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[PMID]:27770364
[Au] Autor:Blevins T
[Ad] Endereço:Institut de Biologie Moléculaire des Plantes, Centre National de la Recherche Scientifique (CNRS) UPR2357, 12 rue du Général Zimmer, Strasbourg Cedex, 67084, USA. todd.blevins@ibmp-cnrs.unistra.fr.
[Ti] Título:Northern Blotting Techniques for Small RNAs.
[So] Source:Methods Mol Biol;1456:141-162, 2017.
[Is] ISSN:1940-6029
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Cells have evolved intricate RNA-directed mechanisms that destroy viruses, silence transposons, and regulate gene expression. These nucleic acid surveillance and gene silencing mechanisms rely upon the selective base-pairing of ~19-25 nt small RNAs to complementary RNA targets. This chapter describes northern blot hybridization techniques for the detection of such small RNAs. Blots spiked with synthetic standards are used to illustrate the detection specificity and sensitivity of DNA oligonucleotide probes. Known endogenous small RNAs are then analyzed in samples prepared from several model plants, including Arabidopsis thaliana, Nicotiana benthamiana, Oryza sativa, Zea mays, and Physcomitrella patens, as well as from the animals Drosophila melanogaster and Mus musculus. Finally, the value of northern blotting for dissecting small RNA biogenesis is shown using an example of virus infection in A. thaliana.
[Mh] Termos MeSH primário: Northern Blotting
Pequeno RNA não Traduzido
[Mh] Termos MeSH secundário: Northern Blotting/métodos
Inativação Gênica
MicroRNAs/genética
Hibridização de Ácido Nucleico/métodos
Interferência de RNA
RNA Interferente Pequeno/genética
Pequeno RNA não Traduzido/isolamento & purificação
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (MicroRNAs); 0 (RNA, Small Interfering); 0 (RNA, Small Untranslated)
[Em] Mês de entrada:1801
[Cu] Atualização por classe:180112
[Lr] Data última revisão:
180112
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161023
[St] Status:MEDLINE


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[PMID]:28827831
[Au] Autor:Wenger C; Oeljeklaus S; Warscheid B; Schneider A; Harsman A
[Ad] Endereço:Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, Bern, Switzerland.
[Ti] Título:A trypanosomal orthologue of an intermembrane space chaperone has a non-canonical function in biogenesis of the single mitochondrial inner membrane protein translocase.
[So] Source:PLoS Pathog;13(8):e1006550, 2017 Aug.
[Is] ISSN:1553-7374
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Mitochondrial protein import is essential for Trypanosoma brucei across its life cycle and mediated by membrane-embedded heterooligomeric protein complexes, which mainly consist of trypanosomatid-specific subunits. However, trypanosomes contain orthologues of small Tim chaperones that escort hydrophobic proteins across the intermembrane space. Here we have experimentally analyzed three novel trypanosomal small Tim proteins, one of which contains only an incomplete Cx3C motif. RNAi-mediated ablation of TbERV1 shows that their import, as in other organisms, depends on the MIA pathway. Submitochondrial fractionation combined with immunoprecipitation and BN-PAGE reveals two pools of small Tim proteins: a soluble fraction forming 70 kDa complexes, consistent with hexamers and a second fraction that is tightly associated with the single trypanosomal TIM complex. RNAi-mediated ablation of the three proteins leads to a growth arrest and inhibits the formation of the TIM complex. In line with these findings, the changes in the mitochondrial proteome induced by ablation of one small Tim phenocopy the effects observed after ablation of TbTim17. Thus, the trypanosomal small Tims play an unexpected and essential role in the biogenesis of the single TIM complex, which for one of them is not linked to import of TbTim17.
