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  • Bacterial Proteins
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  • 1
    Language: English
    In: Proceedings of the National Academy of Sciences of the United States of America, 27 March 2012, Vol.109(13), pp.E757-64
    Description: SgrS RNA is a model for the large class of Hfq-associated small RNAs that act to posttranscriptionally regulate bacterial mRNAs. The function of SgrS is well-characterized in nonpathogenic Escherichia coli, where it was originally shown to counteract glucose-phosphate stress by acting as a repressor of the ptsG mRNA, which encodes the major glucose transporter. We have discovered additional SgrS targets in Salmonella Typhimurium, a pathogen related to E. coli that recently acquired one-quarter of all genes by horizontal gene transfer. We show that the conserved short seed region of SgrS that recognizes ptsG was recruited to target the Salmonella-specific sopD mRNA of a secreted virulence protein. The SgrS-sopD interaction is exceptionally selective; we find that sopD2 mRNA, whose gene arose from sopD duplication during Salmonella evolution, is deaf to SgrS because of a nonproductive G-U pair in the potential SgrS-sopD2 RNA duplex vs. G-C in SgrS-sopD. In other words, SgrS discriminates the two virulence factor mRNAs at the level of a single hydrogen bond. Our study suggests that bacterial pathogens use their large suites of conserved Hfq-associated regulators to integrate horizontally acquired genes into existing posttranscriptional networks, just as conserved transcription factors are recruited to tame foreign genes at the DNA level. The results graphically illustrate the importance of the seed regions of bacterial small RNAs to select new targets with high fidelity and suggest that target predictions must consider all or none decisions by individual seed nucleotides.
    Keywords: Phylogeny ; Base Pairing -- Genetics ; Gene Transfer, Horizontal -- Genetics ; RNA, Bacterial -- Genetics ; Salmonella -- Genetics
    ISSN: 00278424
    E-ISSN: 1091-6490
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  • 2
    In: Nature, 2011, Vol.471(7340), p.602
    Description: CRISPR/Cas systems constitute a widespread class of immunity systems that protect bacteria and archaea against phages and plasmids, and commonly use repeat/spacer-derived short crRNAs to silence foreign nucleic acids in a sequence-specific manner. Although the maturation of crRNAs represents a key event in CRISPR activation, the responsible endoribonucleases (CasE, Cas6, Csy4) are missing in many CRISPR/Cas subtypes. Here, differential RNA sequencing of the human pathogen Streptococcus pyogenes uncovered tracrRNA, a trans-encoded small RNA with 24-nucleotide complementarity to the repeat regions of crRNA precursor transcripts. We show that tracrRNA directs the maturation of crRNAs by the activities of the widely conserved endogenous RNase III and the CRISPR-associated Csn1 protein; all these components are essential to protect S. pyogenes against prophage-derived DNA. Our study reveals a novel pathway of small guide RNA maturation and the first example of a host factor (RNase III) required for bacterial RNA-mediated immunity against invaders. [PUBLICATION ]
    Keywords: Bacterial Proteins–Chemistry ; Bacterial Proteins–Genetics ; Bacterial Proteins–Immunology ; Bacterial Proteins–Metabolism ; Conserved Sequence–Genetics ; DNA, Viral–Metabolism ; DNA, Viral–Genetics ; Escherichia Coli–Genetics ; Models, Biological–Metabolism ; Prophages–Biosynthesis ; RNA Precursors–Genetics ; RNA Precursors–Immunology ; RNA Processing, Post-Transcriptional–Metabolism ; RNA, Bacterial–Genetics ; RNA, Bacterial–Metabolism ; RNA, Bacterial–Genetics ; RNA, Bacterial–Immunology ; RNA, Guide–Metabolism ; Ribonuclease III–Virology ; Streptococcus Pyogenes–Virology ; Streptococcus Pyogenes–Virology ; Streptococcus Pyogenes–Virology ; Streptococcus Pyogenes–Virology ; E Coli ; Bacteria ; Bacteriology ; Plasmids ; Proteins ; Bacterial Proteins ; DNA, Viral ; RNA Precursors ; RNA, Bacterial ; RNA, Guide ; Ribonuclease III;
    ISSN: 0028-0836
    E-ISSN: 14764687
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  • 3
    Language: English
    In: Proceedings of the National Academy of Sciences of the United States of America, 27 June 2017, Vol.114(26), pp.6824-6829
    Description: The functions of many bacterial RNA-binding proteins remain obscure because of a lack of knowledge of their cellular ligands. Although well-studied cold-shock protein A (CspA) family members are induced and function at low temperature, others are highly expressed in infection-relevant conditions. Here, we have profiled transcripts bound in vivo by the CspA family members of serovar Typhimurium to link the constitutively expressed CspC and CspE proteins with virulence pathways. Phenotypic assays in vitro demonstrated a crucial role for these proteins in membrane stress, motility, and biofilm formation. Moreover, double deletion of and fully attenuates in systemic mouse infection. In other words, the RNA ligand-centric approach taken here overcomes a problematic molecular redundancy of CspC and CspE that likely explains why these proteins have evaded selection in previous virulence factor screens in animals. Our results highlight RNA-binding proteins as regulators of pathogenicity and potential targets of antimicrobial therapy. They also suggest that globally acting RNA-binding proteins are more common in bacteria than currently appreciated.
