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1.
Front Immunol ; 10: 2449, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31824476

RESUMO

Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) characterized by mucosa damage associated with an uncontrolled inflammatory response. This immunological impairment leads to altered inflammatory mediators such as IL-33, which is shown to increase in the mucosa of active UC (aUC) patients. MicroRNAs present a distorted feature in inflamed colonic mucosa and are potential IL-33 regulating candidates in UC. Therefore, we studied the microRNA and mRNA profiles in inflamed colonic samples of UC patients, evaluating the effect of a microRNA (selected by in silico analysis and its expression in UC patients), on IL-33 under inflammatory conditions. We found that inflamed mucosa (n = 8) showed increased expression of 40 microRNAs and 2,120 mRNAs, while 49 microRNAs and 1,734 mRNAs were decreased, as determined by microarrays. In particular, IL-33 mRNA showed a 3.8-fold increase and eight members of a microRNA family (miR-378), which targets IL-33 mRNA in the 3'UTR, were decreased (-3.9 to -3.0 times). We selected three members of the miR-378 family (miR-378a-3p, miR-422a, and miR-378c) according to background information and interaction energy analysis, for further correlation analyses with IL-33 expression through qPCR and ELISA, respectively. We determined that aUC (n = 24) showed high IL-33 levels, and decreased expression of miR-378a-3p and miR-422a compared to inactive UC (n = 10) and controls (n = 6). Moreover, both microRNAs were inversely correlated with IL-33 expression, while miR-378c does not show a significant difference. To evaluate the effect of TNFα on the studied microRNAs, aUC patients with anti-TNF therapy were compared to aUC receiving other treatments. The levels of miR-378a-3p and miR-378c were higher in aUC patients with anti-TNF. Based on these findings, we selected miR-378a-3p to exploring the molecular mechanism involved by in vitro assays, showing that over-expression of miR-378a-3p decreased the levels of an IL-33 target sequence ß-gal-reporter gene in HEK293 cells. Stable miR-378a-3p over-expression/inhibition inversely modulated IL-33 content and altered viability of HT-29 cells. Additionally, in an inflammatory context, TNFα decreased miR-378a-3p levels in HT-29 cells enhancing IL-33 expression. Together, our results propose a regulatory mechanism of IL-33 expression exerted by miR-378a-3p in an inflammatory environment, contributing to the understanding of UC pathogenesis.

2.
Artigo em Inglês | MEDLINE | ID: mdl-31681621

RESUMO

Regulation of RNA homeostasis or "RNAstasis" is a central step in eukaryotic gene expression. From transcription to decay, cellular messenger RNAs (mRNAs) associate with specific proteins in order to regulate their entire cycle, including mRNA localization, translation and degradation, among others. The best characterized of such RNA-protein complexes, today named membraneless organelles, are Stress Granules (SGs) and Processing Bodies (PBs) which are involved in RNA storage and RNA decay/storage, respectively. Given that SGs and PBs are generally associated with repression of gene expression, viruses have evolved different mechanisms to counteract their assembly or to use them in their favor to successfully replicate within the host environment. In this review we summarize the current knowledge about the viral regulation of SGs and PBs, which could be a potential novel target for the development of broad-spectrum antiviral therapies.

3.
Cells ; 8(10)2019 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-31614895

RESUMO

Acquired immunodeficiency syndrome (AIDS) has become one of the most devastating pandemics in recorded history. The main causal agent of AIDS is the human immunodeficiency virus (HIV), which infects various cell types of the immune system that express the CD4 receptor on their surfaces. Today, combined antiretroviral therapy (cART) is the standard treatment for all people with HIV; although it has improved the quality of life of people living with HIV (PLWH), it cannot eliminate the latent reservoir of the virus. Therefore HIV/AIDS has turned from a fatal disease to a chronic disease requiring lifelong treatment. Despite significant viral load suppression, it has been observed that at least half of patients under cART present HIV-associated neurocognitive disorders (HAND), which have been related to HIV-1 infection and replication in the central nervous system (CNS). Several studies have focused on elucidating the mechanism by which HIV-1 can invade the CNS and how it can generate the effects seen in HAND. This review summarizes the research on HIV-1 and its interaction with the CNS with an emphasis on the generation of HAND, how the virus enters the CNS, the relationship between HIV-1 and cells of the CNS, and the effect of cART on these cells.

