Base de dados : MEDLINE
Pesquisa : E05.196.401.143.500 [Categoria DeCS]
Referências encontradas : 165 [refinar]
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  1 / 165 MEDLINE  
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[PMID]:28470609
[Au] Autor:Strutton B; Jaffé SRP; Pandhal J; Wright PC
[Ad] Endereço:Department of Chemical and Biological Engineering, ChELSI Institute, University of Sheffield, Mappin Street, Sheffield, S1 3JD, UK.
[Ti] Título:Generation of Recombinant N-Linked Glycoproteins in E. coli.
[So] Source:Methods Mol Biol;1586:233-250, 2017.
[Is] ISSN:1940-6029
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The production of N-linked recombinant glycoproteins is possible in a variety of biotechnology host cells, and more recently in the bacterial workhorse, Escherichia coli. This methods chapter will outline the components and procedures needed to produce N-linked glycoproteins in E. coli, utilizing Campylobacter jejuni glycosylation machinery, although other related genes can be used with minimal tweaks to this methodology. To ensure a successful outcome, various methods will be highlighted that can confirm glycoprotein production to a high degree of confidence, including the gold standard of mass spectrometry analysis.
[Mh] Termos MeSH primário: Campylobacter jejuni/genética
Escherichia coli/genética
Glicoproteínas/genética
Interferon-alfa/genética
[Mh] Termos MeSH secundário: Far-Western Blotting/métodos
Clonagem Molecular/métodos
Eletroforese em Gel de Poliacrilamida/métodos
Genes Bacterianos
Glicoproteínas/química
Glicoproteínas/isolamento & purificação
Glicosilação
Interferon-alfa/química
Interferon-alfa/isolamento & purificação
Espectrometria de Massas/métodos
Plasmídeos/genética
Polissacarídeos/análise
Polissacarídeos/genética
Processamento de Proteína Pós-Traducional
Proteínas Recombinantes/química
Proteínas Recombinantes/genética
Proteínas Recombinantes/isolamento & purificação
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Glycoproteins); 0 (Interferon-alpha); 0 (Polysaccharides); 0 (Recombinant Proteins); 43K1W2T1M6 (interferon alfa-2b)
[Em] Mês de entrada:1802
[Cu] Atualização por classe:180220
[Lr] Data última revisão:
180220
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170505
[St] Status:MEDLINE
[do] DOI:10.1007/978-1-4939-6887-9_15


  2 / 165 MEDLINE  
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[PMID]:28631595
[Au] Autor:Zhong Y; Fei C; Tang X; Zhan W; Sheng X
[Ad] Endereço:1​Laboratory of Pathology and Immunology of Aquatic Animals, KLM, Ocean University of China, 5 Yushan Road, Qingdao 266003, PR China.
[Ti] Título:A 32 kDa viral attachment protein of lymphocystis disease virus (LCDV) specifically interacts with a 27.8 kDa cellular receptor from flounder (Paralichthys olivaceus).
[So] Source:J Gen Virol;98(6):1477-1488, 2017 Jun.
[Is] ISSN:1465-2099
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:The 27.8 kDa protein in flounder gill (FG) cells was previously proved to be a receptor specific for lymphocystis disease virus (LCDV) entry and infection. In this paper, a 32 kDa viral attachment protein (VAP) of LCDV specifically binding to the 27.8 kDa receptor (27.8R) was found by far-Western blotting coupled with monoclonal antibodies (MAbs) against 27.8R. The 32 kDa protein was confirmed to be encoded by the open reading frame (ORF) 038 gene in LCDV-C, and predicted to contain a putative transmembrane region, multiple N-myristoylation and glycosylation sites and phosphorylation motifs. The expression plasmid of pET-32a-ORF038 was constructed and the recombinant VAP (rVAP) was obtained. Rabbit polyclonal antibodies against the rVAP were prepared and could recognize the rVAP and 32 kDa protein in LCDV. Immunogold electron microscopy showed that the 32 kDa protein was located on the surface of LCDV particles. Immunofluorescence assay demonstrated that the rVAP could bind to the 27.8R on the cell membrane of the FG monolayer and the anti-27.8R MAbs could block the rVAP binding. Pre-incubation of the rVAP with FG cells before LCDV infection, or pre-incubation of LCDV with the antibodies against the rVAP, could significantly decrease the LCDV copy numbers (P<0.05) and delay the emergence of cytopathic effects in FG cells in a dose-dependent manner. These results indicated for the first time that the 32 kDa protein functioned as an attachment protein for the initial attachment and entry of LCDV, and the interaction of the 32 kDa VAP with the 27.8R-initiated LCDV infection.
