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1.
Int J Biochem Cell Biol ; 113: 95-102, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31220582

RESUMO

ZnO nanoparticle (ZnO NP) exposure causes oxidative stress in the respiratory system, leading to pulmonary damage. Activating transcription factor 3 (ATF3) participates in a variety of cellular stress responses. However, the role of ATF3 in ZnO NP genotoxicity and cytotoxicity remains to be explored. Here we reported that ZnO NP treatment dramatically induced the expression of ATF3 in human bronchial epithelial (HBE) cells, which was mediated by the nuclear factor erythroid 2-related factor 2 (Nrf2). ATF3 was required for the repair of ZnO NP-induced DNA damage as gamma foci number increased when endogenous ATF3 was silenced. Moreover, ATF3 also contributed to ZnO NP-induced cell apoptosis. Mechanistic study revealed that ATF3 interacted with the p53 protein and upregulated its expression under ZnO NP treatment. Collectively, our findings demonstrated ATF3 as an important regulator of epithelial homeostasis by promoting both DNA repair and the death of damaged cells under ZnO NP-induced genotoxic stress.

2.
J Exp Clin Cancer Res ; 38(1): 217, 2019 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-31122284

RESUMO

BACKGROUND: Upregulation of RNA polymerase (Pol) III products, including tRNAs and 5S rRNA, in tumor cells leads to enhanced protein synthesis and tumor formation, making it a potential target for cancer treatment. In this study, we evaluated the inhibition of Pol III transcription by triptolide and the anti-cancer effect of this drug in colorectal tumorigenesis. METHODS: The effect of triptolide on colorectal cancer development was assessed in colorectal cancer mouse models, 3D organoids, and cultured cells. Colorectal cancer cells were treated with triptolide. Pol III transcription was measured by real-time quantitative polymerase chain reaction (PCR). The formation of TFIIIB, a multi-subunit transcription factor for Pol III, was determined by chromatin immunoprecipitation (ChIP), co-immunoprecipitation (Co-IP), and fluorescence resonance energy transfer (FRET). RESULTS: Triptolide reduced both tumor number and tumor size in adenomatous polyposis coli (Apc) mutated (ApcMin/+) mice as well as AOM/DSS-induced mice. Moreover, triptolide effectively inhibited colorectal cancer cell proliferation, colony formation, and organoid growth in vitro, which was associated with decreased Pol III target genes. Mechanistically, triptolide treatment blocked TBP/Brf1interaction, leading to the reduced formation of TFIIIB at the promoters of tRNAs and 5S rRNA. CONCLUSIONS: Together, our data suggest that inhibition of Pol III transcription with existing drugs such as triptolide provides a new avenue for developing novel therapies for colorectal cancer.

3.
Cell Death Dis ; 10(2): 127, 2019 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-30755584

RESUMO

Silica nanoparticles (SiNPs) have been reported to induce pulmonary fibrosis (PF) with an unknown mechanism. Recently, the activation of autophagy, a lysosome-dependent cell degradation pathway, by SiNPs has been identified in alveolar epithelial cells (AECs). However, the underlying mechanism and the relevance of SiNPs-induced autophagy to the development of PF remain elusive. Here, we report that autophagy dysfunction and subsequent apoptosis in AECs are involved in SiNPs-induced PF. SiNPs engulfed by AECs enhance autophagosome accumulation and apoptosis both in vivo and in vitro. Mechanically, SiNPs block autophagy flux through impairing lysosomal degradation via acidification inhibition. Lysosomal reacidification by cyclic-3',5'-adenosine monophosphate (cAMP) significantly enhances autophagic degradation and attenuate apoptosis. Importantly, enhancement of autophagic degradation by rapamycin protects AECs from apoptosis and attenuates SiNPs-induced PF in the mouse model. Altogether, our data demonstrate a repressive effect of SiNPs on lysosomal acidification, contributing to the decreased autophagic degradation in AECs, thus leading to apoptosis and subsequent PF. These findings may provide an improved understanding of SiNPs-induced PF and molecular targets to antagonize it.

