminal deoxynucleotidyl transferase dUTP nick-end labeling detection further suggested that SjFABP knockdown resulted in increased apoptosis of schistosomes. Taken together, these results suggest that SjFABP may be related to the growth and survival of S. japonicum schistosomula, thereby representing a potential target for the treatment of schistosomiasis.


Assuntos
Schistosoma japonicum , Esquistossomose Japônica , Esquistossomose , Animais , Anticorpos Anti-Helmínticos , Proteínas de Ligação a Ácido Graxo/genética , Marcação In Situ das Extremidades Cortadas , Camundongos , Schistosoma japonicum/genética
12.
Mol Cell ; 46(6): 884-92, 2012 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-22749401

RESUMO

Alternative splicing plays a key role in the expansion of proteomic and regulatory complexity, yet the functions of the vast majority of differentially spliced exons are not known. In this study, we observe that brain and other tissue-regulated exons are significantly enriched in flexible regions of proteins that likely form conserved interaction surfaces. These proteins participate in significantly more interactions in protein-protein interaction (PPI) networks than other proteins. Using LUMIER, an automated PPI assay, we observe that approximately one-third of analyzed neural-regulated exons affect PPIs. Inclusion of these exons stimulated and repressed different partner interactions at comparable frequencies. This assay further revealed functions of individual exons, including a role for a neural-specific exon in promoting an interaction between Bridging Integrator 1 (Bin1)/Amphiphysin II and Dynamin 2 (Dnm2) that facilitates endocytosis. Collectively, our results provide evidence that regulated alternative exons frequently remodel interactions to establish tissue-dependent PPI networks.


Assuntos
Processamento Alternativo , Mapas de Interação de Proteínas , Proteínas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sítios de Ligação , Células Cultivadas , Dinamina II/genética , Dinamina II/metabolismo , Éxons , Células HEK293 , Humanos , Luciferases de Renilla/genética , Luciferases de Renilla/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas/genética , Proteômica , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
13.
Parasite Immunol ; 41(8): e12657, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31125444

RESUMO

AIMS: The aim of this study was to evaluate the effect of anti-CTLA-4 monoclonal antibody (mAb) on 26-kDa glutathione-S-transferase (GST) vaccine-induced immunity against Schistosoma japonicum infection. METHODS AND RESULTS: Mice immunized with GST before infection with S japonicum cercariae were injected with anti-CTLA-4 mAb. Worm reduction rate of GST was increased from 25.41% in mice with GST immunization to 52.48% in mice with GST plus anti-CTLA-4 mAb. The percentages of regulatory T cells (Tregs) were significantly higher following administration of both GST and anti-CTLA-4 mAb, or anti-CTLA-4 mAb alone. Elevated levels of IFN-γ, IL-2, IL-4 and IL-5 were observed. CONCLUSION: These results demonstrated that CTLA-4 may inhibit the protective effect of GST vaccine, and anti-CTLA-4 mAb may be used as an adjuvant to enhance the immune protection conferred by the GST vaccine by enhancing Th1- and Th2-type immune response.


Assuntos
Anticorpos Monoclonais/imunologia , Antígeno CTLA-4/imunologia , Glutationa Transferase/imunologia , Schistosoma japonicum/enzimologia , Esquistossomose Japônica/prevenção & controle , Adjuvantes Imunológicos/administração & dosagem , Animais , Anticorpos Monoclonais/administração & dosagem , Feminino , Glutationa Transferase/administração & dosagem , Glutationa Transferase/genética , Humanos , Imunização , Interleucina-2/genética , Interleucina-2/imunologia , Interleucina-4/genética , Interleucina-4/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Schistosoma japonicum/genética , Schistosoma japonicum/imunologia , Esquistossomose Japônica/imunologia , Esquistossomose Japônica/parasitologia , Linfócitos T Reguladores/imunologia , Vacinas/administração & dosagem , Vacinas/genética , Vacinas/imunologia
14.
Mol Cell ; 43(5): 843-50, 2011 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-21884984

RESUMO

Neurogenesis requires the concerted action of numerous genes that are regulated at multiple levels. However, how different layers of gene regulation are coordinated to promote neurogenesis is not well understood. We show that the neural-specific Ser/Arg repeat-related protein of 100 kDa (nSR100/SRRM4) negatively regulates REST (NRSF), a transcriptional repressor of genes required for neurogenesis. nSR100 directly promotes alternative splicing of REST transcripts to produce a REST isoform (REST4) with greatly reduced repressive activity, thereby activating expression of REST targets in neural cells. Conversely, REST directly represses nSR100 in nonneural cells to prevent the activation of neural-specific splicing events. Consistent with a critical role for nSR100 in the inhibition of REST activity, blocking nSR100 expression in the developing mouse brain impairs neurogenesis. Our results thus reveal a fundamental role for direct regulatory interactions between a splicing activator and transcription repressor in the control of the multilayered regulatory programs required for neurogenesis.