4.
J Fish Dis ; 42(7): 1035-1046, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31049989

RESUMO

We have previously shown that infectious pancreatic necrosis virus (IPNV) enters the embryo cell line CHSE-214 by macropinocytosis. In this study, we have extended our investigation into SHK-1 cells, a macrophage-like cell line derived from the head kidney of Atlantic salmon, the most economically important host of IPNV. We show that IPNV infection stimulated fluid uptake in SHK-1 cells above the constitutive macropinocytosis level. In addition, upon infection of SHK-1 cells, IPNV produced several changes in actin dynamics, such as protrusions and ruffles, which are important features of macropinocytosis. We also observed that the Na+/H+ pump inhibitor EIPA blocked IPNV infection. On the other hand, IPNV entry was independent of clathrin, a possibility that could not be ruled out in CHSE 214 cells. In order to determine the possible role of accessory factors on the macropinocytic process, we tested several inhibitors that affect components of transduction pathways. While pharmacological intervention of PKI3, PAK-1 and Rac1 did not affect IPNV infection, inhibition of Ras and Rho GTPases as well as Cdc42 resulted in a partial decrease in IPNV infection. Further studies will be required to determine the signalling pathway involved in the macropinocytosis-mediated entry of IPNV into its target cells.


Assuntos
Vírus da Necrose Pancreática Infecciosa/fisiologia , Macrófagos/virologia , Pinocitose , Salmão/virologia , Internalização do Vírus , Actinas/metabolismo , Animais , Infecções por Birnaviridae/virologia , Linhagem Celular , Doenças dos Peixes/virologia , Rim Cefálico/virologia , Macrófagos/citologia
5.
Biochim Biophys Acta Gene Regul Mech ; 1862(6): 657-669, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31075539

RESUMO

Variation in Disrupted-in-Schizophrenia 1 (DISC1) increases the risk for neurodegenerative diseases, schizophrenia, and other mental disorders. However, the functions of DISC1 associated with the development of these diseases remain unclear. DISC1 has been reported to inhibit Akt/mTORC1 signaling, a major regulator of translation, and recent studies indicate that DISC1 could exert a direct role in translational regulation. Here, we present evidence of a novel role of DISC1 in the maintenance of protein synthesis during oxidative stress. In order to investigate DISC1 function independently of Akt/mTORC1, we used Tsc2-/- cells, where mTORC1 activation is independent of Akt. DISC1 knockdown enhanced inhibition of protein synthesis in cells treated with sodium arsenite (SA), an oxidative agent used for studying stress granules (SGs) dynamics and translational control. N-acetyl-cysteine inhibited the effect of DISC1, suggesting that DISC1 affects translation in response to oxidative stress. DISC1 decreased SGs number in SA-treated cells, but resided outside SGs and maintained protein synthesis independently of a proper SG nucleation. DISC1-dependent stimulation of translation in SA-treated cells was supported by its interaction with eIF3h, a component of the canonical translation initiation machinery. Consistent with a role in the homeostatic maintenance of translation, DISC1 knockdown or overexpression decreased cell viability after SA exposure. Our data suggest that DISC1 is a relevant component of the cellular response to stress, maintaining certain levels of translation and preserving cell integrity. This novel function of DISC1 might be involved in its association with pathologies affecting tissues frequently exposed to oxidative stress.


