Insights into the pre-initiation events of bacteriophage phi6 RNA-dependent RNA polymerase : towards the assembly of a productive binary complex

The RNA-dependent RNA polymerase (RdRP) of double-stranded RNA (dsRNA) viruses performs both RNA replication and transcription. In order to initiate RNA polymerization, viral RdRPs must be able to interact with the incoming 3 terminus of the template and position it, so that a productive binary complex is formed. Structural studies have revealed that RdRPs of dsRNA viruses that lack helicases have electrostatically charged areas on the polymerase surface, which might facilitate such interactions. In this study, structure-based mutagenesis, enzymatic assays and molecular mapping of bacteriophage 6 RdRP and its RNA were used to elucidate the roles of the negatively charged plough area on the polymerase surface, of the rim of the template tunnel and of the template specificity pocket that is key in the formation of the productive RNA-polymerase binary complex. The positively charged rim of the template tunnel has a significant role in the engagement of highly structured ssRNA molecules, whereas specific interactions further down in the template tunnel promote ssRNA entry to the catalytic site. Hence, we show that by aiding the formation of a stable binary complex with optimized RNA templates, the overall polymerization activity of the 6 RdRP can be greatly enhanced.The RNA-dependent RNA polymerase (RdRP) of double-stranded RNA (dsRNA) viruses performs both RNA replication and transcription. In order to initiate RNA polymerization, viral RdRPs must be able to interact with the incoming 3 terminus of the template and position it, so that a productive binary complex is formed. Structural studies have revealed that RdRPs of dsRNA viruses that lack helicases have electrostatically charged areas on the polymerase surface, which might facilitate such interactions. In this study, structure-based mutagenesis, enzymatic assays and molecular mapping of bacteriophage 6 RdRP and its RNA were used to elucidate the roles of the negatively charged plough area on the polymerase surface, of the rim of the template tunnel and of the template specificity pocket that is key in the formation of the productive RNA-polymerase binary complex. The positively charged rim of the template tunnel has a significant role in the engagement of highly structured ssRNA molecules, whereas specific interactions further down in the template tunnel promote ssRNA entry to the catalytic site. Hence, we show that by aiding the formation of a stable binary complex with optimized RNA templates, the overall polymerization activity of the 6 RdRP can be greatly enhanced.The RNA-dependent RNA polymerase (RdRP) of double-stranded RNA (dsRNA) viruses performs both RNA replication and transcription. In order to initiate RNA polymerization, viral RdRPs must be able to interact with the incoming 3 terminus of the template and position it, so that a productive binary complex is formed. Structural studies have revealed that RdRPs of dsRNA viruses that lack helicases have electrostatically charged areas on the polymerase surface, which might facilitate such interactions. In this study, structure-based mutagenesis, enzymatic assays and molecular mapping of bacteriophage 6 RdRP and its RNA were used to elucidate the roles of the negatively charged plough area on the polymerase surface, of the rim of the template tunnel and of the template specificity pocket that is key in the formation of the productive RNA-polymerase binary complex. The positively charged rim of the template tunnel has a significant role in the engagement of highly structured ssRNA molecules, whereas specific interactions further down in the template tunnel promote ssRNA entry to the catalytic site. Hence, we show that by aiding the formation of a stable binary complex with optimized RNA templates, the overall polymerization activity of the 6 RdRP can be greatly enhanced.Peer reviewe

Plate tectonics of virus shell assembly and reorganization in phage φ8, a distant relative of mammalian reoviruses

The hallmark of a virus is its capsid, which harbors the viral genome and is formed from protein subunits, which assemble following precise geometric rules. dsRNA viruses use an unusual protein multiplicity (120 copies) to form their closed capsids. We have determined the atomic structure of the capsid protein (P1) from the dsRNA cystovirus Φ8. In the crystal P1 forms pentamers, very similar in shape to facets of empty procapsids, suggesting an unexpected assembly pathway that proceeds via a pentameric intermediate. Unlike the elongated proteins used by dsRNA mammalian reoviruses, P1 has a compact trapezoid-like shape and a distinct arrangement in the shell, with two near-identical conformers in nonequivalent structural environments. Nevertheless, structural similarity with the analogous protein from the mammalian viruses suggests a common ancestor. The unusual shape of the molecule may facilitate dramatic capsid expansion during phage maturation, allowing P1 to switch interaction interfaces to provide capsid plasticity