[Mh] Termos MeSH primário: Linguado/virologia
Interações Hospedeiro-Patógeno
Iridoviridae/fisiologia
Receptores de Superfície Celular/metabolismo
Proteínas do Envelope Viral/metabolismo
Ligação Viral
[Mh] Termos MeSH secundário: Animais
Far-Western Blotting
Microscopia Imunoeletrônica
Ligação Proteica
Receptores de Superfície Celular/genética
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Receptors, Cell Surface); 0 (Viral Envelope Proteins)
[Em] Mês de entrada:1707
[Cu] Atualização por classe:170717
[Lr] Data última revisão:
170717
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170621
[St] Status:MEDLINE
[do] DOI:10.1099/jgv.0.000805


  3 / 165 MEDLINE  
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[PMID]:27454323
[Au] Autor:Lee YJ; Chen LL
[Ad] Endereço:Doctoral Degree Program in Marine Biotechnology, National Taiwan Ocean University, Keelung, Taiwan, ROC.
[Ti] Título:WSSV envelope protein VP51B links structural protein complexes and may mediate virus infection.
[So] Source:J Fish Dis;40(4):571-581, 2017 Apr.
[Is] ISSN:1365-2761
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:White spot syndrome virus (WSSV), an enveloped double-stranded DNA virus, is the causative agent of a disease that has led to severe mortalities of cultured shrimps in Taiwan and many other countries. In the previous study, Penaeus monodon chitin-binding protein (CBP) and glucose transporter 1 (Glut1), two cell membrane proteins, were found to at least interact with other 10 WSSV envelope proteins including VP51B. These envelope proteins might form a protein complex. According to the known information, VP51B was used to identify its role in the protein complex. Western blotting of the intact viral particles and fractionation of the viral components confirmed that VP51B is one of WSSV envelope proteins. In this study, the protein-protein interaction between VP51B and other WSSV envelope proteins was identified by far-western blot experiment and VP51B was found to interact with VP24, VP31, VP32, VP39B and VP41A. Furthermore, the in vivo neutralization experiment using recombinant VP51B plus with VP39B showed the best inhibition. These data indicate that VP51B participates in the WSSV protein complex and plays an important role in WSSV infection.
[Mh] Termos MeSH primário: Penaeidae/virologia
Proteínas do Envelope Viral/genética
Vírus 1 da Síndrome da Mancha Branca/fisiologia
[Mh] Termos MeSH secundário: Animais
Far-Western Blotting
Perfilação da Expressão Gênica
Ligação Proteica
Proteínas do Envelope Viral/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Viral Envelope Proteins)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170621
[Lr] Data última revisão:
170621
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160726
[St] Status:MEDLINE
[do] DOI:10.1111/jfd.12538


  4 / 165 MEDLINE  
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[PMID]:27393691
[Au] Autor:Toni M; Cioni C; De Angelis F; di Patti MC
[Ad] Endereço:Department of Biology and Biotechnology "Charles Darwin", Sapienza University, 00161, Rome, Italy. mattia.toni@uniroma1.it.
[Ti] Título:Synuclein expression in the lizard Anolis carolinensis.
[So] Source:J Comp Physiol A Neuroethol Sens Neural Behav Physiol;202(8):577-95, 2016 Aug.