4.
Angiogenesis ; 2018 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-30465119

RESUMO

The miR-200 family, consisting of miR-200a/b/c, miR-141, and miR-429, is well known to inhibit epithelial-to-mesenchymal transition (EMT) in cancer invasion and metastasis. Among the miR-200 family members, miR-200a/b/c and miR-429 have been reported to inhibit angiogenesis. However, the role of miR-141 in angiogenesis remains elusive, as contradicting results have been found in different cancer types and tumor models. Particularly, the effect of miR-141 in vascular endothelial cells has not been defined. In this study, we used several in vitro and in vivo models to demonstrate that miR-141 in endothelial cells inhibits angiogenesis. Additional mechanistic studies showed that miR-141 suppresses angiogenesis through multiple targets, including NRP1, GAB1, CXCL12ß, TGFß2, and GATA6, and bioinformatics analysis indicated that miR-141 and its targets comprise a powerful and precise regulatory network to modulate angiogenesis. Taken together, these data not only demonstrate an anti-angiogenic effect of miR-141, further strengthening the critical role of miR-200 family in the process of angiogenesis, but also provides a valuable cancer therapeutic target to control both angiogenesis and EMT, two essential steps in tumor growth and metastasis.

5.
Eur J Med Chem ; 162: 122-131, 2018 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-30445262

RESUMO

A series of novel fusidic acid (FA) derivatives were synthesized and screened for their in vitro cytotoxicity against the Hela, U87, KBV and MKN45 cancer cell lines. Selected FA derivatives with anti-tumor activity were firstly identified including compound 4, which exhibited good anti-proliferative activity with IC50 values in the range of 1.26-3.57 µM. Further research revealed that compound 4 induced Hela cells to undergo apoptosis by increasing the ratio of the cells in the Sub-G0/G1 phase via decreasing the neo-synthesized proteins in a dose-dependent manner from 1 to 10 µM. Compound 4 also showed good in vivo anti-tumor activity against the xenograft tumor of Hela cells and had no apparent toxicity. This study highlights the advantage of introducing the medium-length amino-terminal groups at the 3-OH position of FA to enhance its anti-tumor activity and suggests that compound 4 provides a starting point for designing more potent derivatives in the future.

6.
Gene ; 639: 111-116, 2018 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-29020616

RESUMO

Follistatin (FST), a single-chain glycosylated protein, is expressed in various tissues. The essential biological function of FST is binding and neutralizing transforming growth factor ß (TGF-ß) superfamily, including activin, myostatin, and bone morphogenetic protein (BMP). Emerging evidence indicates that FST also serves as a stress responsive protein, which plays a protective role under a variety of stresses. In most cases, FST performs the protective function through its neutralization of TGF-ß superfamily. However, under certain circumstances, FST translocates into the nucleus to maintain cellular homeostasis independent of its extracellular antagonism activity. This review provides integrated insight into the most recent advances in understanding the role of FST under various stresses, and the clinical implications corresponding to these findings and discusses the mechanisms to be further studied.


Assuntos
Folistatina/fisiologia , Estresse Fisiológico/fisiologia , Folistatina/genética , Folistatina/metabolismo , Humanos , Processamento de Proteína Pós-Traducional , Processamento Pós-Transcricional do RNA
7.
Mol Neurobiol ; 55(2): 1338-1351, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-28127696

RESUMO

Gene defects have been recognized as prominent factors in the etiology and pathogenesis of neurodegeneration. Among 60 neurodegeneration-related mutations in progranulin (PGRN), a mutation in PGRN gene exon 1 introduces a charged amino acid in the hydrophobic core of its signal peptide at residue 9 (named PGRN A9D) and results in incorrect cytoplasmic sorting. However, the pathogenesis of this mutation remains elusive. To address this issue, we first examined the subcellular distribution of PGRN A9D in human neuronal-like cells (SH-SY5Y). The results showed that PGRN A9D accumulated in cytosolic stress granules. Interestingly, this mis-sorting associated with a cellular redistribution of angiogenin (ANG), a stress-response factor and neurodegenerative disease-related protein, from nucleus to cytoplasmic stress granules, and there existed protein interaction between PGRN A9D and ANG. Further study revealed that the stress granule localization of PGRN A9D was dependent on ANG. Functionally, PGRN A9D abolished the nuclear ANG-mediated biological roles; on the other hand, the relocation of ANG to stress granules activated its cytoprotective stress-response program by cleaving transfer RNAs (tRNAs) to tiRNAs (tRNA-derived, stress-induced small RNAs), thus promoting PGRN A9D cell survival. Taken together, we hypothesize that PGRN A9D leads to the retention of ANG in the cytoplasm to protect cells from PGRN A9D-induced apoptosis, implying that PGRN and ANG act in concert to regulate the progress of neurodegenerative disease.