[Mh] Termos MeSH primário: Proteínas de Membrana Transportadoras/metabolismo
Proteínas de Transporte da Membrana Mitocondrial/metabolismo
Chaperonas Moleculares/metabolismo
Transporte Proteico/fisiologia
Trypanosoma brucei brucei/crescimento & desenvolvimento
[Mh] Termos MeSH secundário: Northern Blotting
Cromatografia Líquida de Alta Pressão
Imunoprecipitação
Estágios do Ciclo de Vida
Espectrometria de Massas
Microscopia de Fluorescência
Membranas Mitocondriais/metabolismo
Trypanosoma brucei brucei/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Membrane Transport Proteins); 0 (Mitochondrial Membrane Transport Proteins); 0 (Molecular Chaperones)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171002
[Lr] Data última revisão:
171002
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170823
[St] Status:MEDLINE
[do] DOI:10.1371/journal.ppat.1006550


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[PMID]:28728902
[Au] Autor:Chandra S; Vimal D; Sharma D; Rai V; Gupta SC; Chowdhuri DK
[Ad] Endereço:Embryotoxicology Laboratory, Environmental Toxicology Group, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Lucknow 226 001, India; Academy of Scientific Innovation & Research (AcSIR), CSIR-IITR Campus, Lucknow, India.
[Ti] Título:Role of miRNAs in development and disease: Lessons learnt from small organisms.
[So] Source:Life Sci;185:8-14, 2017 Sep 15.
[Is] ISSN:1879-0631
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:MicroRNAs (miRNAs) constitute a class of small (18-22 nucleotides) non-coding RNAs that regulate gene expression at the post-transcriptional level. Caenorhabditis elegans, Drosophila melanogaster, and many other small organisms have been instrumental in deciphering the biological functions of miRNAs. While some miRNAs from small organisms are highly conserved across the taxa, others are organism specific. The miRNAs are known to play a crucial role during development and in various cellular functions such as cell survival, cell proliferation, and differentiation. The miRNAs associated with fragile X syndrome, Parkinson's disease, Alzheimer's disease, diabetes, cancer, malaria, infectious diseases and several other human diseases have been identified from small organisms. These organisms have been used as platforms in deciphering the functions of miRNAs in the pathogenesis of human diseases and to study miRNA biogenesis. Small organisms have also been used in the development of miRNA-based diagnostic, prognostic and therapeutic strategies. The molecular techniques such as genome sequencing, northern blot analysis, and quantitative RT-PCR, have been used in deciphering the functions of miRNAs in small organisms. How miRNAs from small organisms especially those from Drosophila and C. elegans regulate development and disease pathogenesis is the focus of this review. The outstanding questions raised by our current understanding are discussed.
[Mh] Termos MeSH primário: Regulação da Expressão Gênica/genética
MicroRNAs/genética
[Mh] Termos MeSH secundário: Animais
Northern Blotting
Caenorhabditis elegans/genética
Diferenciação Celular/genética
Proliferação Celular/genética
Sobrevivência Celular/genética
Drosophila melanogaster/genética
Seres Humanos
Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos
[Pt] Tipo de publicação:JOURNAL ARTICLE; REVIEW
[Nm] Nome de substância:
0 (MicroRNAs)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170908
[Lr] Data última revisão:
170908
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170722
[St] Status:MEDLINE


  4 / 45698 MEDLINE  
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[PMID]:28586482
[Au] Autor:Evans ME; Clark WC; Zheng G; Pan T
[Ad] Endereço:Department of Biochemistry & Molecular Biology, University of Chicago, Chicago, IL 60637, USA.
[Ti] Título:Determination of tRNA aminoacylation levels by high-throughput sequencing.
[So] Source:Nucleic Acids Res;45(14):e133, 2017 Aug 21.