[Is] ISSN:1432-1351
[Cp] País de publicação:Germany
[La] Idioma:eng
[Ab] Resumo:The synuclein (syn) family comprises three proteins: α-, ß- and γ-syns. In humans, they are involved in neurodegenerative diseases such as Parkinson's disease and in tumors. Members of the syn family were sequenced in representative species of all vertebrates and the comparative analysis of amino acid sequences suggests that syns are evolutionarily conserved, but information about their expression in vertebrate lineages is still scarce and completely lacking in reptiles. In this study, the expression of genes coding for α-, ß- and γ-syns was analyzed in the green lizard Anolis carolinensis by semiquantitative RT-PCR and Western blot. Results demonstrate good expression levels of the three syns in the lizard nervous system, similarly to human syns. This, together with the high identity between lizard and human syns, suggests that these proteins fulfill evolutionarily conserved functions. However, differences between lizard and humans in the expression of syn variants (two different variants of γ-syn were detected in A. carolinensis) and differences in some amino acids in key positions for the regulation of protein conformation and affinity for lipid and metal ions also suggest that these proteins may have acquired different functional specializations in the two lineages.
[Mh] Termos MeSH primário: Lagartos/metabolismo
Sinucleínas/metabolismo
[Mh] Termos MeSH secundário: Sequência de Aminoácidos
Animais
Sequência de Bases
Far-Western Blotting
Encéfalo/metabolismo
Evolução Molecular
Olho/metabolismo
Expressão Gênica
Seres Humanos
Intestinos/metabolismo
Fígado/metabolismo
Pulmão/metabolismo
Músculos/metabolismo
Miocárdio/metabolismo
Isoformas de Proteínas
Estrutura Secundária de Proteína
RNA Mensageiro/metabolismo
Reação em Cadeia da Polimerase em Tempo Real
Homologia de Sequência de Aminoácidos
Medula Espinal/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Protein Isoforms); 0 (RNA, Messenger); 0 (Synucleins)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:171014
[Lr] Data última revisão:
171014
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160710
[St] Status:MEDLINE
[do] DOI:10.1007/s00359-016-1108-x


  5 / 165 MEDLINE  
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[PMID]:27193714
[Au] Autor:Portseva TN; Brechalov AV; Dukhanina EA; Stepchenko AG; Pankratova EV; Georgieva SG
[Ad] Endereço:Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, ul. Vavilova 32, Moscow, 119991, Russia. tanjnij86@mail.ru.
[Ti] Título:Transcription factor Oct-1 stimulates the release of Mts1/S100A4 protein by the cancer cells.
[So] Source:Dokl Biochem Biophys;467(1):121-3, 2016 Mar.
[Is] ISSN:1608-3091
[Cp] País de publicação:Russia (Federation)
[La] Idioma:eng
[Ab] Resumo:The effect of the transcription factor Oct-1 (POU2F1) on the expression of the tumor cell marker metastasin (Mts1/S100A4) was studied. Comparative analysis of various tumor lines showed no clear correlation between the expression level of Mts1/S100A4 and the content of Oct-1. However, at stable transfection of tumor cells with Oct-1A, Oct-1L, and Oct-1X isoforms we detected an elevated level of Oct-1, which stimulated Mts1/S100A4 secretion. These findings extend our understanding of the molecular mechanisms of the tumorigenic effect of Oct-1.
[Mh] Termos MeSH primário: Neoplasias/metabolismo
Fator 1 de Transcrição de Octâmero/metabolismo
Proteína A4 de Ligação a Cálcio da Família S100/metabolismo
[Mh] Termos MeSH secundário: Far-Western Blotting
Linhagem Celular Tumoral
Meios de Cultura
Feminino
Seres Humanos
Isoformas de Proteínas
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Culture Media); 0 (Octamer Transcription Factor-1); 0 (POU2F1 protein, human); 0 (Protein Isoforms); 0 (S100 Calcium-Binding Protein A4); 142662-27-9 (S100A4 protein, human)
[Em] Mês de entrada:1701
[Cu] Atualização por classe:171019
[Lr] Data última revisão:
171019
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160520
[St] Status:MEDLINE
[do] DOI:10.1134/S1607672916020125


  6 / 165 MEDLINE  
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[PMID]:27071784
[Au] Autor:Sarro-Ramírez A; Sánchez D; Tejeda-Padrón A; Buenfil-Canto LV; Valladares-García J; Pacheco-Pantoja E; Arias-Carrión O; Murillo-Rodríguez E
[Ti] Título:Characterization of Mitogen-Activated Protein Kinase Expression in Nucleus Accumbens and Hippocampus of Rats Subjected to Food Selection in the Cafeteria Diet Protocol.
