Signal Transduction Pathways in the Pentameric Ligand-Gated Ion Channels

The mechanisms of allosteric action within pentameric ligand-gated ion channels (pLGICs) remain to be determined. Using crystallography, site-directed mutagenesis, and two-electrode voltage clamp measurements, we identified two functionally relevant sites in the extracellular (EC) domain of the bacterial pLGIC from Gloeobacter violaceus (GLIC). One site is at the C-loop region, where the NQN mutation (D91N, E177Q, and D178N) eliminated inter-subunit salt bridges in the open-channel GLIC structure and thereby shifted the channel activation to a higher agonist concentration. The other site is below the C-loop, where binding of the anesthetic ketamine inhibited GLIC currents in a concentration dependent manner. To understand how a perturbation signal in the EC domain, either resulting from the NQN mutation or ketamine binding, is transduced to the channel gate, we have used the Perturbation-based Markovian Transmission (PMT) model to determine dynamic responses of the GLIC channel and signaling pathways upon initial perturbations in the EC domain of GLIC. Despite the existence of many possible routes for the initial perturbation signal to reach the channel gate, the PMT model in combination with Yen's algorithm revealed that perturbation signals with the highest probability flow travel either via the β1-β2 loop or through pre-TM1. The β1-β2 loop occurs in either intra- or inter-subunit pathways, while pre-TM1 occurs exclusively in inter-subunit pathways. Residues involved in both types of pathways are well supported by previous experimental data on nAChR. The direct coupling between pre-TM1 and TM2 of the adjacent subunit adds new insight into the allosteric signaling mechanism in pLGICs. © 2013 Mowrey et al

Extreme genetic fragility of the HIV-1 capsid

Genetic robustness, or fragility, is defined as the ability, or lack thereof, of a biological entity to maintain function in the face of mutations. Viruses that replicate via RNA intermediates exhibit high mutation rates, and robustness should be particularly advantageous to them. The capsid (CA) domain of the HIV-1 Gag protein is under strong pressure to conserve functional roles in viral assembly, maturation, uncoating, and nuclear import. However, CA is also under strong immunological pressure to diversify. Therefore, it would be particularly advantageous for CA to evolve genetic robustness. To measure the genetic robustness of HIV-1 CA, we generated a library of single amino acid substitution mutants, encompassing almost half the residues in CA. Strikingly, we found HIV-1 CA to be the most genetically fragile protein that has been analyzed using such an approach, with 70% of mutations yielding replication-defective viruses. Although CA participates in several steps in HIV-1 replication, analysis of conditionally (temperature sensitive) and constitutively non-viable mutants revealed that the biological basis for its genetic fragility was primarily the need to coordinate the accurate and efficient assembly of mature virions. All mutations that exist in naturally occurring HIV-1 subtype B populations at a frequency >3%, and were also present in the mutant library, had fitness levels that were >40% of WT. However, a substantial fraction of mutations with high fitness did not occur in natural populations, suggesting another form of selection pressure limiting variation in vivo. Additionally, known protective CTL epitopes occurred preferentially in domains of the HIV-1 CA that were even more genetically fragile than HIV-1 CA as a whole. The extreme genetic fragility of HIV-1 CA may be one reason why cell-mediated immune responses to Gag correlate with better prognosis in HIV-1 infection, and suggests that CA is a good target for therapy and vaccination strategies

Evaluation of routinely reported surgical site infections against microbiological culture results: a tool to identify patient groups where diagnosis and treatment may be improved

<p>Abstract</p> <p>Background</p> <p>Surgeons may improve their decision making by assessing the extent to which their initial clinical diagnosis of a surgical site infection (SSI) was supported by culture results. Aim of the present study was to evaluate routinely reported SSI by surgeons against microbiological culture results, to identify patient groups with lower agreement where decision making may be improved.</p> <p>Methods</p> <p>701 admissions with SSI were reported by surgeons in a university medical centre in the period 1997-2005, which were retrospectively checked for microbiological culture results. Reporting a SSI was conditional on treatment being given (e.g. antibiotics) and was classified by severity. To identify specific patient groups, patients were classified according to the surgery group of the first operation during admission (e.g. trauma).</p> <p>Results</p> <p>Of all reported SSI, 523 (74.6%) had a positive culture result, 102 (14.6%) a negative culture result and 76 (10.8%) were classified as unknown culture result (due to no culture taken). Given a known culture result, reported SSI with positive culture results less often concerned trauma patients (16% versus 26%, X²= 4.99 p = 0.03) and less severe SSI (49% versus 85%, X²= 10.11 p < 0.01) suggesting that a more conservative approach may be warranted in these patients. The trauma surgeons themselves perceived to have become too liberal in administering antibiotics (and reporting SSI).</p> <p>Conclusion</p> <p>Routine reporting of SSI was mostly supported by culture results. However, this support was less often found in trauma patients and less severe SSI, thereby giving surgeons feedback that diagnosis and treatment may be improved in these cases.</p