Assuntos
Processamento Alternativo , Neurogênese , Fatores de Transcrição/genética , Animais , Células Cultivadas , Camundongos , Camundongos Endogâmicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Splicing de RNA , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica
15.
Parasitol Res ; 118(7): 2287-2293, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31168702

RESUMO

Schistosomiasis is a devastating disease caused by Schistosoma infection. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has emerged as a candidate vaccine component against Schistosoma japonicum, but only confers partial protection. Cytotoxic T lymphocyte antigen-4 (CTLA-4) regulates T cell activation and shows negative effects on vaccine-induced immune protection; however, its potential influence on the protective effects of a GAPDH vaccine against S. japonicum and the underlying mechanism remain unclear. In this study, we established a mouse model of S. japonicum infection, and the mice were randomly divided into uninfected, infected control, anti-CTLA-4 monoclonal antibody (anti-CTLA-4 mAb), GAPDH, and GAPDH combined with anti-CTLA-4 mAb groups to compare the protective effects against infection and the consequent tissue damage. The worm reduction rate in the GAPDH-treated infected mice was 26.58%, which increased to 54.61% when combined with anti-CTLA-4 mAb. The frequency of regulatory T cells (Tregs) was significantly higher in the anti-CTLA-4 mAb group and was lower in the GAPDH group. However, both anti-CTLA-4 mAb and GAPDH elevated the levels of the cytokines IFN-γ, IL-2, IL-4, and IL-5 in the spleens of infected mice, and their combination further enhanced cytokine production. The diameter of egg granuloma in the anti-CTLA-4 mAb group and combined treatment group increased significantly compared to that of the other groups. These results suggest that anti-CTLA-4 mAb can be used as an adjuvant to enhance the immune protection of the GAPDH vaccine via inducing the Th1 immune response, although this comes at the cost of enhanced body injury.


Assuntos
Antígenos de Helmintos/imunologia , Antígeno CTLA-4/imunologia , Gliceraldeído-3-Fosfato Desidrogenases/imunologia , Schistosoma japonicum/imunologia , Esquistossomose Japônica/imunologia , Vacinas/imunologia , Animais , Anticorpos Monoclonais/imunologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Esquistossomose Japônica/parasitologia , Esquistossomose Japônica/prevenção & controle , Baço/imunologia , Linfócitos T Reguladores/imunologia
16.
Genes Dev ; 25(4): 373-84, 2011 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-21325135

RESUMO

Alternative splicing (AS) plays a major role in the generation of proteomic diversity and in gene regulation. However, the role of the basal splicing machinery in regulating AS remains poorly understood. Here we show that the core snRNP (small nuclear ribonucleoprotein) protein SmB/B' self-regulates its expression by promoting the inclusion of a highly conserved alternative exon in its own pre-mRNA that targets the spliced transcript for nonsense-mediated mRNA decay (NMD). Depletion of SmB/B' in human cells results in reduced levels of snRNPs and a striking reduction in the inclusion levels of hundreds of additional alternative exons, with comparatively few effects on constitutive exon splicing levels. The affected alternative exons are enriched in genes encoding RNA processing and other RNA-binding factors, and a subset of these exons also regulate gene expression by activating NMD. Our results thus demonstrate a role for the core spliceosomal machinery in controlling an exon network that appears to modulate the levels of many RNA processing factors.


Assuntos
Processamento Alternativo/genética , Processamento Alternativo/fisiologia , Precursores de RNA/metabolismo , Spliceossomos/metabolismo , Sequência de Bases , Sequência Conservada , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Modelos Biológicos , Dados de Sequência Molecular , Mutação/fisiologia , Processamento Pós-Transcricional do RNA/genética , Processamento Pós-Transcricional do RNA/fisiologia , Elementos Reguladores de Transcrição/fisiologia , Ribonucleoproteínas Nucleares Pequenas/genética , Ribonucleoproteínas Nucleares Pequenas/metabolismo , Ribonucleoproteínas Nucleares Pequenas/fisiologia , Spliceossomos/fisiologia , Transfecção , Proteínas Centrais de snRNP/genética , Proteínas Centrais de snRNP/metabolismo
17.
Development ; 142(15): 2564-73, 2015 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-26116661

RESUMO

Regulation of the balance between progenitor self-renewal and differentiation is crucial to development. In the mammalian kidney, reciprocal signalling between three lineages (stromal, mesenchymal and ureteric) ensures correct nephron progenitor self-renewal and differentiation. Loss of either the atypical cadherin FAT4 or its ligand Dachsous 1 (DCHS1) results in expansion of the mesenchymal nephron progenitor pool, called the condensing mesenchyme (CM). This has been proposed to be due to misregulation of the Hippo kinase pathway transcriptional co-activator YAP. Here, we use tissue-specific deletions to prove that FAT4 acts non-autonomously in the renal stroma to control nephron progenitors. We show that loss of Yap from the CM in Fat4-null mice does not reduce the expanded CM, indicating that FAT4 regulates the CM independently of YAP. Analysis of Six2(-/-);Fat4(-/-) double mutants demonstrates that excess progenitors in Fat4 mutants are dependent on Six2, a crucial regulator of nephron progenitor self-renewal. Electron microscopy reveals that cell organisation is disrupted in Fat4 mutants. Gene expression analysis demonstrates that the expression of Notch and FGF pathway components are altered in Fat4 mutants. Finally, we show that Dchs1, and its paralogue Dchs2, function in a partially redundant fashion to regulate the number of nephron progenitors. Our data support a model in which FAT4 in the stroma binds to DCHS1/2 in the mouse CM to restrict progenitor self-renewal.