[So] Source:CNS Neurol Disord Drug Targets;15(7):866-72, 2016.
[Is] ISSN:1996-3181
[Cp] País de publicação:United Arab Emirates
[La] Idioma:eng
[Ab] Resumo:Obesity is a world-wide health problem that requires different experimental perspectives to understand the onset of this disease, including the neurobiological basis of food selection. From a molecular perspective, obesity has been related with activity of several endogenous molecules, including the mitogenactivated protein kinases (MAP-K). The aim of this study was to characterize MAP-K expression in hedonic and learning and memory brain-associated areas such as nucleus accumbens (AcbC) and hippocampus (HIPP) after food selection. We show that animals fed with cafeteria diet during 14 days displayed an increase in p38 MAP-K activity in AcbC if chose cheese. Conversely, a diminution was observed in animals that preferred chocolate in AcbC. Also, a decrease of p38 MAP-K phosphorylation was found in HIPP in rats that selected either cheese or chocolate. Our data demonstrate a putative role of MAP-K expression in food selection. These findings advance our understanding of neuromolecular basis engaged in obesity.
[Mh] Termos MeSH primário: Preferências Alimentares/fisiologia
Hipocampo/enzimologia
Núcleo Accumbens/enzimologia
Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
[Mh] Termos MeSH secundário: Animais
Far-Western Blotting
Comportamento de Escolha
Masculino
Modelos Animais
Ratos Wistar
Reação em Cadeia da Polimerase Via Transcriptase Reversa
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
EC 2.7.11.24 (p38 Mitogen-Activated Protein Kinases)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171010
[Lr] Data última revisão:
171010
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160414
[St] Status:MEDLINE


  7 / 165 MEDLINE  
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[PMID]:27071344
[Au] Autor:Jadwin JA; Oh D; Curran TG; Ogiue-Ikeda M; Jia L; White FM; Machida K; Yu J; Mayer BJ
[Ad] Endereço:Raymond and Beverly Sackler Laboratory of Molecular Medicine, Department of Genetics and Genome Sciences, University of Connecticut School of Medicine, Farmington, United States.
[Ti] Título:Time-resolved multimodal analysis of Src Homology 2 (SH2) domain binding in signaling by receptor tyrosine kinases.
[So] Source:Elife;5:e11835, 2016 Apr 12.
[Is] ISSN:2050-084X
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:While the affinities and specificities of SH2 domain-phosphotyrosine interactions have been well characterized, spatio-temporal changes in phosphosite availability in response to signals, and their impact on recruitment of SH2-containing proteins in vivo, are not well understood. To address this issue, we used three complementary experimental approaches to monitor phosphorylation and SH2 binding in human A431 cells stimulated with epidermal growth factor (EGF): 1) phospho-specific mass spectrometry; 2) far-Western blotting; and 3) live cell single-molecule imaging of SH2 membrane recruitment. Far-Western and MS analyses identified both well-established and previously undocumented EGF-dependent tyrosine phosphorylation and binding events, as well as dynamic changes in binding patterns over time. In comparing SH2 binding site phosphorylation with SH2 domain membrane recruitment in living cells, we found in vivo binding to be much slower. Delayed SH2 domain recruitment correlated with clustering of SH2 domain binding sites on the membrane, consistent with membrane retention via SH2 rebinding.
