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[PMID]:28821995
[Au] Autor:Bergallo M; Montanari P; Mareschi K; Merlino C; Berger M; Bini I; Daprà V; Galliano I; Fagioli F
[Ad] Endereço:Department of Public Health and Pediatric Sciences, University of Turin, Medical School, 10136, Turin, Italy. Massimiliano.bergallo@unito.it.
[Ti] Título:Expression of the pol gene of human endogenous retroviruses HERV-K and -W in leukemia patients.
[So] Source:Arch Virol;162(12):3639-3644, 2017 Dec.
[Is] ISSN:1432-8798
[Cp] País de publicação:Austria
[La] Idioma:eng
[Ab] Resumo:The human endogenous retroviruses (HERVs) are a family of endogenous retroviruses that integrated into the germ cell DNA of primates over 30 million years ago. HERV expression seems impaired in several diseases, ranging from autoimmune to neoplastic disorders. The purpose of this study was to evaluate the overall endogenous retroviral transcription profile in bone marrow (BM) samples. A total of 30 paediatric high-risk leukaemia patients (lymphoid and myeloid malignancies) were tested for HERVs virus gene expression. Our findings show that HERV-K expression was significantly higher in leukaemia patients when compared to healthy donors of a similar median age. We observed a significantly high expression of HERV-K in acute lymphoblastic leukemia (ALL) patients. In this study, we also found a relative overexpression of the endogenous retrovirus HERV-K in BM cells from the majority of leukemia samples analyzed, in particular in ALL. This overexpression might be related to lymphatic leukemogenesis and it warrants further investigations.
[Mh] Termos MeSH primário: Retrovirus Endógenos/enzimologia
Expressão Gênica
Produtos do Gene pol/análise
Leucemia/patologia
[Mh] Termos MeSH secundário: Adolescente
Medula Óssea/patologia
Criança
Pré-Escolar
Feminino
Perfilação da Expressão Gênica
Seres Humanos
Lactente
Masculino
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Gene Products, pol)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171113
[Lr] Data última revisão:
171113
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170820
[St] Status:MEDLINE
[do] DOI:10.1007/s00705-017-3526-7


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[PMID]:28700677
[Au] Autor:Saha B; Parks RJ
[Ad] Endereço:Regenerative Medicine Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada.
[Ti] Título:Human adenovirus type 5 vectors deleted of early region 1 (E1) undergo limited expression of early replicative E2 proteins and DNA replication in non-permissive cells.
[So] Source:PLoS One;12(7):e0181012, 2017.
[Is] ISSN:1932-6203
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Adenovirus (Ad) vectors deleted of the early region 1 (E1) are widely used for transgene delivery in preclinical and clinical gene therapy studies. Although proteins encoded within the E1 region are required for efficient virus replication, previous studies have suggested that certain viral or cellular proteins can functionally compensate for E1, leading to expression of the early region 2 (E2)-encoded replicative proteins and subsequent virus replication. We have generated a series of E1-encoding and E1-deficient Ad vectors containing a FLAG-epitope tag on each of the E2-encoded proteins: DNA-binding protein (DBP), terminal protein (TP) and DNA polymerase (Pol). Using these constructs, we show that for the replication-competent virus, the expression level of each E2-encoded protein declines with increasing distance from the E2 promoter, with E2A-encoded DBP expression being ~800-fold higher than E2B-encoded TP. Pol was expressed at extremely low levels in infected cells, and immunoprecipitation from cell lysates was required prior to its detection by immunoblot. We further show that DBP was expressed 200- to 400-fold less efficiently from an E1-deficient virus compared to a replication-competent virus in A549 and HepG2 cells, which was accompanied by a very small increase in genome copy number. For the E1-deficient virus, late gene expression (a marker of virus replication) was only observed at very high multiplicities of infection. These data show that E1-deleted Ad gives rise to limited expression of the E2-encoded genes and replication in infected cells, but highlight the importance of considering viral dose-dependent effects in gene therapy studies.
