The Experts below are selected from a list of 288 Experts worldwide ranked by ideXlab platform
Dieter Haas - One of the best experts on this subject based on the ideXlab platform.
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gac rsm signal transduction pathway of γ proteobacteria from rna recognition to regulation of social behaviour
Molecular Microbiology, 2007Co-Authors: Karine Lapouge, Mario Schubert, Frederic H T Allain, Dieter HaasAbstract:Summary In many g-proteobacteria, the conserved GacS/GacA (BarA/UvrY) two-component system positively con- trols the expression of one to five genes specifying small RNAs (sRNAs) that are characterized by repeated unpaired GGA motifs but otherwise appear to belong to several independent families. The GGA motifs are essential for binding small, dimeric RNA- binding proteins of a single conserved family desig- nated RsmA (CsrA). These proteins, which also occur in bacterial species outside the g-proteobacteria, act as translational repressors of certain mRNAs when these contain an RsmA/CsrA binding site at or near the Shine-Dalgarno Sequence plus additional binding sites located in the 5 untranslated leader mRNA. Recent structural data have established that the RsmA-like protein RsmE of Pseudomonas fluore- scens makes specific contacts with an RNA consen- sus Sequence 5- A /UCANGGANG U /A-3 (where N is any nucleotide). Interaction with an RsmA/CsrA protein promotes the formation of a short stem supporting an ANGGAN loop. This conformation hinders access of 30S ribosomal subunits and hence translation initiation. The output of the Gac/Rsm cascade varies widely in different bacterial species and typically involves management of carbon storage and expres- sion of virulence or biocontrol factors. Unidentified signal molecules co-ordinate the activity of the Gac/ Rsm cascade in a cell population density-dependent manner.
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mechanism of hcna mrna recognition in the gac rsm signal transduction pathway of pseudomonas fluorescens
Molecular Microbiology, 2007Co-Authors: Karine Lapouge, Elena V Sineva, Magnus Lindell, Katja Starke, Carol S Baker, Paul Babitzke, Dieter HaasAbstract:In the plant-beneficial bacterium Pseudomonas fluorescens CHA0, the expression of antifungal exoproducts is controlled by the GacS/GacA two-component system. Two RNA binding proteins (RsmA, RsmE) ensure effective translational repression of exoproduct mRNAs. At high cell population densities, GacA induces three small RNAs (RsmX, RsmY, RsmZ) which sequester both RsmA and RsmE, thereby relieving translational repression. Here we systematically analyse the features that allow the RNA binding proteins to interact strongly with the 5' untranslated leader mRNA of the P. fluorescens hcnA gene (encoding hydrogen cyanide synthase subunit A). We obtained evidence for three major RsmA/RsmE recognition elements in the hcnA leader, based on directed mutagenesis, RsmE footprints and toeprints, and in vivo expression data. Two recognition elements were found in two stem-loop structures whose existence in the 5' leader region was confirmed by lead(II) cleavage analysis. The third recognition element, which overlapped the hcnA Shine-Dalgarno Sequence, was postulated to adopt either an open conformation, which would favour ribosome binding, or a stem-loop structure, which may form upon interaction with RsmA/RsmE and would inhibit access of ribosomes. Effective control of hcnA expression by the Gac/Rsm system appears to result from the combination of the three appropriately spaced recognition elements.
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molecular basis of messenger rna recognition by the specific bacterial repressing clamp rsma csra
Nature Structural & Molecular Biology, 2007Co-Authors: Mario Schubert, Karine Lapouge, Olivier Duss, Florian C Oberstrass, Ilian Jelesarov, Dieter Haas, Frederic H T AllainAbstract:Proteins of the RsmA/CsrA family are global translational regulators in many bacterial species. We have determined the solution structure of a complex formed between the RsmE protein, a member of this family from Pseudomonas fluorescens, and a target RNA encompassing the ribosome-binding site of the hcnA gene. The RsmE homodimer with its two RNA-binding sites makes optimal contact with an 5'-A/UCANGGANGU/A-3' Sequence in the mRNA. When tightly gripped by RsmE, the ANGGAN core folds into a loop, favoring the formation of a 3-base-pair stem by flanking nucleotides. We validated these findings by in vivo and in vitro mutational analyses. The structure of the complex explains well how, by sequestering the Shine-Dalgarno Sequence, the RsmA/CsrA proteins repress translation.
Jonathan S Weissman - One of the best experts on this subject based on the ideXlab platform.
