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At least 55 records · Page 3Linked to original sources
PubMed2006

Bacterial RNA and small antiviral compounds activate caspase-1 through cryopyrin/Nalp3.

Missense mutations in the CIAS1 gene cause three autoinflammatory disorders: familial cold autoinflammatory syndrome, Muckle-Wells syndrome and neonatal-onset multiple-system inflammatory disease. Cryopyrin (also called Nalp3), the product of CIAS1, is a member of the NOD-LRR protein family that has been linked to the activation of intracellular host defence signalling pathways. Cryopyrin forms a multi-protein complex termed 'the inflammasome', which contains the apoptosis-associated speck-like protein (ASC) and caspase-1, and promotes caspase-1 activation and processing of pro-interleukin (IL)-1beta (ref. 4). Here we show the effect of cryopyrin deficiency on inflammasome function and immune responses. Cryopyrin and ASC are essential for caspase-1 activation and IL-1beta and IL-18 production in response to bacterial RNA and the imidazoquinoline compounds R837 and R848. In contrast, secretion of tumour-necrosis factor-alpha and IL-6, as well as activation of NF-kappaB and mitogen-activated protein kinases (MAPKs) were unaffected by cryopyrin deficiency. Furthermore, we show that Toll-like receptors and cryopyrin control the secretion of IL-1beta and IL-18 through different intracellular pathways. These results reveal a critical role for cryopyrin in host defence through bacterial RNA-mediated activation of caspase-1, and provide insights regarding the pathogenesis of autoinflammatory syndromes.

Adaptor Proteins, Signal Transducing↗
PubMed1998

A bacterial RNA that functions as both a tRNA and an mRNA.

Bacterial tmRNA (transfer-messenger RNA, also known as 10Sa RNA) contains a tRNA-like structure in the 5'- and 3'-end sequences and an internal reading frame encoding a 'tag' peptide. The dual function of this molecule as both a tRNA and an mRNA facilitates a trans-translation reaction, in which a ribosome can switch between translation of a truncated mRNA and the tmRNA's tag sequence. The result is a chimeric protein with the tag peptide attached to the C-terminus of the truncated peptide.

Base Sequence↗
PubMed1997

Leaf-specifically expressed genes for polypeptides destined for chloroplasts with domains of sigma70 factors of bacterial RNA polymerases in Arabidopsis thaliana.

Genes for sigma-like factors of bacterial-type RNA polymerase have not been characterized from any multicellular eukaryotes, although they probably play a crucial role in the expression of plastid photosynthesis genes. We have cloned three distinct cDNAs, designated SIG1, SIG2, and SIG3, for polypeptides possessing amino acid sequences for domains conserved in sigma70 factors of bacterial RNA polymerases from the higher plant Arabidopsis thaliana. Each gene is present as one copy per haploid genome without any additional sequences hybridized in the genome. Transient expression assays using green fluorescent protein demonstrated that N-terminal regions of the SIG2 and SIG3 ORFs could function as transit peptides for import into chloroplasts. Transcripts for all three SIG genes were detected in leaves but not in roots, and were induced in leaves of dark-adapted plants in rapid response to light illumination. Together with results of our previous analysis of tissue-specific regulation of transcription of plastid photosynthesis genes, these results indicate that expressed levels of the genes may influence transcription by regulating RNA polymerase activity in a green tissue-specific manner.

Amino Acid Sequence↗
PubMed2001

Structural mechanism for rifampicin inhibition of bacterial rna polymerase.

Rifampicin (Rif) is one of the most potent and broad spectrum antibiotics against bacterial pathogens and is a key component of anti-tuberculosis therapy, stemming from its inhibition of the bacterial RNA polymerase (RNAP). We determined the crystal structure of Thermus aquaticus core RNAP complexed with Rif. The inhibitor binds in a pocket of the RNAP beta subunit deep within the DNA/RNA channel, but more than 12 A away from the active site. The structure, combined with biochemical results, explains the effects of Rif on RNAP function and indicates that the inhibitor acts by directly blocking the path of the elongating RNA when the transcript becomes 2 to 3 nt in length.

Amino Acid Sequence↗
PubMed2003

Bacterial RNA polymerases: the wholo story.

Recent structural and biophysical results have provided unprecedented insights into the structure and function of the bacterial RNA polymerase holoenzyme as it goes through the steps of transcription initiation. Comparisons with structural analyses of evolutionarily unrelated RNA polymerases reveal unexpected general features of the initiation process.

Bacteria↗
PubMed2004

Mode of action of the microbial metabolite GE23077, a novel potent and selective inhibitor of bacterial RNA polymerase.