[Mh] Termos MeSH primário: Adenovírus Humanos/genética
Replicação do DNA/fisiologia
Vetores Genéticos/genética
[Mh] Termos MeSH secundário: Adenovírus Humanos/fisiologia
Linhagem Celular
Replicação do DNA/genética
Proteínas de Ligação a DNA/genética
Proteínas de Ligação a DNA/metabolismo
Produtos do Gene pol/genética
Produtos do Gene pol/metabolismo
Células Hep G2
Seres Humanos
Proteínas Virais/genética
Proteínas Virais/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (DNA-Binding Proteins); 0 (Gene Products, pol); 0 (Viral Proteins); 0 (terminal protein, adenovirus)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170927
[Lr] Data última revisão:
170927
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170713
[St] Status:MEDLINE
[do] DOI:10.1371/journal.pone.0181012


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[PMID]:28314126
[Au] Autor:Spannaus R; Miller C; Lindemann D; Bodem J
[Ad] Endereço:Institut für Virologie und Immunbiologie, Julius-Maximilians-Universität Würzburg, Germany.
[Ti] Título:Purification of foamy viral particles.
[So] Source:Virology;506:28-33, 2017 Jun.
[Is] ISSN:1096-0341
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Foamy viruses are non-pathogenic retroviruses and represent a tool for vector development. For gene therapy applications and for analyses of viral protein composition infectious particles need to be purified, which has been difficult for foamy viruses in the past. Here, we describe a novel, simple, and fast purification method for prototype foamy viruses with high purity using size exclusion and affinity chromatography. More than 99,9% of the contaminating proteins were removed. The purified viruses were used to determine the amount of the incorporated Pol protein relative to Gag. The determined Gag to Pol PR-RT ratio of 30:1 confirmed previous studies suggesting FV virions encapsidate fewer number of Pol molecules than orthoretroviruses.
[Mh] Termos MeSH primário: Cromatografia de Afinidade/métodos
Cromatografia em Gel/métodos
Spumavirus/crescimento & desenvolvimento
Vírion/crescimento & desenvolvimento
[Mh] Termos MeSH secundário: Regulação Viral da Expressão Gênica
Produtos do Gene pol/genética
Produtos do Gene pol/metabolismo
Seres Humanos
Infecções por Retroviridae/virologia
Spumavirus/genética
Spumavirus/isolamento & purificação
Spumavirus/fisiologia
Vírion/genética
Vírion/isolamento & purificação
Vírion/fisiologia
Montagem de Vírus
Cultura de Vírus
[Pt] Tipo de publicação:EVALUATION STUDIES; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Gene Products, pol)
[Em] Mês de entrada:1707
[Cu] Atualização por classe:170705
[Lr] Data última revisão:
170705
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170318
[St] Status:MEDLINE


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[PMID]:27852858
[Au] Autor:Clark DN; Flanagan JM; Hu J
[Ad] Endereço:Department of Microbiology and Immunology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania, USA dnc142@psu.edu.
[Ti] Título:Mapping of Functional Subdomains in the Terminal Protein Domain of Hepatitis B Virus Polymerase.
[So] Source:J Virol;91(3), 2017 Feb 01.
[Is] ISSN:1098-5514
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Hepatitis B virus (HBV) encodes a multifunction reverse transcriptase or polymerase (P), which is composed of several domains. The terminal protein (TP) domain is unique to HBV and related hepadnaviruses and is required for specifically binding to the viral pregenomic RNA (pgRNA). Subsequently, the TP domain is necessary for pgRNA packaging into viral nucleocapsids and the initiation of viral reverse transcription for conversion of the pgRNA to viral DNA. Uniquely, the HBV P protein initiates reverse transcription via a protein priming mechanism using the TP domain as a primer. No structural homologs or high-resolution structure exists for the TP domain. Secondary structure prediction identified three disordered loops in TP with highly conserved sequences. A meta-analysis of mutagenesis studies indicated these predicted loops are almost exclusively where functionally important residues are located. Newly constructed TP mutations revealed a priming loop in TP which plays a specific role in protein-primed DNA synthesis beyond simply harboring the site of priming. Substitutions of potential sites of phosphorylation surrounding the priming site demonstrated that these residues are involved in interactions critical for priming but are unlikely to be phosphorylated during viral replication. Furthermore, the first 13 and 66 TP residues were shown to be dispensable for protein priming and pgRNA binding, respectively. Combining current and previous mutagenesis work with sequence analysis has increased our understanding of TP structure and functions by mapping specific functions to distinct predicted secondary structures and will facilitate antiviral targeting of this unique domain. IMPORTANCE: HBV is a major cause of viral hepatitis, liver cirrhosis, and hepatocellular carcinoma. One important feature of this virus is its polymerase, the enzyme used to create the DNA genome from a specific viral RNA by reverse transcription. One region of this polymerase, the TP domain, is required for association with the viral RNA and production of the DNA genome. Targeting the TP domain for antiviral development is difficult due to the lack of homology to other proteins and high-resolution structure. This study mapped the TP functions according to predicted secondary structure, where it folds into alpha helices or unstructured loops. Three predicted loops were found to be the most important regions functionally and the most conserved evolutionarily. Identification of these functional subdomains in TP will facilitate its targeting for antiviral development.