Assuntos
Arsenitos/farmacologia , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Compostos de Sódio/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Grânulos Citoplasmáticos/metabolismo , DNA Helicases/metabolismo , Fator de Iniciação 3 em Eucariotos/metabolismo , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Proteínas do Tecido Nervoso/genética , Proteína Oncogênica v-akt , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , RNA Helicases/metabolismo , Proteínas com Motivo de Reconhecimento de RNA/metabolismo , Transcriptoma , Proteína 2 do Complexo Esclerose Tuberosa/genética
6.
Nucleic Acids Res ; 46(21): 11539-11552, 2018 11 30.
Artigo em Inglês | MEDLINE | ID: mdl-30239828

RESUMO

Gag synthesis from the full-length unspliced mRNA is critical for the production of the viral progeny during human immunodeficiency virus type-1 (HIV-1) replication. While most spliced mRNAs follow the canonical gene expression pathway in which the recruitment of the nuclear cap-binding complex (CBC) and the exon junction complex (EJC) largely stimulates the rates of nuclear export and translation, the unspliced mRNA relies on the viral protein Rev to reach the cytoplasm and recruit the host translational machinery. Here, we confirm that Rev ensures high levels of Gag synthesis by driving nuclear export and translation of the unspliced mRNA. These functions of Rev are supported by the CBC subunit CBP80, which binds Rev and the unspliced mRNA in the nucleus and the cytoplasm. We also demonstrate that Rev interacts with the DEAD-box RNA helicase eIF4AI, which translocates to the nucleus and cooperates with the viral protein to promote Gag synthesis. Finally, we show that the Rev/RRE axis is important for the assembly of a CBP80-eIF4AI complex onto the unspliced mRNA. Together, our results provide further evidence towards the understanding of the molecular mechanisms by which Rev drives Gag synthesis from the unspliced mRNA during HIV-1 replication.


Assuntos
Fator de Iniciação 4A em Eucariotos/genética , HIV-1/genética , Complexo Proteico Nuclear de Ligação ao Cap/genética , RNA Mensageiro/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética , Produtos do Gene rev do Vírus da Imunodeficiência Humana/genética , Linhagem Celular , Fator de Iniciação 4A em Eucariotos/metabolismo , HIV-1/metabolismo , Células HeLa , Humanos , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Complexo Proteico Nuclear de Ligação ao Cap/metabolismo , Ligação Proteica , Processamento de RNA , RNA Mensageiro/metabolismo , RNA Viral/genética , RNA Viral/metabolismo , Replicação Viral/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/biossíntese , Produtos do Gene rev do Vírus da Imunodeficiência Humana/metabolismo
7.
Front Microbiol ; 9: 576, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29643844

RESUMO

N6-methyladenosine (m6A) is the most abundant internal modification present in Eukaryotic mRNA. The functions of this chemical modification are mediated by m6A-binding proteins (m6A readers) and regulated by methyltransferases (m6A writers) and demethylases (m6A erasers), which together are proposed to be responsible of a new layer of post-transcriptional control of gene expression. Despite the presence of m6A in a retroviral genome was reported more than 40 years ago, the recent development of sequencing-based technologies allowing the mapping of m6A in a transcriptome-wide manner made it possible to identify the topology and dynamics of m6A during replication of HIV-1 as well as other viruses. As such, three independent groups recently reported the presence of m6A along the HIV-1 genomic RNA (gRNA) and described the impact of cellular m6A writers, erasers and readers on different steps of viral RNA metabolism and replication. Interestingly, while two groups reported a positive role of m6A at different steps of viral gene expression it was also proposed that the presence of m6A within the gRNA reduces viral infectivity by inducing the early degradation of the incoming viral genome. This review summarizes the recent advances in this emerging field and discusses the relevance of m6A during HIV-1 replication.

8.
J Virol Methods ; 255: 14-22, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29425681

RESUMO

Self-inactivating VSVG-pseudotyped murine leukemia virus (SIN-VSVG-MLV) has been widely used to generate stable cell lines and produce gene delivery vectors. Despite the broad cellular tropism of the VSVG-pseudotyped MLV, we observed differential viral transduction efficiency depending on the host cell type used. In order to determine the mechanism underlying these differences, we used a GFP-expressing SIN-VSVG-MLV and analyzed the major steps of viral transduction in different cell lines including human epithelial, T-lymphocytes, monocytes and murine fibroblast cells. We observed the better transduction efficiency in HeLa cells, which was 20-fold higher than THP-1 and NIH/3T3 cells. To quantify viral internalization, we determined genomic RNA content by quantifying the early reverse transcription product. Genomic RNA and transduction levels were correlated with HeLa cells showing the higher amount of early RT product followed by tsA201 cells, while NIH/3T3, Jurkat and THP-1 had the lowest amounts. Similar results were observed when the late reverse transcription product was analyzed. Reverse transcription efficiency was 66-85% in HeLa cells and about 30% in tsA201, NIH/3T3, Jurkat and THP-1 cells. Viral integration, determined by Alu-Nested-qPCR, was higher for HeLa and lowerst for Jurkat and THP-1 cells. Interestingly, we observed that viral entry was correlated with the cellular availability of clathrin-mediated endocytosis, which was higher in HeLa and tsA201 cells, potentially explaining the higher rates of SIN-VSVG-MLV transduction and early RT synthesis observed in these cell lines. In conclusion, the SIN-VSVG-MLV vector showed significantly different rates of infectivity depending on the host cell type, possibly due to differential rates of viral internalization.