Yersinia pestis Evolution on a Small Timescale: Comparison of Whole Genome Sequences from North America

Yersinia pestis, the etiologic agent of plague, was responsible for several devastating epidemics throughout history and is currently of global importance to current public heath and biodefense efforts. Y. pestis is widespread in the Western United States. Because Y. pestis was first introduced to this region just over 100 years ago, there has been little time for genetic diversity to accumulate. Recent studies based upon single nucleotide polymorphisms have begun to quantify the genetic diversity of Y. pestis in North America.To examine the evolution of Y. pestis in North America, a gapped genome sequence of CA88-4125 was generated. Sequence comparison with another North American Y. pestis strain, CO92, identified seven regions of difference (six inversions, one rearrangement), differing IS element copy numbers, and several SNPs.The relatively large number of inverted/rearranged segments suggests that North American Y. pestis strains may be undergoing inversion fixation at high rates over a short time span, contributing to higher-than-expected diversity in this region. These findings will hopefully encourage the scientific community to sequence additional Y. pestis strains from North America and abroad, leading to a greater understanding of the evolutionary history of this pathogen

Complete Genomic Characterization of a Pathogenic A.II Strain of Francisella tularensis Subspecies tularensis

Francisella tularensis is the causative agent of tularemia, which is a highly lethal disease from nature and potentially from a biological weapon. This species contains four recognized subspecies including the North American endemic F. tularensis subsp. tularensis (type A), whose genetic diversity is correlated with its geographic distribution including a major population subdivision referred to as A.I and A.II. The biological significance of the A.I – A.II genetic differentiation is unknown, though there are suggestive ecological and epidemiological correlations. In order to understand the differentiation at the genomic level, we have determined the complete sequence of an A.II strain (WY96-3418) and compared it to the genome of Schu S4 from the A.I population. We find that this A.II genome is 1,898,476 bp in size with 1,820 genes, 1,303 of which code for proteins. While extensive genomic variation exists between “WY96” and Schu S4, there is only one whole gene difference. This one gene difference is a hypothetical protein of unknown function. In contrast, there are numerous SNPs (3,367), small indels (1,015), IS element differences (7) and large chromosomal rearrangements (31), including both inversions and translocations. The rearrangement borders are frequently associated with IS elements, which would facilitate intragenomic recombination events. The pathogenicity island duplicated regions (DR1 and DR2) are essentially identical in WY96 but vary relative to Schu S4 at 60 nucleotide positions. Other potential virulence-associated genes (231) varied at 559 nucleotide positions, including 357 non-synonymous changes. Molecular clock estimates for the divergence time between A.I and A.II genomes for different chromosomal regions ranged from 866 to 2131 years before present. This paper is the first complete genomic characterization of a member of the A.II clade of Francisella tularensis subsp. tularensis

GO4genome: A Prokaryotic Phylogeny Based on Genome Organization

Determining the phylogeny of closely related prokaryotes may fail in an analysis of rRNA or a small set of sequences. Whole-genome phylogeny utilizes the maximally available sample space. For a precise determination of genome similarity, two aspects have to be considered when developing an algorithm of whole-genome phylogeny: (1) gene order conservation is a more precise signal than gene content; and (2) when using sequence similarity, failures in identifying orthologues or the in situ replacement of genes via horizontal gene transfer may give misleading results. GO4genome is a new paradigm, which is based on a detailed analysis of gene function and the location of the respective genes. For characterization of genes, the algorithm uses gene ontology enabling a comparison of function independent of evolutionary relationship. After the identification of locally optimal series of gene functions, their length distribution is utilized to compute a phylogenetic distance. The outcome is a classification of genomes based on metabolic capabilities and their organization. Thus, the impact of effects on genome organization that are not covered by methods of molecular phylogeny can be studied. Genomes of strains belonging to Escherichia coli, Shigella, Streptococcus, Methanosarcina, and Yersinia were analyzed. Differences from the findings of classical methods are discussed