[Mh] Termos MeSH primário: Produtos do Gene pol/genética
Produtos do Gene pol/metabolismo
Vírus da Hepatite B/genética
Vírus da Hepatite B/metabolismo
Domínios e Motivos de Interação entre Proteínas
[Mh] Termos MeSH secundário: Sequência de Aminoácidos
Sequência Conservada
Produtos do Gene pol/química
Seres Humanos
Modelos Moleculares
Mutação
Fenótipo
Conformação Proteica em alfa-Hélice
RNA Mensageiro/genética
RNA Viral
Proteínas de Ligação a RNA
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Gene Products, pol); 0 (P protein, Hepatitis B virus); 0 (RNA, Messenger); 0 (RNA, Viral); 0 (RNA-Binding Proteins)
[Em] Mês de entrada:1705
[Cu] Atualização por classe:170718
[Lr] Data última revisão:
170718
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161118
[St] Status:MEDLINE


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[PMID]:27923946
[Au] Autor:Steegen K; Bronze M; Papathanasopoulos MA; van Zyl G; Goedhals D; Van Vuuren C; Macleod W; Sanne I; Stevens WS; Carmona SC
[Ad] Endereço:Department of Molecular Medicine and Haematology.
[Ti] Título:Prevalence of Antiretroviral Drug Resistance in Patients Who Are Not Responding to Protease Inhibitor-Based Treatment: Results From the First National Survey in South Africa.
[So] Source:J Infect Dis;214(12):1826-1830, 2016 Dec 15.
[Is] ISSN:1537-6613
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Limited data exist on human immunodeficiency virus type 1 (HIV-1) resistance in patients who are not responding to protease inhibitor (PI)-based regimens in resource-limited settings. This study assessed resistance profiles in adults across South Africa who were not responding to PI-based regimens. pol sequencing was undertaken and submitted to the Stanford HIV Drug Resistance Database. At least 1 major PI mutation was detected in 16.4% of 350 participants. A total of 53.4% showed intermediate resistance to darunavir/ritonavir, whereas high-level resistance was not observed. Only 5.2% and 32.8% of participants showed high-level and intermediate resistance to etravirine, respectively. Although the prevalence of major PI mutations was within previously reported ranges, most patients will likely experience virological suppression during receipt of currently available South African third-line regimens.
[Mh] Termos MeSH primário: Antirretrovirais/uso terapêutico
Farmacorresistência Viral
Infecções por HIV/tratamento farmacológico
Infecções por HIV/virologia
Inibidores da Protease de HIV/uso terapêutico
HIV-1/efeitos dos fármacos
[Mh] Termos MeSH secundário: Adolescente
Adulto
Idoso
Antirretrovirais/farmacologia
Estudos Transversais
Produtos do Gene pol/genética
Inibidores da Protease de HIV/farmacologia
HIV-1/isolamento & purificação
Seres Humanos
Meia-Idade
Mutação de Sentido Incorreto
Prevalência
Análise de Sequência de DNA
África do Sul/epidemiologia
Falha de Tratamento
Adulto Jovem
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Anti-Retroviral Agents); 0 (Gene Products, pol); 0 (HIV Protease Inhibitors)
[Em] Mês de entrada:1705
[Cu] Atualização por classe:170522
[Lr] Data última revisão:
170522
[Sb] Subgrupo de revista:AIM; IM
[Da] Data de entrada para processamento:161208
[St] Status:MEDLINE


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[PMID]:27492206
[Au] Autor:Wasityastuti W; Yano Y; Widasari DI; Yamani LN; Ratnasari N; Heriyanto DS; Okada R; Tanahashi T; Murakami Y; Azuma T; Hayashi Y
[Ad] Endereço:Division of Infectious Disease Pathology, Department of Microbiology and Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Japan.