GE23077, a novel microbial metabolite recently isolated from Actinomadura sp. culture media, is a potent and selective inhibitor of bacterial RNA polymerase (RNAP). It inhibits Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) RNAPs with IC50 values (i.e. the concentration at which the enzyme activity is inhibited by 50%) in the 10(-8) m range, whereas it is not active on E. coli DNA polymerase or on eukaryotic (wheat germ) RNAP II (IC50 values > 10(-4) m in both cases). In spite of its potent activity on purified bacterial RNAPs, GE23077 shows a narrow spectrum of antimicrobial activity on Gram-positive and Gram-negative bacteria. To investigate the molecular basis of this behaviour, the effects of GE23077 on macromolecular biosynthesis were tested in E. coli cells permeabilized under different conditions. The addition of GE23077 to plasmolyzed cells resulted in an immediate and specific inhibition of intracellular RNA biosynthesis, in a dose-response manner, strongly suggesting that cell penetration is the main obstacle for effective antimicrobial activity of the antibiotic. Biochemical studies were also conducted with purified enzymes to obtain further insights into the mode of action of GE23077. Interestingly, the compound displays a behaviour similar to that of rifampicin, an antibiotic structurally unrelated to GE23077: both compounds act at the level of transcription initiation, but not on the sigma subunit and not on the formation of the promoter DNA-RNAP complex. Tests on different rifampicin-resistant E. coli RNAPs did not show any cross-resistance between the two compounds, indicating distinct binding sites on the target enzyme. In conclusion, GE23077 is an interesting new molecule for future mechanistic studies on bacterial RNAP and for its potential in anti-infective drug discovery.

Actinomycetales↗
PubMed2002

Crystal structure of a bacterial RNA polymerase holoenzyme at 2.6 A resolution.

In bacteria, the binding of a single protein, the initiation factor sigma, to a multi-subunit RNA polymerase core enzyme results in the formation of a holoenzyme, the active form of RNA polymerase essential for transcription initiation. Here we report the crystal structure of a bacterial RNA polymerase holoenzyme from Thermus thermophilus at 2.6 A resolution. In the structure, two amino-terminal domains of the sigma subunit form a V-shaped structure near the opening of the upstream DNA-binding channel of the active site cleft. The carboxy-terminal domain of sigma is near the outlet of the RNA-exit channel, about 57 A from the N-terminal domains. The extended linker domain forms a hairpin protruding into the active site cleft, then stretching through the RNA-exit channel to connect the N- and C-terminal domains. The holoenzyme structure provides insight into the structural organization of transcription intermediate complexes and into the mechanism of transcription initiation.

Amino Acid Sequence↗
PubMed2005

Inhibition of bacterial RNA polymerase by streptolydigin: stabilization of a straight-bridge-helix active-center conformation.

We define the target, mechanism, and structural basis of inhibition of bacterial RNA polymerase (RNAP) by the tetramic acid antibiotic streptolydigin (Stl). Stl binds to a site adjacent to but not overlapping the RNAP active center and stabilizes an RNAP-active-center conformational state with a straight-bridge helix. The results provide direct support for the proposals that alternative straight-bridge-helix and bent-bridge-helix RNAP-active-center conformations exist and that cycling between straight-bridge-helix and bent-bridge-helix RNAP-active-center conformations is required for RNAP function. The results set bounds on models for RNAP function and suggest strategies for design of novel antibacterial agents.

Aminoglycosides↗
PubMed2002

The downstream DNA jaw of bacterial RNA polymerase facilitates both transcriptional initiation and pausing.

Regulation of RNA polymerase during initiation, elongation, and termination of transcription is mediated in part by interactions with intrinsic regulatory signals encoded in the RNA and DNA that contact the enzyme. These interactions include contacts to an 8-9-bp RNA:DNA hybrid within the active-site cleft of the enzyme, contacts to the melted nontemplate DNA strand in the vicinity of the hybrid, contacts to exiting RNA upstream of the hybrid, and contacts to approximately 20 bp of duplex DNA downstream of the active site. Based on characterization of an amino acid substitution (G1161R) and a deletion (Delta1149-1190) in the jaw domain of the bacterial RNA polymerase largest subunit (beta'), we report here that contacts of the jaw domain to downstream DNA at the leading edge of the transcription complex contribute to regulation during all three phases of transcription. The results provide insight into the role of the jaw domain-downstream DNA contact in transcriptional initiation and pausing and suggest possible explanations for the previously reported isolation of the jaw mutants based on reduced ColEI plasmid replication.

Amino Acid Sequence↗
PubMed2001

Bacterial RNA polymerase.

The recently determined crystal structure of a bacterial core RNA polymerase (RNAP) provides the first glimpse of this family of evolutionarily conserved cellular RNAPs. Using the structure as a framework, a consistent picture of protein-nucleic acid interactions in transcription complexes has been accumulated from cross-linking experiments. The molecule can be viewed as a molecular machine, with distinct structural features hypothesized to perform specific functions. Comparison with the alpha-carbon backbone of a eukaryotic RNAP reveals close structural similarity.

Archaeal Proteins↗
PubMed1983

The myxopyronins, new inhibitors of bacterial RNA synthesis from Myxococcus fulvus (Myxobacterales).