[Mh] Termos MeSH primário: Receptores Proteína Tirosina Quinases/metabolismo
Transdução de Sinais
Domínios de Homologia de src
[Mh] Termos MeSH secundário: Far-Western Blotting
Linhagem Celular Tumoral
Membrana Celular/metabolismo
Fator de Crescimento Epidérmico/metabolismo
Células Epiteliais/efeitos dos fármacos
Células Epiteliais/fisiologia
Seres Humanos
Espectrometria de Massas
Imagem Óptica
Ligação Proteica
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, N.I.H., EXTRAMURAL; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
62229-50-9 (Epidermal Growth Factor); EC 2.7.10.1 (Receptor Protein-Tyrosine Kinases)
[Em] Mês de entrada:1612
[Cu] Atualização por classe:171103
[Lr] Data última revisão:
171103
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160414
[St] Status:MEDLINE


  8 / 165 MEDLINE  
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[PMID]:26746014
[Au] Autor:Jun JH; Joo CK
[Ad] Endereço:Department of Ophthalmology, Keimyung University School of Medicine, Dongsan Medical Center, Daegu, Korea.
[Ti] Título:MicroRNA-124 Controls Transforming Growth Factor ß1-Induced Epithelial-Mesenchymal Transition in the Retinal Pigment Epithelium by Targeting RHOG.
[So] Source:Invest Ophthalmol Vis Sci;57(1):12-22, 2016 Jan 01.
[Is] ISSN:1552-5783
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:PURPOSE: MicroRNA-124 (miR-124) is thought to be involved in the epithelial-mesenchymal transition (EMT) of RPE. We investigated the regulation of TGF-ß1-induced EMT by miR-124 in human RPE cells (ARPE-19). METHODS: Expression of miR-124 was evaluated after TGF-ß1 treatment by quantitative RT-PCR. Phenotypic alterations were analyzed by Western blot analysis and immunocytochemical staining. Target validation was performed by a luciferase reporter assay to identify the putative target of miR-124. RESULTS: The expression level of miR-124 was downregulated during the progression of EMT. Overexpression of miR-124 upregulated the levels of zonular occludens 1 and occludin, and downregulated those of fibronectin, α-smooth muscle actin, and vimentin. Furthermore, inhibition of endogenous miR-124 increased and decreased the levels of mesenchymal and epithelial factors, respectively. TargetScan predicted two well-conserved and two vertebrate-only conserved miR-124 target sequences in the 3' untranslated region (UTR) of the Ras homology Growth-related (RHOG) mRNA. Direct targeting of this 3' UTR by miR-124 was demonstrated using a luciferase assay. Silencing of RHOG using a specific siRNA had identical effects on EMT regulation. Overexpression of miR-124 repressed TGF-ß1-induced RPE cell-collagen gel lattice contraction by altering cell spreading/cell-to-cell adhesion. CONCLUSIONS: This study describes the regulation of EMT in RPE cells by TGF-ß1/miR-124/RHOG signaling and suggests that the supplement of miR-124 exogenously would be a valuable therapeutic approach for the prevention or treatment of proliferative vitreoretinopathy.
[Mh] Termos MeSH primário: Transição Epitelial-Mesenquimal/efeitos dos fármacos
Regulação da Expressão Gênica
MicroRNAs/genética
Epitélio Pigmentado da Retina/metabolismo
Fator de Crescimento Transformador beta1/uso terapêutico
Vitreorretinopatia Proliferativa/genética
Proteínas rho de Ligação ao GTP/genética
[Mh] Termos MeSH secundário: Far-Western Blotting
Western Blotting
Linhagem Celular
Seres Humanos
Imuno-Histoquímica
MicroRNAs/biossíntese
RNA Mensageiro/genética
Reação em Cadeia da Polimerase em Tempo Real
Epitélio Pigmentado da Retina/patologia
Vitreorretinopatia Proliferativa/tratamento farmacológico
Vitreorretinopatia Proliferativa/metabolismo
Proteínas rho de Ligação ao GTP/biossíntese
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (MIRN124 microRNA, human); 0 (MicroRNAs); 0 (RNA, Messenger); 0 (Transforming Growth Factor beta1); 147605-13-8 (RHOG protein, human); EC 3.6.5.2 (rho GTP-Binding Proteins)
[Em] Mês de entrada:1605
[Cu] Atualização por classe:160109
[Lr] Data última revisão:
160109
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160110
[St] Status:MEDLINE
[do] DOI:10.1167/iovs.15-17111


  9 / 165 MEDLINE  
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[PMID]:26636044
[Au] Autor:Li Q; Liu H; Du D; Yu Y; Ma C; Jiao F; Yao H; Lu C; Zhang W
[Ad] Endereço:Key Lab of Animal Bacteriology, OIE Reference Lab for Swine Streptococcosis, College of Veterinary Medicine, Ministry of Agriculture, Nanjing Agricultural University Nanjing, China.