Assuntos
Caderinas/metabolismo , Diferenciação Celular/fisiologia , Néfrons/ultraestrutura , Transdução de Sinais/fisiologia , Células-Tronco/citologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Análise de Variância , Animais , Proteínas de Ciclo Celular , Linhagem da Célula/fisiologia , Imunofluorescência , Perfilação da Expressão Gênica , Via de Sinalização Hippo , Immunoblotting , Hibridização In Situ , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Knockout , Microscopia Eletrônica , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Análise de Sequência de RNA , Proteínas de Sinalização YAP
18.
RNA ; 22(9): 1411-26, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27365209

RESUMO

Acinus (apoptotic chromatin condensation inducer in the nucleus) is an RNA-binding protein (RBP) originally identified for its role in apoptosis. It was later found to be an auxiliary component of the exon junction complex (EJC), which is deposited at exon junctions as a consequence of pre-mRNA splicing. To uncover the cellular functions of Acinus and investigate its role in splicing, we mapped its endogenous RNA targets using the cross-linking immunoprecipitation protocol (iCLIP). We observed that Acinus binds to pre-mRNAs, associating specifically to a subset of suboptimal introns, but also to spliced mRNAs. We also confirmed the presence of Acinus as a peripheral factor of the EJC. RNA-seq was used to investigate changes in gene expression and alternative splicing following siRNA-mediated depletion of Acinus in HeLa cells. This analysis revealed that Acinus is preferentially required for the inclusion of specific alternative cassette exons and also controls the faithful splicing of a subset of introns. Moreover, a large number of splicing changes can be related to Acinus binding, suggesting a direct role of Acinus in exon and intron definition. In particular, Acinus regulates the splicing of DFFA/ICAD transcript, a major regulator of DNA fragmentation. Globally, the genome-wide identification of RNA targets of Acinus revealed its role in splicing regulation as well as its involvement in other cellular pathways, including cell cycle progression. Altogether, this study uncovers new cellular functions of an RBP transiently associated with the EJC.


Assuntos
Processamento Alternativo , Proteínas Nucleares/metabolismo , RNA Mensageiro/metabolismo , Ciclo Celular , Células HeLa , Humanos , Proteínas Nucleares/genética , Ligação Proteica , RNA Mensageiro/genética , Transcriptoma
19.
Mol Cell ; 39(6): 925-38, 2010 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-20797886

RESUMO

Alternative splicing (AS) of pre-mRNA is utilized by higher eukaryotes to achieve increased transcriptome and proteomic complexity. The serine/arginine (SR) splicing factors regulate tissue- or cell-type-specific AS in a concentration- and phosphorylation-dependent manner. However, the mechanisms that modulate the cellular levels of active SR proteins remain to be elucidated. In the present study, we provide evidence for a role for the long nuclear-retained regulatory RNA (nrRNA), MALAT1 in AS regulation. MALAT1 interacts with SR proteins and influences the distribution of these and other splicing factors in nuclear speckle domains. Depletion of MALAT1 or overexpression of an SR protein changes the AS of a similar set of endogenous pre-mRNAs. Furthermore, MALAT1 regulates cellular levels of phosphorylated forms of SR proteins. Taken together, our results suggest that MALAT1 regulates AS by modulating the levels of active SR proteins. Our results further highlight the role for an nrRNA in the regulation of gene expression.


Assuntos
Processamento Alternativo/genética , Proteínas Nucleares/metabolismo , RNA não Traduzido/fisiologia , Proteínas de Ligação a RNA/metabolismo , Animais , Sítios de Ligação/genética , Linhagem Celular , Núcleo Celular/genética , Núcleo Celular/patologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células HeLa , Humanos , Espaço Intranuclear/metabolismo , Camundongos , Antígenos de Histocompatibilidade Menor , Mitose/genética , Proteínas Nucleares/genética , Fosforilação/fisiologia , Ligação Proteica/fisiologia , Domínios e Motivos de Interação entre Proteínas/genética , Precursores de RNA/metabolismo , Fatores de Processamento de RNA , RNA não Traduzido/genética , Proteínas de Ligação a RNA/genética , Sequências Reguladoras de Ácido Ribonucleico/genética , Fatores de Processamento de Serina-Arginina , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
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