[Is] ISSN:1362-4962
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Transfer RNA (tRNA) decodes mRNA codons when aminoacylated (charged) with an amino acid at its 3' end. Charged tRNAs turn over rapidly in cells, and variations in charged tRNA fractions are known to be a useful parameter in cellular responses to stress. tRNA charging fractions can be measured for individual tRNA species using acid denaturing gels, or comparatively at the genome level using microarrays. These hybridization-based approaches cannot be used for high resolution analysis of mammalian tRNAs due to their large sequence diversity. Here we develop a high-throughput sequencing method that enables accurate determination of charged tRNA fractions at single-base resolution (Charged DM-tRNA-seq). Our method takes advantage of the recently developed DM-tRNA-seq method, but includes additional chemical steps that specifically remove the 3'A residue in uncharged tRNA. Charging fraction is obtained by counting the fraction of A-ending reads versus A+C-ending reads for each tRNA species in the same sequencing reaction. In HEK293T cells, most cytosolic tRNAs are charged at >80% levels, whereas tRNASer and tRNAThr are charged at lower levels. These low charging levels were validated using acid denaturing gels. Our method should be widely applicable for investigations of tRNA charging as a parameter in biological regulation.
[Mh] Termos MeSH primário: Sequenciamento de Nucleotídeos em Larga Escala/métodos
Aminoacil-RNA de Transferência/genética
RNA de Transferência/genética
Aminoacilação de RNA de Transferência/genética
[Mh] Termos MeSH secundário: Aminoacilação
Northern Blotting
Células HEK293
Seres Humanos
Modelos Genéticos
RNA de Transferência/metabolismo
Aminoacil-RNA de Transferência/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (RNA, Transfer, Amino Acyl); 9014-25-9 (RNA, Transfer)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171023
[Lr] Data última revisão:
171023
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170607
[St] Status:MEDLINE
[do] DOI:10.1093/nar/gkx514


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[PMID]:28586459
[Au] Autor:Huang K; Doyle F; Wurz ZE; Tenenbaum SA; Hammond RK; Caplan JL; Meyers BC
[Ad] Endereço:Bio-Imaging Center, Delaware Biotechnology Institute, University of Delaware, Newark, DE 19711, USA.
[Ti] Título:FASTmiR: an RNA-based sensor for in vitro quantification and live-cell localization of small RNAs.
[So] Source:Nucleic Acids Res;45(14):e130, 2017 Aug 21.
[Is] ISSN:1362-4962
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Small RNAs, including microRNAs (miRNAs) and small interfering RNAs (siRNAs), play a variety of important regulatory roles in many eukaryotes. Their small size has made it challenging to study them directly in live cells. Here we describe an RNA-based fluorescent sensor for small RNA detection both in vitro and in vivo, adaptable for any small RNA. It utilizes an sxRNA switch for detection of miRNA-mRNA interactions combined with a fluorophore-binding sequence 'Spinach', a GFP-like RNA aptamer for which the RNA-fluorophore complex exhibits strong and consistent fluorescence under an excitation wavelength. Two example sensors, FASTmiR171 and FASTmiR122, can rapidly detect and quantify the levels of miR171 and miR122 in vitro. The sensors can determine relative levels of miRNAs in total RNA extracts with sensitivity similar to small RNA sequencing and northern blots. FASTmiR sensors were also used to estimate the copy number range of miRNAs in total RNA extracts. To localize and analyze the spatial distribution of small RNAs in live, single cells, tandem copies of FASTmiR122 were expressed in different cell lines. FASTmiR122 was able to quantitatively detect the differences in miR122 levels in Huh7 and HEK293T cells demonstrating its potential for tracking miRNA expression and localization in vivo.