[Ti] Título:Different Variants in Reverse Transcriptase Domain Determined by Ultra-deep Sequencing in Treatment-naïve and Treated Indonesian Patients Infected with Hepatitis B Virus.
[So] Source:Kobe J Med Sci;62(1):E1-8, 2016 Jun 16.
[Is] ISSN:1883-0498
[Cp] País de publicação:Japan
[La] Idioma:eng
[Ab] Resumo:A nucleos(t)ide analog (NA) is the common antiviral drug available for directly treating hepatitis B virus (HBV) infection. However, its application has led to the emergence of NA-resistant mutations mostly in a conserved region of the reverse transcriptase domain of HBV polymerase. Harboring NA-resistant mutations decreases drug effectiveness and increases the frequency of end-stage liver disease. The invention of next-generation sequencing that can generate thousands of sequences from viral complex mixtures provides opportunities to detect minor changes and early viral evolution under drug stress. The present study used ultra-deep sequencing to evaluate discrepant quasispecies in the reverse transcriptase domain of HBV including NA-resistant hotspots between seven treatment-naïve Indonesian patients infected with HBV and five at the early phase of treatment. The most common sub-genotype was HBV B3 (83.34%). The substitution rate of variants determined among amino acids with a ratio of ≥ 1% changes was higher among the population in conserved regions (23.19% vs. 4.59%, P = 0.001) and in the inter-reverse transcriptase domain (23.95% vs. 2.94%, P = 0.002) in treatment naïve, than in treated patients. Nine hotspots of antiviral resistance were identified in both groups, and the mean frequency of changes in all patients was < 1%. The known rtM204I mutation was the most frequent in both groups. The lower rate of variants in HBV quasispecies in patients undergoing treatment could be associated with virus elimination and the extinction of sensitive species by NA therapy. The present findings imply that HBV quasispecies dynamically change during treatment.
[Mh] Termos MeSH primário: Produtos do Gene pol/genética
Vírus da Hepatite B/enzimologia
Vírus da Hepatite B/genética
DNA Polimerase Dirigida por RNA/genética
[Mh] Termos MeSH secundário: Adolescente
Adulto
Idoso
Sequência de Aminoácidos
Substituição de Aminoácidos
Antivirais/farmacologia
Farmacorresistência Viral/genética
Feminino
Produtos do Gene pol/química
Variação Genética
Vírus da Hepatite B/efeitos dos fármacos
Hepatite B Crônica/tratamento farmacológico
Hepatite B Crônica/virologia
Sequenciamento de Nucleotídeos em Larga Escala
Seres Humanos
Indonésia
Masculino
Meia-Idade
Domínios Proteicos
DNA Polimerase Dirigida por RNA/química
Análise de Sequência de DNA
Adulto Jovem
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Antiviral Agents); 0 (Gene Products, pol); 0 (P protein, Hepatitis B virus); EC 2.7.7.49 (RNA-Directed DNA Polymerase)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170608
[Lr] Data última revisão:
170608
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160806
[St] Status:MEDLINE


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[PMID]:27399760
[Au] Autor:Sabino Cunha M; Lima Sampaio T; Peterlin BM; Jesus da Costa L
[Ad] Endereço:Departamento de Virologia-Instituto de Microbiologia, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373-CCS-Bloco I, Rio de Janeiro 21941-902, Brazil. marcela.scw@gmail.com.
[Ti] Título:A Truncated Nef Peptide from SIVcpz Inhibits the Production of HIV-1 Infectious Progeny.