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the coding and noncoding architecture of the caulobacter crescentus genome
PLOS Genetics, 2014Co-Authors: Jared M Schrader, Sean Crosson, Harley H Mcadams, Keren Lasker, Bo Zhou, Seth W Childers, Brandon Williams, Tao Long, Jonathan S Weissman, Lucy ShapiroAbstract:Caulobacter crescentus undergoes an asymmetric cell division controlled by a genetic circuit that cycles in space and time. We provide a universal strategy for defining the coding potential of bacterial genomes by applying ribosome profiling, RNA-seq, global 5′-RACE, and liquid chromatography coupled with tandem mass spectrometry (LC-MS) data to the 4-megabase C. crescentus genome. We mapped transcript units at single base-pair resolution using RNA-seq together with global 5′-RACE. Additionally, using ribosome profiling and LC-MS, we mapped translation start sites and coding regions with near complete coverage. We found most start codons lacked corresponding Shine-Dalgarno sites although ribosomes were observed to pause at internal Shine-Dalgarno sites within the coding DNA Sequence (CDS). These data suggest a more prevalent use of the Shine-Dalgarno Sequence for ribosome pausing rather than translation initiation in C. crescentus. Overall 19% of the transcribed and translated genomic elements were newly identified or significantly improved by this approach, providing a valuable genomic resource to elucidate the complete C. crescentus genetic circuitry that controls asymmetric cell division.
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the anti shine dalgarno Sequence drives translational pausing and codon choice in bacteria
Nature, 2012Co-Authors: Jonathan S WeissmanAbstract:Protein synthesis by ribosomes takes place on a linear substrate but at non-uniform speeds. Transient pausing of ribosomes can affect a variety of co-translational processes, including protein targeting and folding. These pauses are influenced by the Sequence of the messenger RNA. Thus, redundancy in the genetic code allows the same protein to be translated at different rates. However, our knowledge of both the position and the mechanism of translational pausing in vivo is highly limited. Here we present a genome-wide analysis of translational pausing in bacteria by ribosome profiling--deep sequencing of ribosome-protected mRNA fragments. This approach enables the high-resolution measurement of ribosome density profiles along most transcripts at unperturbed, endogenous expression levels. Unexpectedly, we found that codons decoded by rare transfer RNAs do not lead to slow translation under nutrient-rich conditions. Instead, Shine-Dalgarno-(SD)-like features within coding Sequences cause pervasive translational pausing. Using an orthogonal ribosome possessing an altered anti-SD Sequence, we show that pausing is due to hybridization between the mRNA and 16S ribosomal RNA of the translating ribosome. In protein-coding Sequences, internal SD Sequences are disfavoured, which leads to biased usage, avoiding codons and codon pairs that resemble canonical SD sites. Our results indicate that internal SD-like Sequences are a major determinant of translation rates and a global driving force for the coding of bacterial genomes.
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the anti shine dalgarno Sequence drives translational pausing and codon choice in bacteria
Nature, 2012Co-Authors: Genewei Li, Eugene Oh, Jonathan S WeissmanAbstract:Internal Shine–Dalgarno-like Sequences in bacterial messenger RNA determine the elongation rate of protein synthesis and synonymous codon usage.
Karine Lapouge - One of the best experts on this subject based on the ideXlab platform.
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gac rsm signal transduction pathway of γ proteobacteria from rna recognition to regulation of social behaviour
Molecular Microbiology, 2007Co-Authors: Karine Lapouge, Mario Schubert, Frederic H T Allain, Dieter HaasAbstract:Summary In many g-proteobacteria, the conserved GacS/GacA (BarA/UvrY) two-component system positively con- trols the expression of one to five genes specifying small RNAs (sRNAs) that are characterized by repeated unpaired GGA motifs but otherwise appear to belong to several independent families. The GGA motifs are essential for binding small, dimeric RNA- binding proteins of a single conserved family desig- nated RsmA (CsrA). These proteins, which also occur in bacterial species outside the g-proteobacteria, act as translational repressors of certain mRNAs when these contain an RsmA/CsrA binding site at or near the Shine-Dalgarno Sequence plus additional binding sites located in the 5 untranslated leader mRNA. Recent structural data have established that the RsmA-like protein RsmE of Pseudomonas fluore- scens makes specific contacts with an RNA consen- sus Sequence 5- A /UCANGGANG U /A-3 (where N is any nucleotide). Interaction with an RsmA/CsrA protein promotes the formation of a short stem supporting an ANGGAN loop. This conformation hinders access of 30S ribosomal subunits and hence translation initiation. The output of the Gac/Rsm cascade varies widely in different bacterial species and typically involves management of carbon storage and expres- sion of virulence or biocontrol factors. Unidentified signal molecules co-ordinate the activity of the Gac/ Rsm cascade in a cell population density-dependent manner.