    Keywords: RNA-Binding Protein ; Salmonella ; Bacterial Pathogenesis ; Cold-Shock Protein ; Stress Response ; Bacterial Proteins ; Cold Shock Proteins and Peptides ; Heat-Shock Proteins ; RNA-Binding Proteins ; Salmonella Infections ; Salmonella Typhimurium ; Virulence Factors
    ISSN: 00278424
    E-ISSN: 1091-6490
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  • 4
    In: Molecular Microbiology, December 2010, Vol.78(6), pp.1327-1331
    Description: Although most bacterial small RNAs act to repress target mRNAs, some also activate messengers. The predominant mode of activation has been seen in ‘anti‐antisense’ regulation whereby a small RNA prevents the formation of an inhibitory 5′ mRNA structure that otherwise impairs translational initiation and protein synthesis. The translational activation might also stabilize the target yet this was considered a secondary effect in the examples known thus far. Two recent papers in investigate post‐transcriptional activation of collagenase mRNA by VR‐RNA, and streptokinase mRNA by FasX RNA, to suggest that small RNAs exert positive regulation of virulence genes primarily at the level of mRNA stabilization.
    Keywords: Protein Synthesis ; Messenger Rna;
    ISSN: 0950-382X
    E-ISSN: 1365-2958
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  • 5
    Language: English
    In: PLoS ONE, 2011, Vol.6(3), p.e17296
    Description: P-bodies are dynamic aggregates of RNA and proteins involved in several post-transcriptional regulation processes. P-bodies have been shown to play important roles in regulating viral infection, whereas their interplay with bacterial pathogens, specifically intracellular bacteria that extensively manipulate host cell pathways, remains unknown. Here, we report that Salmonella infection induces P-body disassembly in a cell type-specific manner, and independently of previously characterized pathways such as inhibition of host cell RNA synthesis or microRNA-mediated gene silencing. We show that the Salmonella -induced P-body disassembly depends on the activation of the SPI-2 encoded type 3 secretion system, and that the secreted effector protein SpvB plays a major role in this process. P-body disruption is also induced by the related pathogen, Shigella flexneri , arguing that this might be a new mechanism by which intracellular bacterial pathogens subvert host cell function.
    Keywords: Research Article ; Biology ; Medicine ; Infectious Diseases ; Microbiology ; Molecular Biology ; Cell Biology
    E-ISSN: 1932-6203
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  • 6
    Language: English
    In: Proceedings of the National Academy of Sciences of the United States of America, 11 October 2016, Vol.113(41), pp.11591-11596
    Description: The functional annotation of transcriptomes and identification of noncoding RNA (ncRNA) classes has been greatly facilitated by the advent of next-generation RNA sequencing which, by reading the nucleotide order of transcripts, theoretically allows the rapid profiling of all transcripts in a cell. However, primary sequence per se is a poor predictor of function, as ncRNAs dramatically vary in length and structure and often lack identifiable motifs. Therefore, to visualize an informative RNA landscape of organisms with potentially new RNA biology that are emerging from microbiome and environmental studies requires the use of more functionally relevant criteria. One such criterion is the association of RNAs with functionally important cognate RNA-binding proteins. Here we analyze the full ensemble of cellular RNAs using gradient profiling by sequencing (Grad-seq) in the bacterial pathogen Salmonella enterica, partitioning its coding and noncoding transcripts based on their network of RNA-protein interactions. In addition to capturing established RNA classes based on their biochemical profiles, the Grad-seq approach enabled the discovery of an overlooked large collective of structured small RNAs that form stable complexes with the conserved protein ProQ. We show that ProQ is an abundant RNA-binding protein with a wide range of ligands and a global influence on Salmonella gene expression. Given its generic ability to chart a functional RNA landscape irrespective of transcript length and sequence diversity, Grad-seq promises to define functional RNA classes and major RNA-binding proteins in both model species and genetically intractable organisms.