[So] Source:Viruses;8(7), 2016 Jul 07.
[Is] ISSN:1999-4915
[Cp] País de publicação:Switzerland
[La] Idioma:eng
[Ab] Resumo:Nef proteins from all primate Lentiviruses, including the simian immunodeficiency virus of chimpanzees (SIVcpz), increase viral progeny infectivity. However, the function of Nef involved with the increase in viral infectivity is still not completely understood. Nonetheless, until now, studies investigating the functions of Nef from SIVcpz have been conducted in the context of the HIV-1 proviruses. In an attempt to investigate the role played by Nef during the replication cycle of an SIVcpz, a Nef-defective derivative was obtained from the SIVcpzWTGab2 clone by introducing a frame shift mutation at a unique restriction site within the nef sequence. This nef-deleted clone expresses an N-terminal 74-amino acid truncated peptide of Nef and was named SIVcpz-tNef. We found that the SIVcpz-tNef does not behave as a classic nef-deleted HIV-1 or simian immunodeficiency virus of macaques SIVmac. Markedly, SIVcpz-tNef progeny from both Hek-293T and Molt producer cells were completely non-infectious. Moreover, the loss in infectivity of SIVcpz-tNef correlated with the inhibition of Gag and GagPol processing. A marked accumulation of Gag and very low levels of reverse transcriptase were detected in viral lysates. Furthermore, these observations were reproduced once the tNef peptide was expressed in trans both in SIVcpzΔNef and HIV-1WT expressing cells, demonstrating that the truncated peptide is a dominant negative for viral processing and infectivity for both SIVcpz and HIV-1. We demonstrated that the truncated Nef peptide binds to GagPol outside the protease region and by doing so probably blocks processing of both GagPol and Gag precursors at a very early stage. This study demonstrates for the first time that naturally-occurring Nef peptides can potently block lentiviral processing and infectivity.
[Mh] Termos MeSH primário: Produtos do Gene nef/metabolismo
HIV-1/fisiologia
Vírus da Imunodeficiência Símia/fisiologia
Replicação Viral
[Mh] Termos MeSH secundário: Animais
Linhagem Celular
Mutação da Fase de Leitura
Técnicas de Inativação de Genes
Produtos do Gene gag/metabolismo
Produtos do Gene nef/genética
Produtos do Gene pol/metabolismo
Seres Humanos
Pan troglodytes
Ligação Proteica
Vírus da Imunodeficiência Símia/genética
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Gene Products, gag); 0 (Gene Products, nef); 0 (Gene Products, pol)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170926
[Lr] Data última revisão:
170926
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160712
[St] Status:MEDLINE


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[PMID]:27211560
[Au] Autor:Faucard R; Madeira A; Gehin N; Authier FJ; Panaite PA; Lesage C; Burgelin I; Bertel M; Bernard C; Curtin F; Lang AB; Steck AJ; Perron H; Kuntzer T; Créange A
[Ad] Endereço:GeNeuro Innovation, France.
[Ti] Título:Human Endogenous Retrovirus and Neuroinflammation in Chronic Inflammatory Demyelinating Polyradiculoneuropathy.
[So] Source:EBioMedicine;6:190-198, 2016 Apr.