8.
Int J Biochem Cell Biol ; 95: 27-34, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29246685

RESUMO

Follistatin (FST) plays a protective role during silica nanoparticle (SiO2 NP) exposure. SiO2 NP treatment induces FST transcription with an unknown mechanism. We herein reported that SIRT6, one of the sirtuin family members, induced epigenetic silencing of FST. The expression of FST was elevated after SIRT6 knockdown while reduced after SIRT6 overexpression. Chromatin immunoprecipitation revealed a direct interaction between SIRT6 with FST promoter. Knockdown of SIRT6 increased both Ac-H3K9 level and Ac-H3K56 level at FST promoter region. SiO2 NP treatment de-stabilized SIRT6 mRNA and reduced SIRT6 expression, leading to the activation of FST transcription. Finally, over-expression of SIRT6 increased SiO2 NP-induced apoptosis. Collectively, this study provided evidence that SIRT6 is a negative regulator of FST transcription and participates in the regulation of cell survival during silica nanoparticle exposure.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Repressão Epigenética/efeitos dos fármacos , Folistatina/antagonistas & inibidores , Nanopartículas/toxicidade , Dióxido de Silício/toxicidade , Sirtuínas/metabolismo , Células A549 , Acetilação/efeitos dos fármacos , Células Epiteliais Alveolares/metabolismo , Animais , Folistatina/agonistas , Folistatina/genética , Folistatina/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Genes Reporter/efeitos dos fármacos , Células HEK293 , Histonas/metabolismo , Humanos , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Regiões Promotoras Genéticas/efeitos dos fármacos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Interferência de RNA , Estabilidade de RNA/efeitos dos fármacos , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Sirtuínas/antagonistas & inibidores , Sirtuínas/química , Sirtuínas/genética
9.
Sci Rep ; 6: 21133, 2016 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-26878911

RESUMO

Silica nanoparticles (SiO2 NPs) cause oxidative stress in respiratory system. Meanwhile, human cells launch adaptive responses to overcome SiO2 NP toxicity. However, besides a few examples, the regulation of SiO2 NP-responsive proteins and their functions in SiO2 NP response remain largely unknown. In this study, we demonstrated that SiO2 NP induced the expression of follistatin (FST), a stress responsive gene, in mouse lung tissue as well as in human lung epithelial cells (A549). The levels of Ac-H3(K9/18) and H3K4me2, two active gene markers, at FST promoter region were significantly increased during SiO2 NP treatment. The induction of FST transcription was mediated by the nuclear factor erythroid 2-related factor 2 (Nrf2), as evidenced by the decreased FST expression in Nrf2-deficient cells and the direct binding of Nrf2 to FST promoter region. Down-regulation of FST promoted SiO2 NP-induced apoptosis both in cultured cells and in mouse lung tissue. Furthermore, knockdown of FST increased while overexpression of FST decreased the expression level of NADPH oxidase 1 (NOX1) and NOX5 as well as the production of cellular reactive oxygen species (ROS). Taken together, these findings demonstrated a protective role of FST in SiO2 NP-induced oxidative stress and shed light on the interaction between SiO2 NPs and biological systems.


Assuntos
Células Epiteliais Alveolares/metabolismo , Folistatina/genética , Fator 2 Relacionado a NF-E2/metabolismo , Nanopartículas , Estresse Oxidativo , Dióxido de Silício , Ativação Transcricional , Animais , Apoptose/genética , Linhagem Celular , Expressão Gênica , Humanos , Masculino , Proteínas de Membrana/genética , Camundongos , NADPH Oxidase 1 , NADPH Oxidase 5 , NADPH Oxidases/genética , Nanopartículas/efeitos adversos , Espécies Reativas de Oxigênio/metabolismo , Dióxido de Silício/efeitos adversos
10.
Methods Mol Biol ; 1358: 303-16, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26463392

RESUMO

Regulation of translation initiation is a central control point in protein synthesis. Variations of start codon selection contribute to protein diversity and complexity. Systemic mapping of start codon positions and precise measurement of the corresponding initiation rate would transform our understanding of translational control. Here we describe a ribosome profiling approach that enables identification of translation initiation sites on a genome-wide scale. By capturing initiating ribosomes using lactimidomycin, this approach permits qualitative and quantitative analysis of alternative translation initiation.