Assuntos
Vetores Genéticos/genética , Vírus da Leucemia Murina/genética , Vírus da Leucemia Murina/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas do Envelope Viral/metabolismo , Internalização do Vírus , Animais , Linhagem Celular , Humanos , Vírus da Leucemia Murina/fisiologia , Glicoproteínas de Membrana/genética , Camundongos , Transdução Genética , Proteínas do Envelope Viral/genética
9.
Sci Rep ; 7(1): 3068, 2017 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-28596575

RESUMO

Infectious pancreatic necrosis virus (IPNV) is a non-enveloped virus belonging to the Birnaviridae family. IPNV produces an acute disease in salmon fingerlings, with high mortality rates and persistent infection in survivors. Although there are reports of IPNV binding to various cells, the viral receptor and entry pathways remain unknown. The aim of this study was to determine the endocytic pathway that allows for IPNV entry. We observed that IPNV stimulated fluid uptake and virus particles co-localysed with the uptake marker dextran in intracellular compartments, suggesting a role for macropinocytosis in viral entry. Consistent with this idea, viral infection was significantly reduced when the Na+/H+ exchanger NHE1 was inhibited with 5-(N-Ethyl-N-isopropyl) amiloride (EIPA). Neither chlorpromazine nor filipin complex I affected IPNV infection. To examine the role of macropinocytosis regulators, additional inhibitors were tested. Inhibitors of the EGFR pathway and the effectors Pak1, Rac1 and PKC reduced viral infection. Together, our results indicate that IPNV is mainly internalized into CHSE-214 cells by macropinocytosis.


Assuntos
Vírus da Necrose Pancreática Infecciosa/fisiologia , Pinocitose , Internalização do Vírus , Animais , Infecções por Birnaviridae/virologia , Caveolinas/metabolismo , Linhagem Celular , Clorpromazina/farmacologia , Dinaminas/metabolismo , Endocitose , Doenças dos Peixes/tratamento farmacológico , Doenças dos Peixes/virologia , Microdomínios da Membrana , Pinocitose/efeitos dos fármacos , Salmão/virologia , Internalização do Vírus/efeitos dos fármacos
10.
Biochim Biophys Acta Gene Regul Mech ; 1860(4): 460-471, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28219769

RESUMO

RNA plays central roles in biology and novel functions and regulation mechanisms are constantly emerging. To accomplish some of their functions within the cell, RNA molecules undergo hundreds of chemical modifications from which N6-methyladenosine (m6A), inosine (I), pseudouridine (ψ) and 5-methylcytosine (5mC) have been described in eukaryotic mRNA. Interestingly, the m6A modification was shown to be reversible, adding novel layers of regulation of gene expression through what is now recognized as epitranscriptomics. The development of molecular mapping strategies coupled to next generation sequencing allowed the identification of thousand of modified transcripts in different tissues and under different physiological conditions such as viral infections. As intracellular parasites, viruses are confronted to cellular RNA modifying enzymes and, as a consequence, viral RNA can be chemically modified at some stages of the replication cycle. This review focuses on the chemical modifications of viral RNA and the impact that these modifications have on viral gene expression and the output of infection. A special emphasis is given to m6A, which was recently shown to play important yet controversial roles in different steps of the HIV-1, HCV and ZIKV replication cycles.