[Is] ISSN:2352-3964
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: Human endogenous retroviruses HERV-W encode a pro-inflammatory protein, named MSRV-Env from its original identification in Multiple Sclerosis. Though not detected in various neurological controls, MSRV-Env was found in patients with chronic inflammatory demyelinating polyradiculoneuropathies (CIDPs). This study investigated the expression of MSRV in CIDP and evaluated relevant MSRV-Env pathogenic effects. METHODS: 50 CIDP patients, 19 other neurological controls (ONDs) and 65 healthy blood donors (HBDs) were recruited from two different countries. MSRV-env and -pol transcripts, IL6 and CXCL10 levels were quantified from blood samples. MSRV-Env immunohistology was performed in distal sensory nerves from CIDP and neurological controls biopsies. MSRV-Env pathogenic effects and mode of action were assayed in cultured primary human Schwann cells (HSCs). FINDINGS: In both cohorts, MSRV-env and -pol transcripts, IL6 positivity prevalence and CXCL10 levels were significantly elevated in CIDP patients when compared to HBDs and ONDs (statistically significant in all comparisons). MSRV-Env protein was detected in Schwann cells in 5/7 CIDP biopsies. HSC exposed to or transfected with MSRV-env presented a strong increase of IL6 and CXCL10 transcripts and protein secretion. These pathogenic effects on HSC were inhibited by GNbAC1, a highly specific and neutralizing humanized monoclonal antibody targeting MSRV-Env. INTERPRETATION: The present study showed that MSRV-Env may trigger the release of critical immune mediators proposed as instrumental factors involved in the pathophysiology of CIDP. Significant MSRV-Env expression was detected in a significant proportion of patients with CIDP, in which it may play a role according to its presently observed effects on Schwann cells along with previously known effects on immune cells. Experimental results also suggest that a biomarker-driven therapeutic strategy targeting this protein with a neutralizing antibody such as GNbAC1 may offer new perspectives for treating CIDP patients with positive detection of MSRV-Env expression. FUNDING: Geneuro-Innovation, France.
[Mh] Termos MeSH primário: Quimiocina CXCL10/genética
Retrovirus Endógenos/patogenicidade
Produtos do Gene env/genética
Interleucina-6/genética
Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/imunologia
[Mh] Termos MeSH secundário: Adulto
Idoso
Anticorpos Monoclonais Humanizados/farmacologia
Linhagem Celular
Retrovirus Endógenos/genética
Retrovirus Endógenos/imunologia
Feminino
França
Produtos do Gene pol/genética
Seres Humanos
Masculino
Meia-Idade
Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/genética
Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/virologia
Células de Schwann/efeitos dos fármacos
Células de Schwann/virologia
Adulto Jovem
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Antibodies, Monoclonal, Humanized); 0 (CXCL10 protein, human); 0 (Chemokine CXCL10); 0 (GNbAC1 monoclonal antibody); 0 (Gene Products, env); 0 (Gene Products, pol); 0 (IL6 protein, human); 0 (Interleukin-6)
[Em] Mês de entrada:1703
[Cu] Atualização por classe:170625
[Lr] Data última revisão:
170625
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160524
[St] Status:MEDLINE


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[PMID]:27122388
[Au] Autor:Luan H; Wang Y; Li Y; Cui Z; Chang S; Zhao P
[Ad] Endereço:College of Veterinary Medicine, Shandong Agricultural University, Tai'an, 271018.
[Ti] Título:Development of a real-time quantitative RT-PCR to detect REV contamination in live vaccine.
[So] Source:Poult Sci;95(9):2023-9, 2016 Sep 01.
[Is] ISSN:1525-3171
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Based on the published Avian reticuloendotheliosis virus (REV) whole genome sequence, primers and TaqMan probes were designed and synthesized, and the TaqMan probe fluorescence real-time quantitative RT-PCR (qRT-PCR) method for detecting the REV pol gene was established by optimizing the reaction conditions. Sensitivity analysis showed that the qRT-PCR method had a sensitivity that was 1,000-fold higher than conventional PCR. Additionally, no amplification signals were obtained when we attempted to detect DNA or cDNA of ALV-A/B/J, MDV, CIAV, IBDV, ARV, NDV, AIV, or other viruses, suggesting a high specificity for our method. Various titers of REV were artificially "spiked" into the FPV and MDV vaccines to simulate REV contamination in attenuated vaccines to validate this qRT-PCR method. Our findings indicated that this qRT-PCR method could detect REV contamination at a dose of 1 TCID50/1,000 feathers, which was 10,000-fold more sensitive than the regular RT-PCR detection (10(4) TCID50/1000 feathers).