Assuntos
Códon de Iniciação/genética , Biologia Molecular/métodos , Iniciação Traducional da Cadeia Peptídica , Biossíntese de Proteínas/genética , Processamento Alternativo/genética , RNA Mensageiro/genética , Ribossomos/genética
11.
Nature ; 526(7574): 591-4, 2015 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-26458103

RESUMO

The most abundant mRNA post-transcriptional modification is N(6)-methyladenosine (m(6)A), which has broad roles in RNA biology. In mammalian cells, the asymmetric distribution of m(6)A along mRNAs results in relatively less methylation in the 5' untranslated region (5'UTR) compared to other regions. However, whether and how 5'UTR methylation is regulated is poorly understood. Despite the crucial role of the 5'UTR in translation initiation, very little is known about whether m(6)A modification influences mRNA translation. Here we show that in response to heat shock stress, certain adenosines within the 5'UTR of newly transcribed mRNAs are preferentially methylated. We find that the dynamic 5'UTR methylation is a result of stress-induced nuclear localization of YTHDF2, a well-characterized m(6)A 'reader'. Upon heat shock stress, the nuclear YTHDF2 preserves 5'UTR methylation of stress-induced transcripts by limiting the m(6)A 'eraser' FTO from demethylation. Remarkably, the increased 5'UTR methylation in the form of m(6)A promotes cap-independent translation initiation, providing a mechanism for selective mRNA translation under heat shock stress. Using Hsp70 mRNA as an example, we demonstrate that a single m(6)A modification site in the 5'UTR enables translation initiation independent of the 5' end N(7)-methylguanosine cap. The elucidation of the dynamic features of 5'UTR methylation and its critical role in cap-independent translation not only expands the breadth of physiological roles of m(6)A, but also uncovers a previously unappreciated translational control mechanism in heat shock response.


Assuntos
Adenosina/análogos & derivados , Regulação da Expressão Gênica , Resposta ao Choque Térmico , Metilação , Iniciação Traducional da Cadeia Peptídica , RNA Mensageiro/metabolismo , Regiões 5' não Traduzidas/genética , Adenosina/metabolismo , Dioxigenase FTO Dependente de alfa-Cetoglutarato , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Proteínas de Choque Térmico HSP70/genética , Resposta ao Choque Térmico/genética , Camundongos , Oxigenases de Função Mista/antagonistas & inibidores , Oxigenases de Função Mista/metabolismo , Oxo-Ácido-Liases/antagonistas & inibidores , Oxo-Ácido-Liases/metabolismo , Capuzes de RNA/metabolismo , RNA Mensageiro/genética , Proteínas de Ligação a RNA/metabolismo , Transcrição Genética/genética
12.
Int J Biochem Cell Biol ; 65: 12-9, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25980932

RESUMO

Ribonuclease like 5 (Rnasel5) is a novel member of the zebrafish ribonuclease A family and its expression is increased during early embryogenesis. However, the in vivo biological function of Rnasel5 remains to be elucidated. Here, we report that knockdown of Rnasel5 by morhpolinos caused shrunken yolk extension as well as increased DNA damage at yolk syncytial layer and external tissue layers via the activation of p53 pathway. In addition, the morphological defects caused by Rnasel5 knockdown can be partially rescued by mRNA injection. Our findings provide the first functional characterization of Rnasel5 in zebrafish development and reveal its critical role in yolk extension by modulation of the p53 pathway.