From the culture supernatant of the myxobacterium, Myxococcus fulvus strain Mx f50, an antibiotic activity was isolated which blocked growth of many Gram-positive and several Gram-negative bacteria, but not of yeasts and fungi. The activity consisted of two closely related compounds, myxopyronins A and B. The myxopyronins appear to be new antibiotics, and seem to specifically inhibit bacterial RNA polymerase.

Animals↗
PubMed1975

The 5'-termini of bacterial RNA. I. The monophosphorylated termini.

A two-dimensional fingerprint technique is described which separated oligonucleotides by length and number of phosphates present at the termini. The diversity of terminal nucleotide sequences was found to be quite restricted in bacterial 4-5 S RNA: in the strain used here the predominant dinucleotide species was pG-C, and the only trinucleotides produced were pG-G-Up and pG-G-Cp. Examination of total pulse-labeled RNA revealed the same restricted set of terminal trinucleotides.

Animals↗
PubMed2005

Structure and function of lineage-specific sequence insertions in the bacterial RNA polymerase beta' subunit.

The large beta and beta' subunits of the bacterial core RNA polymerase (RNAP) are highly conserved throughout evolution. Nevertheless, large sequence insertions in beta and beta' characterize specific evolutionary lineages of bacteria. The Thermus aquaticus RNAP beta' subunit contains a 283 residue insert between conserved regions A and B that is found in only four bacterial species. The Escherichia coli RNAP beta' subunit contains a 188 residue insert in the middle of conserved region G that is found in a wide range of bacterial species. Here, we present structural studies of these two beta' insertions. We show that the inserts comprise repeats of a previously characterized fold, the sandwich-barrel hybrid motif (as predicted from previous sequence analysis) and that the inserts serve significant roles in facilitating protein/protein and/or protein/nucleic acid interactions.

Amino Acid Sequence↗
PubMed1985

The corallopyronins, new inhibitors of bacterial RNA synthesis from Myxobacteria.

From the culture broth of the myxobacterium, Corallococcus (Myxococcus) coralloides, three new antibiotics have been isolated: corallopyronin A, B and C. The compounds, which are chemically related to the recently discovered myxopyronins, act mainly on Gram-positive bacteria, with MIC values between 0.1 and 10 micrograms/ml, and only exceptionally or at much higher concentrations (MIC values; 100 and more micrograms/ml) on Gram-negatives. They do not inhibit eukaryotic organisms and show no toxicity for mice (sc). The corallopyronins appear to block specifically eubacterial RNA polymerase.

Animals↗
PubMed2000

Selective amplification of bacterial RNA: use of a DNA primer containing mismatched bases near its 3' terminus to reduce false-positive signals.

A reverse transcription PCR (RT-PCR) method designed to reduce false-positive results due to the co-amplification of contaminating genomic DNA is reported. Feasibility of the method was evaluated using 16S rRNA sequences specific to Bacillus cereus. A DNA oligonucleotide primer, consisting of 22-bases containing three consecutive mismatched bases near its 3' terminus (primer B16RT), was used for reverse transcription and in subsequent cDNA amplification. Specific rRNA was reverse transcribed at low temperature (40 degrees C or 45 degrees C) in the presence of primer B16RT. As subsequent PCR using primer B16RT at high (62 degrees C) annealing temperatures is not nearly as efficient as amplification using the specific primer, amplification of genomic DNA was hindered relative to the amplification of cDNA. The method was readily adapted to the selective amplification of mRNA of the Listeria monocytogenes listeriolysin O (hly) gene.

3' Untranslated Regions↗
PubMed2004

A small bacterial RNA regulates a putative ABC transporter.

A small noncoding bacterial ribonucleic acid of 62-64 nucleotides, RydC, was identified in the genomes of Escherichia coli, Salmonella, and Shigella. In vivo, RydC binds to the RNA-binding protein Hfq, and it is unstable when Hfq is absent. Mobility assays reveal that complex formation between RydC and Hfq is specific, with an apparent binding constant of approximately 300 nm. Sequence alignments combined with structural probing demonstrate that RydC folds as a pseudoknot. Hfq binds the loops crossing the deep and shallow grooves of the pseudoknotted RNA and reorganizes its overall conformation. An interaction with a polycistronic mRNA, yejABEF, which encodes a putative ABC transporter, was detected by affinity purification of immobilized RNA-Hfq complexes. In vivo, the yejABEF operon is expressed on minimal medium. Remarkably, its expression is reduced when RydC is absent, and the operon is degraded when RydC expression is stimulated. This observation correlates with the growth defects associated with a stimulation of its expression in vivo, generating a thermosensitive phenotype that affects growth on minimal media supplemented with glycerol, maltose, or ribose. We conclude that RydC regulates the yejABEF-encoded ABC permease at the mRNA level. This small RNA may contribute to optimal adaptation of some Enterobacteria to environmental conditions.

ATP-Binding Cassette Transporters↗