[Mh] Termos MeSH primário: Produtos do Gene pol/isolamento & purificação
Doenças das Aves Domésticas/prevenção & controle
Reação em Cadeia da Polimerase em Tempo Real/veterinária
Vírus da Reticuloendoteliose/imunologia
Infecções por Retroviridae/veterinária
Infecções Tumorais por Vírus/veterinária
Vacinas Virais/análise
[Mh] Termos MeSH secundário: Animais
Doenças das Aves Domésticas/virologia
Vírus da Reticuloendoteliose/isolamento & purificação
Infecções por Retroviridae/prevenção & controle
Infecções por Retroviridae/virologia
Infecções Tumorais por Vírus/prevenção & controle
Infecções Tumorais por Vírus/virologia
Vacinas Atenuadas/análise
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Gene Products, pol); 0 (Vaccines, Attenuated); 0 (Viral Vaccines)
[Em] Mês de entrada:1704
[Cu] Atualização por classe:170411
[Lr] Data última revisão:
170411
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160429
[St] Status:MEDLINE
[do] DOI:10.3382/ps/pew147


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[PMID]:26939125
[Au] Autor:Rowe CL; Wagstaff KM; Oksayan S; Glover DJ; Jans DA; Moseley GW
[Ad] Endereço:Viral Pathogenesis Laboratory, Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Parkville, Victoria, Australia.
[Ti] Título:Nuclear Trafficking of the Rabies Virus Interferon Antagonist P-Protein Is Regulated by an Importin-Binding Nuclear Localization Sequence in the C-Terminal Domain.
[So] Source:PLoS One;11(3):e0150477, 2016.
[Is] ISSN:1932-6203
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Rabies virus P-protein is expressed as five isoforms (P1-P5) which undergo nucleocytoplasmic trafficking important to roles in immune evasion. Although nuclear import of P3 is known to be mediated by an importin (IMP)-recognised nuclear localization sequence in the N-terminal region (N-NLS), the mechanisms underlying nuclear import of other P isoforms in which the N-NLS is inactive or has been deleted have remained unresolved. Based on the previous observation that mutation of basic residues K214/R260 of the P-protein C-terminal domain (P-CTD) can result in nuclear exclusion of P3, we used live cell imaging, protein interaction analysis and in vitro nuclear transport assays to examine in detail the nuclear trafficking properties of this domain. We find that the effect of mutation of K214/R260 on P3 is largely dependent on nuclear export, suggesting that nuclear exclusion of mutated P3 involves the P-CTD-localized nuclear export sequence (C-NES). However, assays using cells in which nuclear export is pharmacologically inhibited indicate that these mutations significantly inhibit P3 nuclear accumulation and, importantly, prevent nuclear accumulation of P1, suggestive of effects on NLS-mediated import activity in these isoforms. Consistent with this, molecular binding and transport assays indicate that the P-CTD mediates IMPα2/IMPß1-dependent nuclear import by conferring direct binding to the IMPα2/IMPß1 heterodimer, as well as to a truncated form of IMPα2 lacking the IMPß-binding autoinhibitory domain (ΔIBB-IMPα2), and IMPß1 alone. These properties are all dependent on K214 and R260. This provides the first evidence that P-CTD contains a genuine IMP-binding NLS, and establishes the mechanism by which P-protein isoforms other than P3 can be imported to the nucleus. These data underpin a refined model for P-protein trafficking that involves the concerted action of multiple NESs and IMP-binding NLSs, and highlight the intricate regulation of P-protein subcellular localization, consistent with important roles in infection.
[Mh] Termos MeSH primário: Produtos do Gene pol/genética
Genômica
Carioferinas/metabolismo
Mapas de Interação de Proteínas/genética
Vírus da Raiva/metabolismo
[Mh] Termos MeSH secundário: Transporte Ativo do Núcleo Celular/genética
Citoplasma/metabolismo
Produtos do Gene pol/metabolismo
Interferons/antagonistas & inibidores
Interferons/genética
Sinais de Localização Nuclear/metabolismo
Imagem Óptica
Ligação Proteica
Estrutura Terciária de Proteína
Vírus da Raiva/patogenicidade
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Gene Products, pol); 0 (Karyopherins); 0 (Nuclear Localization Signals); 9008-11-1 (Interferons)
[Em] Mês de entrada:1608
[Cu] Atualização por classe:170220
[Lr] Data última revisão:
170220
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160304
[St] Status:MEDLINE
[do] DOI:10.1371/journal.pone.0150477



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