Assuntos
Epigênese Genética , Transcriptoma/genética , Replicação Viral/genética , Animais , Humanos , Modelos Genéticos , Edição de RNA/genética
11.
Nucleic Acids Res ; 45(8): 4810-4824, 2017 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-28077561

RESUMO

MicroRNAs (miRNAs) are small non-coding RNAs that control gene expression by recognizing and hybridizing to a specific sequence generally located in the 3΄ untranslated region (UTR) of targeted mRNAs. miRNA-induced inhibition of translation occurs during the initiation step, most probably at the level of ribosome scanning. In this process, the RNA-induced silencing complex interacts both with PABP and the 43S pre-initiation complex to disrupt scanning of the 40S ribosome. However, in some specific cases, miRNAs can stimulate translation. Although the mechanism of miRNA-mediated upregulation is unknown, it appears that the poly(A) tail and the lack of availability of the TNRC6 proteins are amongst major determinants. The genomic RNA of the Hepatitis C Virus is uncapped, non-polyadenylated and harbors a peculiar internal ribosome entry site (IRES) that binds the ribosome directly to the AUG codon. Thus, we have exploited the unique properties of the HCV IRES and other related IRESes (HCV-like) to study how translation initiation can be modulated by miRNAs on these elements. Here, we report that miRNA binding to the 3΄ UTR can stimulate translation of a reporter gene given that its expression is driven by an HCV-like IRES and that it lacks a poly(A) tail at its 3΄ extremity.


Assuntos
Hepacivirus/genética , Sítios Internos de Entrada Ribossomal/genética , MicroRNAs/genética , Iniciação Traducional da Cadeia Peptídica , Códon/genética , Regulação da Expressão Gênica , Hepatite C/genética , Hepatite C/virologia , Humanos , MicroRNAs/biossíntese , Proteína I de Ligação a Poli(A)/genética , Biossíntese de Proteínas/genética , RNA Viral/biossíntese , RNA Viral/genética , Subunidades Ribossômicas Menores de Eucariotos/genética
12.
Viruses ; 8(11)2016 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-27886048

RESUMO

The human immunodeficiency virus type-1 (HIV-1) unspliced transcript is used both as mRNA for the synthesis of structural proteins and as the packaged genome. Given the presence of retained introns and instability AU-rich sequences, this viral transcript is normally retained and degraded in the nucleus of host cells unless the viral protein REV is present. As such, the stability of the HIV-1 unspliced mRNA must be particularly controlled in the nucleus and the cytoplasm in order to ensure proper levels of this viral mRNA for translation and viral particle formation. During its journey, the HIV-1 unspliced mRNA assembles into highly specific messenger ribonucleoproteins (mRNPs) containing many different host proteins, amongst which are well-known regulators of cytoplasmic mRNA decay pathways such as up-frameshift suppressor 1 homolog (UPF1), Staufen double-stranded RNA binding protein 1/2 (STAU1/2), or components of miRNA-induced silencing complex (miRISC) and processing bodies (PBs). More recently, the HIV-1 unspliced mRNA was shown to contain N6-methyladenosine (m6A), allowing the recruitment of YTH N6-methyladenosine RNA binding protein 2 (YTHDF2), an m6A reader host protein involved in mRNA decay. Interestingly, these host proteins involved in mRNA decay were shown to play positive roles in viral gene expression and viral particle assembly, suggesting that HIV-1 interacts with mRNA decay components to successfully accomplish viral replication. This review summarizes the state of the art in terms of the interactions between HIV-1 unspliced mRNA and components of different host mRNA decay machineries.


Assuntos
HIV-1/genética , Interações Hospedeiro-Patógeno , Estabilidade de RNA , RNA Viral/metabolismo , Humanos
13.
Viruses ; 8(7)2016 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-27367717

RESUMO

After viral infection, host cells respond by mounting an anti-viral stress response in order to create a hostile atmosphere for viral replication, leading to the shut-off of mRNA translation (protein synthesis) and the assembly of RNA granules. Two of these RNA granules have been well characterized in yeast and mammalian cells, stress granules (SGs), which are translationally silent sites of RNA triage and processing bodies (PBs), which are involved in mRNA degradation. This review discusses the role of these RNA granules in the evasion of anti-viral stress responses through virus-induced remodeling of cellular ribonucleoproteins (RNPs).