Assuntos
Dano ao DNA , Ribonuclease Pancreático/metabolismo , Proteína Supressora de Tumor p53/genética , Saco Vitelino/fisiologia , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/fisiologia , Animais , Embrião não Mamífero , Desenvolvimento Embrionário/fisiologia , Técnicas de Silenciamento de Genes , Ribonuclease Pancreático/genética , Proteína Supressora de Tumor p53/metabolismo , Saco Vitelino/crescimento & desenvolvimento , Saco Vitelino/metabolismo , Proteínas de Peixe-Zebra/metabolismo
13.
Nat Struct Mol Biol ; 22(5): 404-10, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25866880

RESUMO

In response to stress, cells attenuate global protein synthesis but permit efficient translation of mRNAs encoding heat-shock proteins (HSPs). Although decades have passed since the first description of the heat-shock response, how cells achieve translational control of HSP synthesis remains enigmatic. Here we report an unexpected role for mitochondrial ribosomal protein L18 (MRPL18) in the mammalian cytosolic stress response. MRPL18 bears a downstream CUG start codon and generates a cytosolic isoform in a stress-dependent manner. Cytosolic MRPL18 incorporates into the 80S ribosome and facilitates ribosome engagement on mRNAs selected for translation during stress. MRPL18 knockdown has minimal effects on mitochondrial function but substantially dampens cytosolic HSP expression at the level of translation. Our results uncover a hitherto-uncharacterized stress-adaptation mechanism in mammalian cells, which involves formation of a 'hybrid' ribosome responsible for translational regulation during the cytosolic stress response.


Assuntos
Proteínas de Choque Térmico/biossíntese , Biossíntese de Proteínas/genética , Proteínas Ribossômicas/genética , Estresse Fisiológico/fisiologia , Linhagem Celular Tumoral , Códon de Iniciação/genética , Regulação da Expressão Gênica , Células HeLa , Resposta ao Choque Térmico/genética , Humanos , Fosforilação , Isoformas de Proteínas/genética , Interferência de RNA , RNA Interferente Pequeno , Proteínas Ribossômicas/metabolismo , Ribossomos/genética , eIF-2 Quinase/metabolismo
14.
Nat Methods ; 12(2): 147-53, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25486063

RESUMO

Cells have evolved exquisite mechanisms to fine-tune the rate of protein synthesis in response to stress. Systemic mapping of start-codon positions and precise measurement of the corresponding initiation rate would transform our understanding of translational control. Here we present quantitative translation initiation sequencing (QTI-seq), with which the initiating ribosomes can be profiled in real time at single-nucleotide resolution. Resultant initiation maps not only delineated variations of start-codon selection but also highlighted a dynamic range of initiation rates in response to nutrient starvation. The integrated data set provided unique insights into principles of alternative translation and mechanisms controlling different aspects of translation initiation. With RiboTag mice, QTI-seq permitted tissue-specific profiling of initiating ribosomes in vivo. Liver cell-specific ribosome profiling uncovered a robust translational reprogramming of the proteasome system in fasted mice. Our findings illuminated the prevalence and dynamic nature of translational regulation pivotal to physiological adaptation in vivo.


Assuntos
Biossíntese de Proteínas , RNA Mensageiro/metabolismo , Ribossomos/metabolismo , Animais , Técnicas de Cultura de Células , Meios de Cultura , Células HEK293 , Humanos , Fígado/metabolismo , Camundongos Transgênicos , Iniciação Traducional da Cadeia Peptídica/genética , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Biossíntese de Proteínas/genética , RNA Mensageiro/genética , Estresse Fisiológico/genética
15.
Nucleic Acids Res ; 42(16): 10720-30, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25159612

RESUMO

Follistatin (FST) performs several vital functions in the cells, including protection from apoptosis during stress. The expression of FST is up-regulated in response to glucose deprivation by an unknown mechanism. We herein showed that the induction of FST by glucose deprivation was due to an increase in the half-life of its mRNA. We further identified an AU-rich element (ARE) in the 3'UTR of FST mRNA that mediated its decay. The expression of FST was elevated after knocking down AUF1 and reduced when AUF1 was further expressed. In vitro binding assays and RNA pull-down assays revealed that AUF1 interacted with FST mRNA directly via its ARE. During glucose deprivation, a majority of AUF1 shuttled from cytoplasm to nucleus, resulting in dissociation of AUF1 from FST mRNA and thus stabilization of FST mRNA. Finally, knockdown of AUF1 decreased whereas overexpression of AUF1 increased glucose deprivation-induced apoptosis. The apoptosis promoting effect of AUF1 was eliminated in FST expressing cells. Collectively, this study provided evidence that AUF1 is a negative regulator of FST expression and participates in the regulation of cell survival under glucose deprivation.