    Keywords: Hfq ; Proq ; RNA–Protein Interaction ; Noncoding RNA ; Small RNA ; Bacterial Proteins -- Metabolism ; High-Throughput Nucleotide Sequencing -- Methods ; RNA, Bacterial -- Metabolism ; RNA-Binding Proteins -- Metabolism ; Salmonella Enterica -- Metabolism
    ISSN: 00278424
    E-ISSN: 1091-6490
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  • 7
    In: EMBO Journal, 03 June 2015, Vol.34(11), pp.1478-1492
    Description: There is an expanding list of examples by which one can posttranscriptionally influence the expression of others. This can involve sponges that sequester regulatory s of s in the same regulon, but the underlying molecular mechanism of such cross talk remains little understood. Here, we report sponge‐mediated cross talk in the posttranscriptional network of GcvB, a conserved Hfq‐dependent small with one of the largest regulons known in bacteria. We show that decay from the locus encoding an amino acid transporter generates a stable fragment (SroC) that base‐pairs with GcvB. This interaction triggers the degradation of GcvB by ase E, alleviating the GcvB‐mediated repression of other amino acid‐related transport and metabolic genes. Intriguingly, since the itself is a target of GcvB, the SroC sponge seems to enable both an internal feed‐forward loop to activate its parental in and activation of many ‐encoded s in the same pathway. Disabling this cross talk affects bacterial growth when peptides are the sole carbon and nitrogen sources. Decay of the bacterial GcvB , which keeps it from regulating its targets, is triggered by a 3′‐‐derived fragment from a target . This ability of s to compete for regulatory interaction presents a new mode of cross talk in bacteria. . Decay of the bacterial GcvB s, which keeps it from regulating its m targets, is triggered by a 3′‐‐derived fragment from a target m. This ability of ms to compete for regulatory interaction presents a new mode of cross talk in bacteria.
    Keywords: G Cv B ; H Fq ; Noncoding Rna ; Rn Ase E ; S Ro C
    ISSN: 0261-4189
    E-ISSN: 1460-2075
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  • 8
    Language: English
    In: Nature, 28 January 2016, Vol.529(7587), pp.496-501
    Description: Bacteria express many small RNAs for which the regulatory roles in pathogenesis have remained poorly understood due to a paucity of robust phenotypes in standard virulence assays. Here we use a generic 'dual RNA-seq' approach to profile RNA expression simultaneously in pathogen and host during Salmonella enterica serovar Typhimurium infection and reveal the molecular impact of bacterial riboregulators. We identify a PhoP-activated small RNA, PinT, which upon bacterial internalization temporally controls the expression of both invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity causes pervasive changes in coding and noncoding transcripts of the host. Interspecies correlation analysis links PinT to host cell JAK-STAT signalling, and we identify infection-specific alterations in multiple long noncoding RNAs. Our study provides a paradigm for a sensitive RNA-based analysis of intracellular bacterial pathogens and their hosts without physical separation, as well as a new discovery route for hidden functions of pathogen genes.
    Keywords: Gene Expression Regulation -- Genetics ; Host-Pathogen Interactions -- Genetics ; RNA, Bacterial -- Genetics ; RNA, Untranslated -- Genetics ; Salmonella Typhimurium -- Genetics
    ISSN: 00280836
    E-ISSN: 1476-4687
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  • 9
    Language: English
    In: Nucleic acids research, 02 June 2017, Vol.45(10), pp.6147-6167
    Description: Neisseria meningitidis is a human commensal that can also cause life-threatening meningitis and septicemia. Despite growing evidence for RNA-based regulation in meningococci, their transcriptome structure and output of regulatory small RNAs (sRNAs) are incompletely understood. Using dRNA-seq, we have mapped at single-nucleotide resolution the primary transcriptome of N. meningitidis strain 8013. Annotation of 1625 transcriptional start sites defines transcription units for most protein-coding genes but also reveals a paucity of classical σ70-type promoters, suggesting the existence of activators that compensate for the lack of -35 consensus sequences in N. meningitidis. The transcriptome maps also reveal 65 candidate sRNAs, a third of which were validated by northern blot analysis. Immunoprecipitation with the RNA chaperone Hfq drafts an unexpectedly large post-transcriptional regulatory network in this organism, comprising 23 sRNAs and hundreds of potential mRNA targets. Based on this data, using a newly developed gfp reporter system we validate an Hfq-dependent mRNA repression of the putative colonization factor PrpB by the two trans-acting sRNAs RcoF1/2. Our genome-wide RNA compendium will allow for a better understanding of meningococcal transcriptome organization and riboregulation with implications for colonization of the human nasopharynx.
    Keywords: Gene Expression Regulation, Bacterial ; Transcriptome ; Bacterial Proteins -- Metabolism ; Host Factor 1 Protein -- Metabolism ; Micrornas -- Genetics ; Molecular Chaperones -- Metabolism ; Neisseria Meningitidis -- Genetics ; RNA, Bacterial -- Genetics ; RNA, Messenger -- Genetics
    ISSN: 03051048
    E-ISSN: 1362-4962
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  • 10
    Language: English
    In: Science (New York, N.Y.), 30 May 2014, Vol.344(6187), pp.972-3
    Description: Argonaute proteins have emerged as the key effectors in virtually all eukaryotic small RNA-mediated gene silencing pathways . Central to all their activities is their association with the small guide RNAs that allow them to recognize through sequence complementarity, and in some cases also cleave, cellular transcripts. [PUBLICATION ]
    Keywords: DNA Cleavage ; Gene Silencing ; Argonaute Proteins -- Metabolism ; DNA -- Metabolism ; DNA, Bacterial -- Genetics ; Plasmids -- Genetics ; Prokaryotic Cells -- Metabolism ; Rhodobacter Sphaeroides -- Genetics ; Thermus Thermophilus -- Genetics
    ISSN: 00368075
    E-ISSN: 1095-9203
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