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mechanism of hcna mrna recognition in the gac rsm signal transduction pathway of pseudomonas fluorescens
Molecular Microbiology, 2007Co-Authors: Karine Lapouge, Elena V Sineva, Magnus Lindell, Katja Starke, Carol S Baker, Paul Babitzke, Dieter HaasAbstract:In the plant-beneficial bacterium Pseudomonas fluorescens CHA0, the expression of antifungal exoproducts is controlled by the GacS/GacA two-component system. Two RNA binding proteins (RsmA, RsmE) ensure effective translational repression of exoproduct mRNAs. At high cell population densities, GacA induces three small RNAs (RsmX, RsmY, RsmZ) which sequester both RsmA and RsmE, thereby relieving translational repression. Here we systematically analyse the features that allow the RNA binding proteins to interact strongly with the 5' untranslated leader mRNA of the P. fluorescens hcnA gene (encoding hydrogen cyanide synthase subunit A). We obtained evidence for three major RsmA/RsmE recognition elements in the hcnA leader, based on directed mutagenesis, RsmE footprints and toeprints, and in vivo expression data. Two recognition elements were found in two stem-loop structures whose existence in the 5' leader region was confirmed by lead(II) cleavage analysis. The third recognition element, which overlapped the hcnA Shine-Dalgarno Sequence, was postulated to adopt either an open conformation, which would favour ribosome binding, or a stem-loop structure, which may form upon interaction with RsmA/RsmE and would inhibit access of ribosomes. Effective control of hcnA expression by the Gac/Rsm system appears to result from the combination of the three appropriately spaced recognition elements.
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molecular basis of messenger rna recognition by the specific bacterial repressing clamp rsma csra
Nature Structural & Molecular Biology, 2007Co-Authors: Mario Schubert, Karine Lapouge, Olivier Duss, Florian C Oberstrass, Ilian Jelesarov, Dieter Haas, Frederic H T AllainAbstract:Proteins of the RsmA/CsrA family are global translational regulators in many bacterial species. We have determined the solution structure of a complex formed between the RsmE protein, a member of this family from Pseudomonas fluorescens, and a target RNA encompassing the ribosome-binding site of the hcnA gene. The RsmE homodimer with its two RNA-binding sites makes optimal contact with an 5'-A/UCANGGANGU/A-3' Sequence in the mRNA. When tightly gripped by RsmE, the ANGGAN core folds into a loop, favoring the formation of a 3-base-pair stem by flanking nucleotides. We validated these findings by in vivo and in vitro mutational analyses. The structure of the complex explains well how, by sequestering the Shine-Dalgarno Sequence, the RsmA/CsrA proteins repress translation.
Franz Narberhaus - One of the best experts on this subject based on the ideXlab platform.
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rna mediated thermoregulation of iron acquisition genes in shigella dysenteriae and pathogenic escherichia coli
PLOS ONE, 2013Co-Authors: Andrew B Kouse, Franz Narberhaus, Francesco Righetti, Jens Kortmann, Erin R MurphyAbstract:The initiation, progression and transmission of most bacterial infections is dependent upon the ability of the invading pathogen to acquire iron from each of the varied environments encountered during the course of a natural infection. In total, 95% of iron within the human body is complexed within heme, making heme a potentially rich source of host-associated nutrient iron for invading bacteria. As heme is encountered only within the host, pathogenic bacteria often regulate synthesis of heme utilization factors such that production is maximal under host-associated environmental conditions. This study examines the regulated production of ShuA, an outer-membrane receptor required for the utilization of heme as a source of nutrient iron by Shigella dysenteriae, a pathogenic bacterium that causes severe diarrheal diseases in humans. Specifically, the impact of the distinct environmental temperatures encountered during infection within a host (37°C) and transmission between hosts (25°C) on shuA expression is investigated. We show that shuA expression is subject to temperature-dependent post-transcriptional regulation resulting in increased ShuA production at 37°C. The observed thermoregulation is mediated by nucleic acid Sequences within the 5′ untranslated region. In addition, we have identified similar nucleotide Sequences within the 5′ untranslated region of the orthologous chuA transcript of enteropathogenic E. coli and have demonstrated that it also functions to confer temperature-dependent post-transcriptional regulation. In both function and predicted structure, the regulatory element within the shuA and chuA 5′ untranslated regions closely resembles a FourU RNA thermometer, a zipper-like RNA structure that occludes the Shine-Dalgarno Sequence at low temperatures. Increased production of ShuA and ChuA in response to the host body temperature allows for maximal production of these heme acquisition factors within the environment where S. dysenteriae and pathogenic E. coli strains would encounter heme, a host-specific iron source.