First Measurement of Transferred Polarization in the Exclusive e p --> e' K+ Lambda Reaction

The first measurements of the transferred polarization for the exclusive ep --> e'K+ Lambda reaction have been performed in Hall B at the Thomas Jefferson National Accelerator Facility using the CLAS spectrometer. A 2.567 GeV electron beam was used to measure the hyperon polarization over a range of Q2 from 0.3 to 1.5 (GeV/c)2, W from 1.6 to 2.15 GeV, and over the full center-of-mass angular range of the K+ meson. Comparison with predictions of hadrodynamic models indicates strong sensitivity to the underlying resonance contributions. A non-relativistic quark model interpretation of our data suggests that the s-sbar quark pair is produced with spins predominantly anti-aligned. Implications for the validity of the widely used 3P0 quark-pair creation operator are discussed.Comment: 6 pages, 4 figure

The interaction between vaginal microbiota, cervical length, and vaginal progesterone treatment for preterm birth risk

© The Author(s) 2017. Background: Preterm birth is the primary cause of infant death worldwide. A short cervix in the second trimester of pregnancy is a risk factor for preterm birth. In specific patient cohorts, vaginal progesterone reduces this risk. Using 16S rRNA gene sequencing, we undertook a prospective study in women at risk of preterm birth (n = 161) to assess (1) the relationship between vaginal microbiota and cervical length in the second trimester and preterm birth risk and (2) the impact of vaginal progesterone on vaginal bacterial communities in women with a short cervix. Results: Lactobacillus iners dominance at 16 weeks of gestation was significantly associated with both a short cervix < 25 mm (n = 15, P < 0.05) and preterm birth < 34+0 weeks (n = 18; P < 0.01; 69% PPV). In contrast, Lactobacillus crispatus dominance was highly predictive of term birth (n = 127, 98% PPV). Cervical shortening and preterm birth were not associated with vaginal dysbiosis. A longitudinal characterization of vaginal microbiota (< 18, 22, 28, and 34 weeks) was then undertaken in women receiving vaginal progesterone (400 mg/OD, n = 25) versus controls (n = 42). Progesterone did not alter vaginal bacterial community structure nor reduce L. iners-associated preterm birth (< 34 weeks). Conclusions: L. iners dominance of the vaginal microbiota at 16 weeks of gestation is a risk factor for preterm birth, whereas L. crispatus dominance is protective against preterm birth. Vaginal progesterone does not appear to impact the pregnancy vaginal microbiota. Patients and clinicians who may be concerned about "infection risk" associated with the use of a vaginal pessary during high-risk pregnancy can be reassured

Measurement of the prompt J/psi and psi(2S) polarizations in pp collisions at sqrt(s) = 7 TeV

The polarizations of prompt J/psi and psi(2S) mesons are measured in proton-proton collisions at sqrt(s) = 7 TeV, using a dimuon data sample collected by the CMS experiment at the LHC, corresponding to an integrated luminosity of 4.9 inverse femtobarns. The prompt J/psi and psi(2S) polarization parameters lambda[theta], lambda[phi], and lambda[theta, phi], as well as the frame-invariant quantity lambda(tilde), are measured from the dimuon decay angular distributions in three different polarization frames. The J/psi results are obtained in the transverse momentum range 14 &lt; pt &lt; 70 GeV, in the rapidity intervals abs(y) &lt; 0.6 and 0.6 &lt; abs(y) &lt; 1.2. The corresponding psi(2S) results cover 14 &lt; pt &lt; 50 GeV and include a third rapidity bin, 1.2 &lt; abs(y) &lt; 1.5. No evidence of large transverse or longitudinal polarizations is seen in these kinematic regions, which extend much beyond those previously explored