Assuntos
Grânulos Citoplasmáticos/metabolismo , Interações Hospedeiro-Patógeno , RNA Viral/metabolismo , Estresse Fisiológico , Vírus/crescimento & desenvolvimento , Vírus/imunologia , Biossíntese de Proteínas
14.
Biochim Biophys Acta ; 1859(5): 719-30, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27012366

RESUMO

DEAD-box RNA helicase DDX3 is a host factor essential for HIV-1 replication and thus, a potential target for novel therapies aimed to overcome viral resistance. Previous studies have shown that DDX3 promotes nuclear export and translation of the HIV-1 unspliced mRNA. Although the function of DDX3 during both processes requires its catalytic activity, it is unknown whether other domains surrounding the helicase core are involved. Here, we show the involvement of the N- and C-terminal domains of DDX3 in the regulation of HIV-1 unspliced mRNA translation. Our results suggest that the intrinsically disordered N-terminal domain of DDX3 regulates its functions in translation by acting prior to the recruitment of the 43S pre-initiation complex onto the viral 5'-UTR. Interestingly, this regulation was conserved in HIV-2 and was dependent on the CRM1-dependent nuclear export pathway suggesting a role of the RNA helicase in interconnecting nuclear export with ribosome recruitment of the viral unspliced mRNA. This specific function of DDX3 during HIV gene expression could be exploited as an alternative target for pharmaceutical intervention.


Assuntos
RNA Helicases DEAD-box/genética , Infecções por HIV/genética , HIV-1/genética , Carioferinas/genética , Receptores Citoplasmáticos e Nucleares/genética , Transporte Ativo do Núcleo Celular/genética , Regulação Viral da Expressão Gênica , Infecções por HIV/terapia , Infecções por HIV/virologia , HIV-1/patogenicidade , Interações Hospedeiro-Patógeno/genética , Humanos , Biossíntese de Proteínas , Estrutura Terciária de Proteína , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Viral/genética , Replicação Viral/genética
15.
Biomolecules ; 5(4): 2808-39, 2015 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-26492277

RESUMO

Unspliced, genomic HIV-1 RNA (vRNA) is a component of several ribonucleoprotein complexes (RNP) during the viral replication cycle. In earlier work, we demonstrated that the host upframeshift protein 1 (UPF1), a key factor in nonsense-mediated mRNA decay (NMD), colocalized and associated to the viral structural protein Gag during viral egress. In this work, we demonstrate a new function for UPF1 in the regulation of vRNA nuclear export. OPEN ACCESS Biomolecules 2015, 5 2809 We establish that the nucleocytoplasmic shuttling of UPF1 is required for this function and demonstrate that UPF1 exists in two essential viral RNPs during the late phase of HIV-1 replication: the first, in a nuclear export RNP that contains Rev, CRM1, DDX3 and the nucleoporin p62, and the second, which excludes these nuclear export markers but contains Gag in the cytoplasm. Interestingly, we observed that both UPF2 and the long isoform of UPF3a, UPF3aL, but not the shorter isoforms UPF3aS and UPF3b, are excluded from the UPF1-Rev-CRM1-DDX3 complex as they are negative regulators of vRNA nuclear export. In silico protein-protein docking analyses suggest that Rev binds UPF1 in a region that overlaps the UPF2 binding site, thus explaining the exclusion of this negative regulatory factor by HIV-1 that is necessary for vRNA trafficking. This work uncovers a novel and unique regulatory circuit involving several UPF proteins that ultimately regulate vRNA nuclear export and trafficking.