Measurement of the inclusive and dijet cross-sections of b-jets in pp collisions at sqrt(s) = 7 TeV with the ATLAS detector

The inclusive and dijet production cross-sections have been measured for jets containing b-hadrons (b-jets) in proton-proton collisions at a centre-of-mass energy of sqrt(s) = 7 TeV, using the ATLAS detector at the LHC. The measurements use data corresponding to an integrated luminosity of 34 pb^-1. The b-jets are identified using either a lifetime-based method, where secondary decay vertices of b-hadrons in jets are reconstructed using information from the tracking detectors, or a muon-based method where the presence of a muon is used to identify semileptonic decays of b-hadrons inside jets. The inclusive b-jet cross-section is measured as a function of transverse momentum in the range 20 < pT < 400 GeV and rapidity in the range |y| < 2.1. The bbbar-dijet cross-section is measured as a function of the dijet invariant mass in the range 110 < m_jj < 760 GeV, the azimuthal angle difference between the two jets and the angular variable chi in two dijet mass regions. The results are compared with next-to-leading-order QCD predictions. Good agreement is observed between the measured cross-sections and the predictions obtained using POWHEG + Pythia. MC@NLO + Herwig shows good agreement with the measured bbbar-dijet cross-section. However, it does not reproduce the measured inclusive cross-section well, particularly for central b-jets with large transverse momenta.Comment: 10 pages plus author list (21 pages total), 8 figures, 1 table, final version published in European Physical Journal

Observation of associated near-side and away-side long-range correlations in √sNN=5.02 TeV proton-lead collisions with the ATLAS detector

Two-particle correlations in relative azimuthal angle (Δϕ) and pseudorapidity (Δη) are measured in √sNN=5.02  TeV p+Pb collisions using the ATLAS detector at the LHC. The measurements are performed using approximately 1  μb-1 of data as a function of transverse momentum (pT) and the transverse energy (ΣETPb) summed over 3.1<η<4.9 in the direction of the Pb beam. The correlation function, constructed from charged particles, exhibits a long-range (2<|Δη|<5) “near-side” (Δϕ∼0) correlation that grows rapidly with increasing ΣETPb. A long-range “away-side” (Δϕ∼π) correlation, obtained by subtracting the expected contributions from recoiling dijets and other sources estimated using events with small ΣETPb, is found to match the near-side correlation in magnitude, shape (in Δη and Δϕ) and ΣETPb dependence. The resultant Δϕ correlation is approximately symmetric about π/2, and is consistent with a dominant cos⁡2Δϕ modulation for all ΣETPb ranges and particle pT

Measurement of the cross-section of high transverse momentum vector bosons reconstructed as single jets and studies of jet substructure in pp collisions at √s = 7 TeV with the ATLAS detector

This paper presents a measurement of the cross-section for high transverse momentum W and Z bosons produced in pp collisions and decaying to all-hadronic final states. The data used in the analysis were recorded by the ATLAS detector at the CERN Large Hadron Collider at a centre-of-mass energy of √s = 7 TeV;{\rm Te}{\rm V}$and correspond to an integrated luminosity of$4.6\;{\rm f}{{{\rm b}}^{-1}}$. The measurement is performed by reconstructing the boosted W or Z bosons in single jets. The reconstructed jet mass is used to identify the W and Z bosons, and a jet substructure method based on energy cluster information in the jet centre-of-mass frame is used to suppress the large multi-jet background. The cross-section for events with a hadronically decaying W or Z boson, with transverse momentum${{p}_{{\rm T}}}\gt 320\;{\rm Ge}{\rm V}$and pseudorapidity$|\eta |\lt 1.9$, is measured to be${{\sigma }_{W+Z}}=8.5\pm 1.7$ pb and is compared to next-to-leading-order calculations. The selected events are further used to study jet grooming techniques