Schadevergoeding bij overlijden: een stoel die een soort tafeltje is

In het aansprakelijkheidsrecht heeft een benadeelde in beginsel recht op volledige vergoeding van zijn schade. De gevolgen van de schadetoebrengende gebeurtenis dienen zoveel als mogelijk te worden weggenomen of te worden gecompenseerd. Daarbij wordt gekeken naar de situatie waarin de benadeelde zou hebben verkeerd indien de schadetoebrengende gebeurtenis niet zou hebben plaatsgevonden. Dat is bij overlijden per definitie problematisch. Er is iemand weggevallen, wat vele gevolgen heeft. De schade als gevolg van het overlijden komt maar beperkt voor vergoeding in aanmerking. In artikel 6:108 BW is een drietal beperkingen te vinden. Het gaat hier om beperkingen ten aanzien van de aard van de schade, de kring van gerechtigden en de omvang van de schade. Daarbij hinkt het recht op schadevergoeding bij overlijden op twee gedachten. Aan de ene kant is er het aansprakelijkheidsrecht, maar de geleden schade komt niet volledig voor vergoeding in aanmerking. Aan de andere kant is er het recht op alimentatie uit het familierecht, maar dat wordt bij overlijden niet consequent toegepast. De motieven voor de beperkingen van het recht op schadevergoeding zijn achterhaald en niet (langer) overtuigend. Het recht is niet bij de tijd, het sluit niet aan bij de maatschappelijke ontwikkelingen. De beperkingen die het recht op schadevergoeding bij overlijden in de huidige samenleving met zich meebrengt zorgen voor complexe methoden om de nabestaanden tegemoet te komen en oogsten daardoor veel kritiek. In deze bijdrage wordt die kritiek besproken. De bijdrage wordt afgesloten met enkele denkrichtingen voor nader onderzoek

Alignment of the CMS tracker with LHC and cosmic ray data

© CERN 2014 for the benefit of the CMS collaboration, published under the terms of the Creative Commons Attribution 3.0 License by IOP Publishing Ltd and Sissa Medialab srl. Any further distribution of this work must maintain attribution to the author(s) and the published article's title, journal citation and DOI.The central component of the CMS detector is the largest silicon tracker ever built. The precise alignment of this complex device is a formidable challenge, and only achievable with a significant extension of the technologies routinely used for tracking detectors in the past. This article describes the full-scale alignment procedure as it is used during LHC operations. Among the specific features of the method are the simultaneous determination of up to 200 000 alignment parameters with tracks, the measurement of individual sensor curvature parameters, the control of systematic misalignment effects, and the implementation of the whole procedure in a multi-processor environment for high execution speed. Overall, the achieved statistical accuracy on the module alignment is found to be significantly better than 10μm

Genetic mechanisms of critical illness in COVID-19.

Host-mediated lung inflammation is present1, and drives mortality2, in the critical illness caused by coronavirus disease 2019 (COVID-19). Host genetic variants associated with critical illness may identify mechanistic targets for therapeutic development3. Here we report the results of the GenOMICC (Genetics Of Mortality In Critical Care) genome-wide association study in 2,244 critically ill patients with COVID-19 from 208 UK intensive care units. We have identified and replicated the following new genome-wide significant associations: on chromosome 12q24.13 (rs10735079, P = 1.65 × 10-8) in a gene cluster that encodes antiviral restriction enzyme activators (OAS1, OAS2 and OAS3); on chromosome 19p13.2 (rs74956615, P = 2.3 × 10-8) near the gene that encodes tyrosine kinase 2 (TYK2); on chromosome 19p13.3 (rs2109069, P = 3.98 ×  10-12) within the gene that encodes dipeptidyl peptidase 9 (DPP9); and on chromosome 21q22.1 (rs2236757, P = 4.99 × 10-8) in the interferon receptor gene IFNAR2. We identified potential targets for repurposing of licensed medications: using Mendelian randomization, we found evidence that low expression of IFNAR2, or high expression of TYK2, are associated with life-threatening disease; and transcriptome-wide association in lung tissue revealed that high expression of the monocyte-macrophage chemotactic receptor CCR2 is associated with severe COVID-19. Our results identify robust genetic signals relating to key host antiviral defence mechanisms and mediators of inflammatory organ damage in COVID-19. Both mechanisms may be amenable to targeted treatment with existing drugs. However, large-scale randomized clinical trials will be essential before any change to clinical practice