Assuntos
Núcleo Celular/metabolismo , HIV-1/fisiologia , RNA Viral/metabolismo , Transativadores/metabolismo , Replicação Viral , Transporte Ativo do Núcleo Celular , Núcleo Celular/virologia , RNA Helicases DEAD-box/metabolismo , HIV-1/metabolismo , Células HeLa , Humanos , Carioferinas/metabolismo , Glicoproteínas de Membrana/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Ligação Proteica , RNA Helicases , Proteínas de Ligação a RNA/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Liberação de Vírus , Produtos do Gene rev do Vírus da Imunodeficiência Humana/metabolismo
16.
Viruses ; 7(8): 4326-51, 2015 Aug 04.
Artigo em Inglês | MEDLINE | ID: mdl-26247956

RESUMO

Post-transcriptional control in both HIV-1 and HIV-2 is a highly regulated process that commences in the nucleus of the host infected cell and finishes by the expression of viral proteins in the cytoplasm. Expression of the unspliced genomic RNA is particularly controlled at the level of RNA splicing, export, and translation. It appears increasingly obvious that all these steps are interconnected and they result in the building of a viral ribonucleoprotein complex (RNP) that must be efficiently translated in the cytosolic compartment. This review summarizes our knowledge about the genesis, localization, and expression of this viral RNP.


Assuntos
Regulação Viral da Expressão Gênica , HIV-1/fisiologia , HIV-2/fisiologia , Interações Hospedeiro-Patógeno , Biossíntese de Proteínas , RNA Viral/metabolismo , Replicação Viral , Núcleo Celular/metabolismo , Citoplasma/metabolismo , HIV-1/genética , HIV-2/genética , Humanos , Ribonucleoproteínas/metabolismo , Proteínas Virais/metabolismo
17.
Rev Med Virol ; 25(5): 286-99, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26174373

RESUMO

Asp-Glu-Ala-Asp (DEAD)-box polypeptide 3, or DDX3, belongs to the DEAD-box family of ATP-dependent RNA helicases and is known to play different roles in RNA metabolism ranging from transcription to nuclear export, translation, and assembly of stress granules. In addition, there is growing evidence that DDX3 is a component of the innate immune response against viral infections. As such, DDX3 has been shown to play roles both upstream and downstream of I-kappa beta kinase ε (IKKε)/TANK-binding kinase 1, leading to IFN-ß production. Interestingly, several RNA viruses, including human threats such as HIV-1 and hepatitis C virus, hijack DDX3 to accomplish various steps of their replication cycles. Thus, it seems that viruses have evolved to exploit DDX3's functions while threatening the innate immune response. Understanding this interesting dichotomy in DDX3 function will help us not only to improve our knowledge of virus-host interactions but also to develop novel antiviral drugs targeting the multifaceted roles of DDX3 in viral replication.


Assuntos
RNA Helicases DEAD-box/metabolismo , Interações Hospedeiro-Patógeno , Interferon beta/metabolismo , RNA Helicases/metabolismo , Vírus de RNA/imunologia , Vírus de RNA/fisiologia , Replicação Viral , Humanos , Imunidade Inata
18.
PLoS One ; 10(4): e0123029, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25830243

RESUMO

We have previously shown a functional interaction between human papillomavirus type 16 (HPV-16) E6 and E7 oncoproteins and cigarette smoke condensate (CSC) in lung cells suggesting cooperation during carcinogenesis. The molecular mechanisms of such interaction, however, remain to be elucidated. Here we first present evidence showing that cigarette smoke condensate (CSC) has the ability to activate the HPV-16 p97 promoter by acting on the long control region (LCR) in lung epithelial cells. Interestingly, we observed that CSC-induced p97 promoter activation occurs in a dose-dependent manner in both tumor A-549 (lung adenocarcinoma), H-2170 (bronchial carcinoma), SiHa or Hela (cervical carcinoma) cells but not in non-tumor BEAS-2B (bronchial) or NL-20 (alveolar) lung cells unless they ectopically expressed the HPV-16 E6 and E7 oncogenes. In addition, we also observed a significant increase of primary DNA damage in tumor and non-tumor CSC-treated lung cells expressing HPV-16 E6 and E7 oncogenes suggesting a cooperative effect in this process, even though the contribution of E7 was significantly higher. Taken together, our results strongly suggest that tobacco smoke is able to induce the activation of the HPV-16 p97 promoter in cooperation with HPV-16 E6 and E7 oncogenes that, in turn, sensitize lung cells to tobacco smoke-induced DNA damage.