[Mh] Termos MeSH primário: Técnicas Biossensoriais/métodos
MicroRNAs/genética
RNA de Plantas/genética
Spinacia oleracea/genética
[Mh] Termos MeSH secundário: Sequência de Bases
Northern Blotting
Linhagem Celular Tumoral
Núcleo Celular/metabolismo
Citoplasma/metabolismo
Corantes Fluorescentes/química
Proteínas de Fluorescência Verde/genética
Proteínas de Fluorescência Verde/metabolismo
Células HEK293
Seres Humanos
Hibridização in Situ Fluorescente
MicroRNAs/metabolismo
Microscopia de Fluorescência
RNA de Plantas/metabolismo
Reprodutibilidade dos Testes
Reação em Cadeia da Polimerase Via Transcriptase Reversa
Spinacia oleracea/citologia
Spinacia oleracea/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Fluorescent Dyes); 0 (MIRN122 microRNA, human); 0 (MicroRNAs); 0 (RNA, Plant); 147336-22-9 (Green Fluorescent Proteins)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171023
[Lr] Data última revisão:
171023
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170607
[St] Status:MEDLINE
[do] DOI:10.1093/nar/gkx504


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[PMID]:28446464
[Au] Autor:Huang B; Yang H; Cheng X; Wang D; Fu S; Shen W; Zhang Q; Zhang L; Xue Z; Li Y; Da Y; Yang Q; Li Z; Liu L; Qiao L; Kong Y; Yao Z; Zhao P; Li M; Zhang R
[Ad] Endereço:Department of Immunology, Research Center of Basic Medical Sciences, Key Laboratory of Immune Microenvironment and Diseases of Educational Ministry of China, Tianjin Medical University, Tianjin, China.
[Ti] Título:tRF/miR-1280 Suppresses Stem Cell-like Cells and Metastasis in Colorectal Cancer.
[So] Source:Cancer Res;77(12):3194-3206, 2017 Jun 15.
[Is] ISSN:1538-7445
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Several studies have shown that tRNAs can be enzymatically cleaved to generate distinct classes of tRNA-derived fragments (tRF). Here, we report that tRF/miR-1280, a 17-bp fragment derived from tRNA and pre-miRNA, influences Notch signaling pathways that support the function of cancer stem-like cells (CSC) in colorectal cancer progression. tRF/miR-1280 expression was decreased in human specimens of colorectal cancer. Ectopic expression of tRF/miR-1280 reduced cell proliferation and colony formation, whereas its suppression reversed these effects. Mechanistic investigations implicated the Notch ligand JAG2 as a direct target of tRF/miR-1280 binding through which it reduced tumor formation and metastasis. Notably, tRF/miR-1280-mediated inactivation of Notch signaling suppressed CSC phenotypes, including by direct transcriptional repression of the Gata1/3 and miR-200b genes. These results were consistent with findings of decreased levels of miR-200b and elevated levels of JAG2, Gata1, Gata3, Zeb1, and Suz12 in colorectal cancer tissue specimens. Taken together, our results established that tRF/miR-1280 suppresses colorectal cancer growth and metastasis by repressing Notch signaling pathways that support CSC phenotypes. Furthermore, they provide evidence that functionally active miRNA can be derived from tRNA, offering potential biomarker and therapeutic uses. .
[Mh] Termos MeSH primário: Neoplasias Colorretais/patologia
MicroRNAs
Invasividade Neoplásica/genética
Células-Tronco Neoplásicas/patologia
RNA de Transferência
[Mh] Termos MeSH secundário: Animais
Northern Blotting
Western Blotting
Imunoprecipitação da Cromatina
Neoplasias Colorretais/genética
Citometria de Fluxo
Regulação Neoplásica da Expressão Gênica/fisiologia
Xenoenxertos
Seres Humanos
Camundongos
Camundongos Endogâmicos BALB C
Camundongos Nus
Invasividade Neoplásica/patologia
Reação em Cadeia da Polimerase
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (MIRN-1280 microRNA, human); 0 (MicroRNAs); 9014-25-9 (RNA, Transfer)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170914
[Lr] Data última revisão:
170914
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170428
[St] Status:MEDLINE
[do] DOI:10.1158/0008-5472.CAN-16-3146


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[PMID]:28400331
[Au] Autor:Thiruketheeswaran P; Thomalla P; Krüger E; Hinssen H; D'Haese J
[Ad] Endereço:Institute for Cell Biology, Department Biology, Heinrich-Heine-University Düsseldorf, Universitätsstrasse 1, D-40225 Düsseldorf, Germany.