[Ti] Título:Identification of Novel Laminin- and Fibronectin-binding Proteins by Far-Western Blot: Capturing the Adhesins of Streptococcus suis Type 2.
[So] Source:Front Cell Infect Microbiol;5:82, 2015.
[Is] ISSN:2235-2988
[Cp] País de publicação:Switzerland
[La] Idioma:eng
[Ab] Resumo:Bacterial cell wall (CW) and extracellular (EC) proteins are often involved in interactions with extracellular matrix (ECM) proteins such as laminin (LN) and fibronectin (FN), which play important roles in adhesion and invasion. In this study, an efficient method combining proteomic analysis and Far-Western blotting assays was developed to screen directly for bacterial surface proteins with LN- and FN-binding capacity. With this approach, fifteen potential LN-binding proteins and five potential FN-binding proteins were identified from Streptococcus suis serotype 2 (SS2) CW and EC proteins. Nine newly identified proteins, including oligopeptide-binding protein OppA precursor (OppA), elongation factor Tu (EF-Tu), enolase, lactate dehydrogenase (LDH), fructose-bisphosphate aldolase (FBA), 3-ketoacyl-ACP reductase (KAR), Gly ceraldehyde-3-phosphate dehydrogenase (GAPDH), Inosine 5'-monophosphate dehydrogenase (IMPDH), and amino acid ABC transporter permease (ABC) were cloned, expressed, purified and further confirmed by Far-Western blotting and ELISA. Five proteins (OppA, EF-Tu, enolase, LDH, and FBA) exhibited specifically binding activity to both human LN and human FN. Furthermore, seven important recombinant proteins were selected and identified to have the ability to bind Hep-2 cells by the indirect immunofluorescent assay. In addition, four recombinant proteins, and their corresponding polyclonal antibodies, were observed to decrease SS2 adhesion to Hep-2 cells, which indicates that these proteins contribute to the adherence of SS2 to host cell surface. Collectively, these results show that the approach described here represents a useful tool for investigating the host-pathogen interactions.
[Mh] Termos MeSH primário: Adesinas Bacterianas/metabolismo
Far-Western Blotting/métodos
Fibronectinas/metabolismo
Laminina/metabolismo
Streptococcus suis/fisiologia
[Mh] Termos MeSH secundário: Linhagem Celular
Células Epiteliais/metabolismo
Seres Humanos
Ligação Proteica
Proteômica/métodos
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Adhesins, Bacterial); 0 (Fibronectins); 0 (Laminin)
[Em] Mês de entrada:1606
[Cu] Atualização por classe:151207
[Lr] Data última revisão:
151207
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:151205
[St] Status:MEDLINE
[do] DOI:10.3389/fcimb.2015.00082


  10 / 165 MEDLINE  
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[PMID]:26530384
[Au] Autor:Mohan Kumar D; Lin M; Xiong Q; Webber MJ; Kural C; Rikihisa Y
[Ad] Endereço:Department of Veterinary Biosciences, The Ohio State University, Columbus, Ohio, USA.
[Ti] Título:EtpE Binding to DNase X Induces Ehrlichial Entry via CD147 and hnRNP-K Recruitment, Followed by Mobilization of N-WASP and Actin.
[So] Source:MBio;6(6):e01541-15, 2015 Nov 03.