Assuntos
Apoptose , Folistatina/genética , Glucose/fisiologia , Ribonucleoproteínas Nucleares Heterogêneas Grupo D/metabolismo , Estabilidade de RNA , RNA Mensageiro/metabolismo , Regiões 3' não Traduzidas , Proteínas Quinases Ativadas por AMP/metabolismo , Transporte Ativo do Núcleo Celular , Núcleo Celular/metabolismo , Folistatina/metabolismo , Células HeLa , Humanos
16.
Cell Res ; 24(7): 842-51, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24903108

RESUMO

The journey of a newly synthesized polypeptide starts in the peptidyltransferase center of the ribosome, from where it traverses the exit tunnel. The interior of the ribosome exit tunnel is neither straight nor smooth. How the ribosome dynamics in vivo is influenced by the exit tunnel is poorly understood. Genome-wide ribosome profiling in mammalian cells reveals elevated ribosome density at the start codon and surprisingly the downstream 5th codon position as well. We found that the highly focused ribosomal pausing shortly after initiation is attributed to the geometry of the exit tunnel, as deletion of the loop region from ribosome protein L4 diminishes translational pausing at the 5th codon position. Unexpectedly, the ribosome variant undergoes translational abandonment shortly after initiation, suggesting that there exists an obligatory step between initiation and elongation commitment. We propose that the post-initiation pausing of ribosomes represents an inherent signature of the translation machinery to ensure productive translation.


Assuntos
Iniciação Traducional da Cadeia Peptídica/fisiologia , Ribossomos/genética , Códon , Células HEK293 , Humanos , Proteínas Ribossômicas/genética
17.
J Cell Physiol ; 229(4): 521-9, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24122807

RESUMO

Angiogenin (ANG) undergoes nuclear translocation and promotes ribosomal RNA (rRNA) transcription thereby enhancing cell growth and proliferation. However, the mode of action of ANG in stimulating rRNA transcription is unclear. Here, we show that ANG enhances the formation of RNA polymerase I (Pol I) pre-initiation complex at the ribosomal DNA (rDNA) promoter. ANG binds at the upstream control element (UCE) of the promoter and enhances promoter occupancy of RNA Pol I as well as the selectivity factor SL1 components TAFI 48 and TAFI 110. We also show that ANG increases the number of actively transcribing rDNA by epigenetic activation through promoter methylation and histone modification. ANG binds to histone H3, inhibits H3K9 methylation, and activates H3K4 methylation as well as H4 acetylation at the rDNA promoter. These data suggest that one of the mechanisms by which ANG stimulates rRNA transcription is through an epigenetic activation of rDNA promoter.


Assuntos
DNA Ribossômico/metabolismo , Epigênese Genética/fisiologia , RNA Ribossômico/metabolismo , Ribonuclease Pancreático/metabolismo , Transcrição Genética/fisiologia , Metilação de DNA , DNA Ribossômico/genética , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HeLa , Histonas/metabolismo , Humanos , Regiões Promotoras Genéticas , Ligação Proteica , RNA Ribossômico/genética , Ribonuclease Pancreático/genética
18.
PLoS One ; 8(8): e71487, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23977052

RESUMO

Nuclear translocation of angiogenin (ANG) is essential for the proliferation of its target cells. ANG promotes rRNA synthesis, while whether it regulates mRNA transcription remains unknown. Using the chromatin immunoprecipitation method, we have identified 12 ANG-binding sequences. One of these sequences lies in the estrogen receptor-related receptor gamma (ERRγ) gene which we designated as ANG-Binding Sequence within ERRγ (ABSE). ABSE exhibited ANG-dependent repressor activity in the luciferase reporter system. Down-regulation of ANG increased ERRγ expression, and active gene marker level at the ABSE region. The expression levels of ERRγ targets genes, p21(WAF/CIP) and p27(KIP1), and the occupation of ERRγ on their promoter regions were increased in ANG-deficient cells accordingly. Furthermore, knockdown of ERRγ promoted the proliferation rate in ANG-deficient breast cancer cells. Finally, immunohistochemistry staining showed negative correlation between ANG and ERRγ in breast cancer tissue. Altogether, our study provides evidence that nuclear ANG directly binds to the ABSE of ERRγ gene and inhibits ERRγ transcription to promote breast cancer cell proliferation.