[Ti] Título:Four paralog gelsolin genes are differentially expressed in the earthworm Lumbricus terrestris.
[So] Source:Comp Biochem Physiol B Biochem Mol Biol;208-209:58-67, 2017 Jun.
[Is] ISSN:1879-1107
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:We have identified and characterized four distinct variants of the gelsolin-related protein (EWAM P1-P4) in the earthworm L. terrestris. All of these proteins biochemically qualify as gelsolins since they sever actin filaments in a calcium dependent manner. P1, P2 and P3 are present in the Lumbricus body wall muscle whereas in the gizzard muscle P3 and P4 were found. P1-P4 are encoded by four paralog genes and are differentially expressed in various muscle cell tissues. While the genes for P1 and P2 contain one intron, there was no intron in both P3 and P4 genes. The coding sequences consist of 1104bp (368 amino acids) for P1/P4 and 1101bp (367 amino acids) for P2/P3. Corresponding genes were confirmed by northern blot analysis which revealed three (calculated lengths: 3100, 2300 and 2100 nucleotides) and two (calculated lengths: 2300 and 1700 nucleotides) mRNA transcripts in the body wall and the gizzard, respectively. EWAM mRNA was localized by fluorescence in situ hybridization in the body wall and the gizzard muscle. P1 mRNA was detected in the inner proximal layers of both the circular and longitudinal muscle of the body wall whereas in the gizzard no significant staining was observed for P1. P2-P4 mRNAs were abundant in the outer distal layers of both the circular and the longitudinal muscles of both body wall and gizzard. The differential expression of four paralog gelsolin genes suggests a functional adaptation of different muscle cells with respect to actin filament turnover and modulation of its polymer state.
[Mh] Termos MeSH primário: Éxons/genética
Gelsolina/genética
Íntrons/genética
Músculos/metabolismo
Oligoquetos/genética
[Mh] Termos MeSH secundário: Animais
Northern Blotting
Gelsolina/classificação
Hibridização in Situ Fluorescente
Oligoquetos/crescimento & desenvolvimento
RNA Mensageiro/genética
Reação em Cadeia da Polimerase em Tempo Real
Reação em Cadeia da Polimerase Via Transcriptase Reversa
[Pt] Tipo de publicação:COMPARATIVE STUDY; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Gelsolin); 0 (RNA, Messenger)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170623
[Lr] Data última revisão:
170623
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170413
[St] Status:MEDLINE


  8 / 45698 MEDLINE  
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[PMID]:28348459
[Au] Autor:Dong Y; Wu Y; Cui MZ; Xu X
[Ad] Endereço:Department of Biomedical & Diagnostic Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996, USA; Vascular Biology Program, Boston Children's Hospital, Harvard Medical School, Boston, MA 02115, USA.
[Ti] Título:Lysophosphatidic Acid Triggers Apoptosis in HeLa Cells through the Upregulation of Tumor Necrosis Factor Receptor Superfamily Member 21.
[So] Source:Mediators Inflamm;2017:2754756, 2017.
[Is] ISSN:1466-1861
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Lysophosphatidic acid (LPA), a naturally occurring bioactive phospholipid, activates G protein-coupled receptors (GPCRs), leading to regulation of diverse cellular events including cell survival and apoptosis. Despite extensive studies of the signaling pathways that mediate LPA-regulated cell growth and survival, the mechanisms underlying the apoptotic effect of LPA remain largely unclear. In this study, we investigated this issue in HeLa cells. Our data demonstrate that LPA induces apoptosis in HeLa cells at pathologic concentrations with a concomitant upregulation of the expression of TNFRSF21 (tumor necrosis factor receptor superfamily member 21), also known as death receptor number 6 (DR6) involved in inflammation. Moreover, treatment of cells with LPA receptor (LPAR) antagonist abolished the DR6 upregulation by LPA. LPA-induced DR6 expression was also abrogated by pertussis toxin (PTX), an inhibitor of GPCRs, and by inhibitors of PI3K, PKC, MEK, and ERK. Intriguingly, LPA-induced DR6 expression was specifically blocked by dominant-negative form of PKC (PKC -DN). LPA-induced DR6 expression was also dramatically inhibited by knockdown of ERK or CREB. These results suggest that activation of the MEK/ERK pathway and the transcription factor CREB mediate LPA-induced DR6 expression. More interestingly, knockdown of DR6 using siRNA approach remarkably attenuated LPA-induced apoptosis. In conclusion, our results suggest that LPA-induced apoptosis in HeLa cells is mediated by the upregulation of DR6 expression.