[Is] ISSN:2150-7511
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:UNLABELLED: Obligate intracellular bacteria, such as Ehrlichia chaffeensis, perish unless they can enter eukaryotic cells. E. chaffeensis is the etiological agent of human monocytic ehrlichiosis, an emerging infectious disease. To infect cells, Ehrlichia uses the C terminus of the outer membrane invasin entry-triggering protein (EtpE) of Ehrlichia (EtpE-C), which directly binds the mammalian cell surface glycosylphosphatidyl inositol-anchored protein, DNase X. How this binding drives Ehrlichia entry is unknown. Here, using affinity pulldown of host cell lysates with recombinant EtpE-C (rEtpE-C), we identified two new human proteins that interact with EtpE-C: CD147 and heterogeneous nuclear ribonucleoprotein K (hnRNP-K). The interaction of CD147 with rEtpE-C was validated by far-Western blotting and coimmunoprecipitation of native EtpE with endogenous CD147. CD147 was ubiquitous on the cell surface and also present around foci of rEtpE-C-coated-bead entry. Functional neutralization of surface-exposed CD147 with a specific antibody inhibited Ehrlichia internalization and infection but not binding. Downregulation of CD147 by short hairpin RNA (shRNA) impaired E. chaffeensis infection. Functional ablation of cytoplasmic hnRNP-K by a nanoscale intracellular antibody markedly attenuated bacterial entry and infection but not binding. EtpE-C also interacted with neuronal Wiskott-Aldrich syndrome protein (N-WASP), which is activated by hnRNP-K. Wiskostatin, which inhibits N-WASP activation, and cytochalasin D, which inhibits actin polymerization, inhibited Ehrlichia entry. Upon incubation with host cell lysate, EtpE-C but not an EtpE N-terminal fragment stimulated in vitro actin polymerization in an N-WASP- and DNase X-dependent manner. Time-lapse video images revealed N-WASP recruitment at EtpE-C-coated bead entry foci. Thus, EtpE-C binding to DNase X drives Ehrlichia entry by engaging CD147 and hnRNP-K and activating N-WASP-dependent actin polymerization. IMPORTANCE: Ehrlichia chaffeensis, an obligate intracellular bacterium, causes a blood-borne disease called human monocytic ehrlichiosis, one of the most prevalent life-threatening emerging tick-transmitted infectious diseases in the United States. The survival of Ehrlichia bacteria, and hence, their ability to cause disease, depends on their specific mode of entry into eukaryotic host cells. Understanding the mechanism by which E. chaffeensis enters cells will create new opportunities for developing effective therapies to prevent bacterial entry and disease in humans. Our findings reveal a novel cellular signaling pathway triggered by an ehrlichial surface protein called EtpE to induce its infectious entry. The results are also important from the viewpoint of human cell physiology because three EtpE-interacting human proteins, DNase X, CD147, and hnRNP-K, are hitherto unknown partners that drive the uptake of small particles, including bacteria, into human cells.
[Mh] Termos MeSH primário: Proteínas da Membrana Bacteriana Externa/metabolismo
Basigina/metabolismo
Desoxirribonucleases/metabolismo
Ehrlichia chaffeensis/fisiologia
Endocitose
Ribonucleoproteínas Nucleares Heterogêneas Grupo K/metabolismo
Interações Hospedeiro-Patógeno
[Mh] Termos MeSH secundário: Actinas/metabolismo
Animais
Far-Western Blotting
Ehrlichia chaffeensis/metabolismo
Seres Humanos
Imunoprecipitação
Camundongos Endogâmicos C57BL
Camundongos Knockout
Ligação Proteica
Mapeamento de Interação de Proteínas
Proteína Neuronal da Síndrome de Wiskott-Aldrich/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Actins); 0 (BSG protein, human); 0 (Bacterial Outer Membrane Proteins); 0 (Heterogeneous-Nuclear Ribonucleoprotein K); 0 (WASL protein, human); 0 (Wiskott-Aldrich Syndrome Protein, Neuronal); 136894-56-9 (Basigin); EC 3.1.- (Deoxyribonucleases)
[Em] Mês de entrada:1608
[Cu] Atualização por classe:171116
[Lr] Data última revisão:
171116
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:151105
[St] Status:MEDLINE



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