Assuntos
Receptores Estrogênicos/metabolismo , Ribonuclease Pancreático/metabolismo , Transcrição Genética , Sequência de Bases , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Imunoprecipitação da Cromatina , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Ligação Proteica/genética , Processamento de Proteína Pós-Traducional , RNA Polimerase II/metabolismo
19.
Arterioscler Thromb Vasc Biol ; 32(11): 2662-9, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22922963

RESUMO

OBJECTIVE: Abnormal proliferation and migration of vascular smooth muscle cells (SMCs) are the key events in the progression of neointima formation in response to vascular injury. The goal of this study is to investigate the functional role of a potent oncogene yes-associated protein (YAP) in SM phenotypic modulation in vitro and in vivo. METHODS AND RESULTS: In vitro cell culture and in vivo in both mouse and rat arterial injury models YAP expression is significantly induced and correlated with the vascular SMC synthetic phenotype. Overexpression of YAP promotes SMC migration and proliferation while attenuating SM contractile gene expression. Conversely, knocking down endogenous YAP in SMCs upregulates SM gene expression but attenuates SMC proliferation and migration. Consistent with this, knocking down YAP expression in a rat carotid balloon injury model and genetic deletion of YAP, specifically, in vascular SMCs in mouse after carotid artery ligation injury attenuates injury-induced SM phenotypic switch and neointima formation. CONCLUSIONS: YAP plays a novel integrative role in SM phenotypic modulation by inhibiting SM-specific gene expression while promoting SM proliferation and migration in vitro and in vivo. Blocking the induction of YAP would be a potential therapeutic approach for ameliorating vascular occlusive diseases.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Lesões das Artérias Carótidas/enzimologia , Músculo Liso Vascular/enzimologia , Miócitos de Músculo Liso/enzimologia , Fosfoproteínas/metabolismo , Lesões do Sistema Vascular/enzimologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Aorta/enzimologia , Aorta/patologia , Proteínas Reguladoras de Apoptose/genética , Artérias Carótidas/enzimologia , Artérias Carótidas/patologia , Lesões das Artérias Carótidas/genética , Lesões das Artérias Carótidas/patologia , Lesões das Artérias Carótidas/prevenção & controle , Movimento Celular , Proliferação de Células , Células Cultivadas , Modelos Animais de Doenças , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Genótipo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Neointima , Fenótipo , Fosfoproteínas/genética , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Fatores de Tempo , Transfecção , Lesões do Sistema Vascular/genética , Lesões do Sistema Vascular/patologia , Lesões do Sistema Vascular/prevenção & controle
20.
PLoS One ; 6(12): e28797, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22194915

RESUMO

Angiogenin (ANG) acts on both vascular endothelial cells and cancer cells, but the underlying mechanism remains elusive. In this study, we carried out a co-immunoprecipitation assay in HeLa cells and identified 14 potential ANG-interacting proteins. Among these proteins, ß-actin, α-actinin 4, and non-muscle myosin heavy chain 9 are stress fiber components and involved in cytoskeleton organization and movement, which prompted us to investigate the mechanism of action of ANG in cell migration. Upon confirmation of the interactions between ANG and the three proteins, further studies revealed that ANG co-localized with ß-actin and α-actinin 4 at the leading edge of migrating cells. Down-regulation of ANG resulted in fewer but thicker stress fibers with less dynamics, which was associated with the enlargements of focal adhesions. The focal adhesion kinase activity and cell migration capacity were significantly decreased in ANG-deficient cells. Taken together, our data demonstrated that the existence of ANG in the cytoplasm optimizes stress fiber assembly and focal adhesion formation to accommodate cell migration. The finding that ANG promoted cancer cell migration might provide new clues for tumor metastasis research.


Assuntos
Movimento Celular , Adesões Focais/metabolismo , Ribonuclease Pancreático/metabolismo , Fibras de Estresse/metabolismo , Citoesqueleto/metabolismo , Regulação para Baixo , Ativação Enzimática , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Células HeLa , Células Endoteliais da Veia Umbilical Humana/citologia , Humanos , Espectrometria de Massas , Anotação de Sequência Molecular , Fosforilação , Ligação Proteica , Pseudópodes/metabolismo , Ribonuclease Pancreático/deficiência
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