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direct observation of the temperature induced melting process of the salmonella fouru rna thermometer at base pair resolution
Nucleic Acids Research, 2010Co-Authors: Jorg Rinnenthal, Franz Narberhaus, Birgit Klinkert, Harald SchwalbeAbstract:In prokaryotes, RNA thermometers regulate a number of heat shock and virulence genes. These temperature sensitive RNA elements are usually located in the 5′-untranslated regions of the regulated genes. They repress translation initiation by base pairing to the Shine–Dalgarno Sequence at low temperatures. We investigated the thermodynamic stability of the temperature labile hairpin 2 of the Salmonella fourU RNA thermometer over a broad temperature range and determined free energy, enthalpy and entropy values for the base-pair opening of individual nucleobases by measuring the temperature dependence of the imino proton exchange rates via NMR spectroscopy. Exchange rates were analyzed for the wild-type (wt) RNA and the A8C mutant. The wt RNA was found to be stabilized by the extraordinarily stable G14–C25 base pair. The mismatch base pair in the wt RNA thermometer (A8–G31) is responsible for the smaller cooperativity of the unfolding transition in the wt RNA. Enthalpy and entropy values for the base-pair opening events exhibit linear correlation for both RNAs. The slopes of these correlations coincide with the melting points of the RNAs determined by CD spectroscopy. RNA unfolding occurs at a temperature where all nucleobases have equal thermodynamic stabilities. Our results are in agreement with a consecutive zipper-type unfolding mechanism in which the stacking interaction is responsible for the observed cooperativity. Furthermore, remote effects of the A8C mutation affecting the stability of nucleobase G14 could be identified. According to our analysis we deduce that this effect is most probably transduced via the hydration shell of the RNA.
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rna thermometers are common in α and γ proteobacteria
Biological Chemistry, 2005Co-Authors: Torsten Waldminghaus, Anja Fippinger, Juliane Alfsmann, Franz NarberhausAbstract:: Expression of many rhizobial small heat-shock genes is controlled by the ROSE element, a thermoresponsive structure in the 5'-untranslated region of the corresponding mRNAs. Using a bioinformatics approach, we found more than 20 new potential ROSE-like RNA thermometers upstream of small heat-shock genes in a wide variety of alpha- and gamma-proteobacteria. Northern blot analyses revealed heat-inducible transcripts of the representative candidate Caulobacter crescentus CC2258, Escherichia coli ibpA and Salmonella typhimurium ibpA genes. Typical sigma(32)-type promoters were mapped upstream of the potential RNA thermometers by primer extension. Additional translational control was demonstrated in a lacZ reporter system and by site-directed mutagenesis. RNA secondary structure predictions strongly suggest that the Shine-Dalgarno Sequence in the RNA thermometers is masked at low temperatures. Combining two regulatory modules, a sigma(32) promoter and a ROSE-type RNA thermometer, provides a novel stringent mechanism to control expression of small heat-shock genes.
Masahiro Sugiura - One of the best experts on this subject based on the ideXlab platform.
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translation initiation of cyanobacterial rbcs mrnas requires the 38 kda ribosomal protein s1 but not the shine dalgarno Sequence development of a cyanobacterial in vitro translation system
Journal of Biological Chemistry, 2006Co-Authors: Michinori Mutsuda, Masahiro SugiuraAbstract:Little is known about the biochemical mechanism of translation in cyanobacteria though substantial studies have been made on photosynthesis, nitrogen fixation, circadian rhythm, and genome structure. To analyze the mechanism of cyanobacterial translation, we have developed an in vitro translation system from Synechococcus cells using a psbAI-lacZ fusion mRNA as a model template. This in vitro system supports accurate translation from the authentic initiation site of a variety of Synechococcus mRNAs. In Synechococcus cells, rbcL and rbcS encoding the large and small subunits, respectively, of ribulose-1,5-bisphosphate carboxylase/oxygenase are co-transcribed as a dicistronic mRNA, and the downstream rbcS mRNA possesses two possible initiation codons separated by three nucleotides. Using this in vitro system and mutated mRNAs, we demonstrated that translation starts exclusively from the upstream AUG codon. Although there are Shine-Dalgarno-like Sequences in positions similar to those of the functional Shine-Dalgarno elements in Escherichia coli, mutation analysis indicated that these Sequences are not required for translation. Assays with deletions within the 5'-untranslated region showed that a pyrimidine-rich Sequence in the -46 to -15 region is necessary for efficient translation. Synechococcus cells contain two ribosomal protein S1 homologues of 38 and 33 kDa in size. UV cross-linking and immunoprecipitation experiments suggested that the 38-kDa S1 is involved in efficient translation via associating with the pyrimidine-rich Sequence. The present in vitro translation system will be a powerful tool to analyze the basic mechanism of translation in cyanobacteria.