Assuntos
Papillomavirus Humano 16/genética , Neoplasias Pulmonares/virologia , Proteínas Oncogênicas Virais/genética , Proteínas E7 de Papillomavirus/genética , Proteínas Repressoras/genética , Fumar/efeitos adversos , Células Epiteliais Alveolares/patologia , Células Epiteliais Alveolares/virologia , Linhagem Celular Tumoral , Dano ao DNA , Regulação Viral da Expressão Gênica , Humanos , Neoplasias Pulmonares/genética , Proteínas Oncogênicas Virais/metabolismo , Oxirredução , Proteínas E7 de Papillomavirus/metabolismo , Infecções por Papillomavirus/virologia , Regiões Promotoras Genéticas , Proteínas Repressoras/metabolismo , Risco
19.
Nucleic Acids Res ; 42(20): 12861-75, 2014 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-25352557

RESUMO

During the post-transcriptional events of the HIV-2 replication cycle, the full-length unspliced genomic RNA (gRNA) is first used as an mRNA to synthesize Gag and Gag-Pol proteins and then packaged into progeny virions. However, the mechanisms responsible for the coordinate usage of the gRNA during these two mutually exclusive events are poorly understood. Here, we present evidence showing that HIV-2 expression induces stress granule assembly in cultured cells. This contrasts with HIV-1, which interferes with stress granules assembly even upon induced cellular stress. Moreover, we observed that the RNA-binding protein and stress granules assembly factor TIAR associates with the gRNA to form a TIAR-HIV-2 ribonucleoprotein (TH2RNP) complex localizing diffuse in the cytoplasm or aggregated in stress granules. Although the assembly of TH2RNP in stress granules did not require the binding of the Gag protein to the gRNA, we observed that increased levels of Gag promoted both translational arrest and stress granule assembly. Moreover, HIV-2 Gag also localizes to stress granules in the absence of a 'packageable' gRNA. Our results indicate that the HIV-2 gRNA is compartmentalized in stress granules in the absence of active translation prior to being selected for packaging by the Gag polyprotein.


Assuntos
Grânulos Citoplasmáticos/virologia , HIV-2/genética , RNA Viral/metabolismo , Montagem de Vírus , Grânulos Citoplasmáticos/metabolismo , Fator de Iniciação 2 em Eucariotos/metabolismo , Genoma Viral , HIV-2/fisiologia , Células HeLa , Humanos , Biossíntese de Proteínas , RNA Viral/análise , RNA Viral/biossíntese , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Ribonucleoproteínas/metabolismo , Estresse Fisiológico , Replicação Viral , Produtos do Gene gag do Vírus da Imunodeficiência Humana/biossíntese , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética
20.
Nucleic Acids Res ; 41(12): 6286-99, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23630313

RESUMO

Here, we show a novel molecular mechanism promoted by the DEAD-box RNA helicase DDX3 for translation of the HIV-1 genomic RNA. This occurs through the adenosine triphosphate-dependent formation of a translation initiation complex that is assembled at the 5' m(7)GTP cap of the HIV-1 mRNA. This is due to the property of DDX3 to substitute for the initiation factor eIF4E in the binding of the HIV-1 m(7)GTP 5' cap structure where it nucleates the formation of a core DDX3/PABP/eIF4G trimeric complex on the HIV-1 genomic RNA. By using RNA fluorescence in situ hybridization coupled to indirect immunofluorescence, we further show that this viral ribonucleoprotein complex is addressed to compartmentalized cytoplasmic foci where the translation initiation complex is assembled.


Assuntos
RNA Helicases DEAD-box/fisiologia , HIV-1/genética , Iniciação Traducional da Cadeia Peptídica , RNA Viral/biossíntese , Trifosfato de Adenosina/metabolismo , Compartimento Celular , Grânulos Citoplasmáticos/química , RNA Helicases DEAD-box/análise , Fator de Iniciação 4E em Eucariotos/metabolismo , Genoma Viral , HIV-1/fisiologia , Células HeLa , Humanos , Células Jurkat , Capuzes de RNA/metabolismo , RNA Viral/análise , Replicação Viral
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