[Mh] Termos MeSH primário: Lisofosfolipídeos/farmacologia
Receptores do Fator de Necrose Tumoral/metabolismo
[Mh] Termos MeSH secundário: Apoptose/efeitos dos fármacos
Northern Blotting
Western Blotting
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo
MAP Quinases Reguladas por Sinal Extracelular/genética
MAP Quinases Reguladas por Sinal Extracelular/metabolismo
Células HeLa
Seres Humanos
Marcação In Situ das Extremidades Cortadas
Sistema de Sinalização das MAP Quinases/efeitos dos fármacos
Toxina Pertussis/farmacologia
Fosfatidilinositol 3-Quinases/antagonistas & inibidores
Fosfatidilinositol 3-Quinases/metabolismo
RNA Interferente Pequeno/genética
RNA Interferente Pequeno/fisiologia
Transdução de Sinais/efeitos dos fármacos
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (CREB1 protein, human); 0 (Cyclic AMP Response Element-Binding Protein); 0 (Lysophospholipids); 0 (RNA, Small Interfering); 0 (Receptors, Tumor Necrosis Factor); 0 (TNFRSF21 protein, human); EC 2.4.2.31 (Pertussis Toxin); EC 2.7.1.- (Phosphatidylinositol 3-Kinases); EC 2.7.11.24 (Extracellular Signal-Regulated MAP Kinases); PG6M3969SG (lysophosphatidic acid)
[Em] Mês de entrada:1707
[Cu] Atualização por classe:170802
[Lr] Data última revisão:
170802
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170329
[St] Status:MEDLINE
[do] DOI:10.1155/2017/2754756


  9 / 45698 MEDLINE  
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[PMID]:28346491
[Au] Autor:Anjuwon-Foster BR; Tamayo R
[Ad] Endereço:Department of Microbiology and Immunology, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina, United States of America.
[Ti] Título:A genetic switch controls the production of flagella and toxins in Clostridium difficile.
[So] Source:PLoS Genet;13(3):e1006701, 2017 Mar.
[Is] ISSN:1553-7404
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:In the human intestinal pathogen Clostridium difficile, flagella promote adherence to intestinal epithelial cells. Flagellar gene expression also indirectly impacts production of the glucosylating toxins, which are essential to diarrheal disease development. Thus, factors that regulate the expression of the flgB operon will likely impact toxin production in addition to flagellar motility. Here, we report the identification a "flagellar switch" that controls the phase variable production of flagella and glucosylating toxins. The flagellar switch, located upstream of the flgB operon containing the early stage flagellar genes, is a 154 bp invertible sequence flanked by 21 bp inverted repeats. Bacteria with the sequence in one orientation expressed flagellum and toxin genes, produced flagella, and secreted the toxins ("flg phase ON"). Bacteria with the sequence in the inverse orientation were attenuated for flagellar and toxin gene expression, were aflagellate, and showed decreased toxin secretion ("flg phase OFF"). The orientation of the flagellar switch is reversible during growth in vitro. We provide evidence that gene regulation via the flagellar switch occurs post-transcription initiation and requires a C. difficile-specific regulatory factor to destabilize or degrade the early flagellar gene mRNA when the flagellar switch is in the OFF orientation. Lastly, through mutagenesis and characterization of flagellar phase locked isolates, we determined that the tyrosine recombinase RecV, which catalyzes inversion at the cwpV switch, is also responsible for inversion at the flagellar switch in both directions. Phase variable flagellar motility and toxin production suggests that these important virulence factors have both advantageous and detrimental effects during the course of infection.
[Mh] Termos MeSH primário: Proteínas de Bactérias/genética
Toxinas Bacterianas/genética
Clostridium difficile/genética
Flagelos/genética
Regulação Bacteriana da Expressão Gênica
[Mh] Termos MeSH secundário: Proteínas de Bactérias/metabolismo
Toxinas Bacterianas/metabolismo
Sequência de Bases
Northern Blotting
Western Blotting
Clostridium difficile/metabolismo
Clostridium difficile/fisiologia
Enterocolite Pseudomembranosa/microbiologia
Flagelos/metabolismo
Flagelos/fisiologia
Seres Humanos
Intestinos/microbiologia
Microscopia de Fluorescência
Movimento/fisiologia
Óperon/genética
RNA Mensageiro/genética
RNA Mensageiro/metabolismo
Sequências Reguladoras de Ácido Nucleico/genética
Reação em Cadeia da Polimerase Via Transcriptase Reversa
Homologia de Sequência do Ácido Nucleico
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Bacterial Toxins); 0 (RNA, Messenger)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170607
[Lr] Data última revisão:
170607
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170328
[St] Status:MEDLINE
[do] DOI:10.1371/journal.pgen.1006701


  10 / 45698 MEDLINE  
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[PMID]:28278298
[Au] Autor:Wang F; Fang Q; Wang B; Yan Z; Hong J; Bao Y; Kuhn JH; Werren JH; Song Q; Ye G
[Ad] Endereço:State Key Laboratory of Rice Biology & Ministry of Agriculture Key Lab of Molecular Biology of Crop Pathogens and Insects, Institute of Insect Sciences, Zhejiang University, Hangzhou, China.
[Ti] Título:A novel negative-stranded RNA virus mediates sex ratio in its parasitoid host.
[So] Source:PLoS Pathog;13(3):e1006201, 2017 Mar.
[Is] ISSN:1553-7374
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Parasitoid wasps are important natural enemies of arthropod hosts in natural and agricultural ecosystems and are often associated with viruses or virion-like particles. Here, we report a novel negative-stranded RNA virus from a parasitoid wasp (Pteromalus puparum). The complete viral genome is 12,230 nucleotides in length, containing five non-overlapping, linearly arranged open reading frames. Phylogenetically, the virus clusters with and is a novel member of the mononegaviral family Nyamiviridae, here designated as Pteromalus puparum negative-strand RNA virus 1 (PpNSRV-1). PpNSRV-1 is present in various tissues and life stages of the parasitoid wasp, and is transmitted vertically through infected females and males. Virus infections in field populations of P. puparum wasps ranged from 16.7 to 37.5%, without linearly correlating with temperature. PpNSRV-1 increased adult longevity and impaired several fitness parameters of the wasp, but had no influence on successful parasitism. Strikingly, PpNSRV-1 mediated the offspring sex ratio by decreasing female offspring numbers. RNA interference knockdown of virus open reading frame I eliminated these PpNSRV-1-induced effects. Thus, we infer that PpNSRV-1 has complex effects on its insect host including sex ratio distortion towards males, as well as possible mutualistic benefits through increasing wasp longevity.
[Mh] Termos MeSH primário: Infecções por Vírus de RNA/transmissão
Vírus de RNA/fisiologia
Vespas/virologia
[Mh] Termos MeSH secundário: Animais
Northern Blotting
Feminino
Imuno-Histoquímica
Masculino
Microscopia Eletrônica de Transmissão
Filogenia
Reação em Cadeia da Polimerase
Razão de Masculinidade
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Em] Mês de entrada:1707
[Cu] Atualização por classe:170718
[Lr] Data última revisão:
170718
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170310
[St] Status:MEDLINE
[do] DOI:10.1371/journal.ppat.1006201



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