Asymmetry of 13C labeled 3-pyruvate affords improved site specific labeling of RNA for NMR spectroscopy

Selective isotopic labeling provides an unparalleled window within which to study the structure and dynamics of RNAs by high resolution NMR spectroscopy. Unlike commonly used carbon sources, the asymmetry of 13C-labeled pyruvate provides selective labeling in both the ribose and base moieties of nucleotides using E. coli variants, that until now were not feasible. Here we show that an E. coli mutant strain that lacks succinate and malate dehydrogenases (DL323) and grown on [3-13C]-pyruvate affords ribonucleotides with site specific labeling at C5′ (~95%) and C1′ (~42%) and minimal enrichment elsewhere in the ribose ring. Enrichment is also achieved at purine C2 and C8 (~95%) and pyrimidine C5 (~100%) positions with minimal labeling at pyrimidine C6 and purine C5 positions. These labeling patterns contrast with those obtained with DL323 E. coli grown on [1, 3-13C]-glycerol for which the ribose ring is labeled in all but the C4′ carbon position, leading to multiplet splitting of the C1′, C2′ and C3′ carbon atoms. The usefulness of these labeling patterns is demonstrated with a 27-nt RNA fragment derived from the 30S ribosomal subunit. Removal of the strong magnetic coupling within the ribose and base leads to increased sensitivity, substantial simplification of NMR spectra, and more precise and accurate dynamic parameters derived from NMR relaxation measurements. Thus these new labels offer valuable probes for characterizing the structure and dynamics of RNA that were previously limited by the constraint of uniformly labeled nucleotides

Biomass production of site selective 13C/15N nucleotides using wild type and a transketolase E. coli mutant for labeling RNA for high resolution NMR

Characterization of the structure and dynamics of nucleic acids by NMR benefits significantly from position specifically labeled nucleotides. Here an E. coli strain deficient in the transketolase gene (tktA) and grown on glucose that is labeled at different carbon sites is shown to facilitate cost-effective and large scale production of useful nucleotides. These nucleotides are site specifically labeled in C1′ and C5′ with minimal scrambling within the ribose ring. To demonstrate the utility of this labeling approach, the new site-specific labeled and the uniformly labeled nucleotides were used to synthesize a 36-nt RNA containing the catalytically essential domain 5 (D5) of the brown algae group II intron self-splicing ribozyme. The D5 RNA was used in binding and relaxation studies probed by NMR spectroscopy. Key nucleotides in the D5 RNA that are implicated in binding Mg2+ ions are well resolved. As a result, spectra obtained using selectively labeled nucleotides have higher signal-to-noise ratio compared to those obtained using uniformly labeled nucleotides. Thus, compared to the uniformly 13C/15N-labeled nucleotides, these specifically labeled nucleotides eliminate the extensive 13C–13C coupling within the nitrogenous base and ribose ring, give rise to less crowded and more resolved NMR spectra, and accurate relaxation rates without the need for constant-time or band-selective decoupled NMR experiments. These position selective labeled nucleotides should, therefore, find wide use in NMR analysis of biologically interesting RNA molecules

Genomic screening of 16 UK native bat species through conservationist networks uncovers coronaviruses with zoonotic potential

There has been limited characterisation of bat-borne coronaviruses in Europe. Here, we screened for coronaviruses in 48 faecal samples from 16 of the 17 bat species breeding in the UK, collected through a bat rehabilitation and conservationist network. We recovered nine complete genomes, including two novel coronavirus species, across six bat species: four alphacoronaviruses, a MERS-related betacoronavirus, and four closely related sarbecoviruses. We demonstrate that at least one of these sarbecoviruses can bind and use the human ACE2 receptor for infecting human cells, albeit suboptimally. Additionally, the spike proteins of these sarbecoviruses possess an R-A-K-Q motif, which lies only one nucleotide mutation away from a furin cleavage site (FCS) that enhances infectivity in other coronaviruses, including SARS-CoV-2. However, mutating this motif to an FCS does not enable spike cleavage. Overall, while UK sarbecoviruses would require further molecular adaptations to infect humans, their zoonotic risk warrants closer surveillance

Global, regional, and national comparative risk assessment of 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks, 1990-2015: a systematic analysis for the Global Burden of Disease Study 2015

SummaryBackground The Global Burden of Diseases, Injuries, and Risk Factors Study 2015 provides an up-to-date synthesis of the evidence for risk factor exposure and the attributable burden of disease. By providing national and subnational assessments spanning the past 25 years, this study can inform debates on the importance of addressing risks in context. Methods We used the comparative risk assessment framework developed for previous iterations of the Global Burden of Disease Study to estimate attributable deaths, disability-adjusted life-years (DALYs), and trends in exposure by age group, sex, year, and geography for 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks from 1990 to 2015. This study included 388 risk-outcome pairs that met World Cancer Research Fund-defined criteria for convincing or probable evidence. We extracted relative risk and exposure estimates from randomised controlled trials, cohorts, pooled cohorts, household surveys, census data, satellite data, and other sources. We used statistical models to pool data, adjust for bias, and incorporate covariates. We developed a metric that allows comparisons of exposure across risk factors—the summary exposure value. Using the counterfactual scenario of theoretical minimum risk level, we estimated the portion of deaths and DALYs that could be attributed to a given risk. We decomposed trends in attributable burden into contributions from population growth, population age structure, risk exposure, and risk-deleted cause-specific DALY rates. We characterised risk exposure in relation to a Socio-demographic Index (SDI). Findings Between 1990 and 2015, global exposure to unsafe sanitation, household air pollution, childhood underweight, childhood stunting, and smoking each decreased by more than 25%. Global exposure for several occupational risks, high body-mass index (BMI), and drug use increased by more than 25% over the same period. All risks jointly evaluated in 2015 accounted for 57·8% (95% CI 56·6–58·8) of global deaths and 41·2% (39·8–42·8) of DALYs. In 2015, the ten largest contributors to global DALYs among Level 3 risks were high systolic blood pressure (211·8 million [192·7 million to 231·1 million] global DALYs), smoking (148·6 million [134·2 million to 163·1 million]), high fasting plasma glucose (143·1 million [125·1 million to 163·5 million]), high BMI (120·1 million [83·8 million to 158·4 million]), childhood undernutrition (113·3 million [103·9 million to 123·4 million]), ambient particulate matter (103·1 million [90·8 million to 115·1 million]), high total cholesterol (88·7 million [74·6 million to 105·7 million]), household air pollution (85·6 million [66·7 million to 106·1 million]), alcohol use (85·0 million [77·2 million to 93·0 million]), and diets high in sodium (83·0 million [49·3 million to 127·5 million]). From 1990 to 2015, attributable DALYs declined for micronutrient deficiencies, childhood undernutrition, unsafe sanitation and water, and household air pollution; reductions in risk-deleted DALY rates rather than reductions in exposure drove these declines. Rising exposure contributed to notable increases in attributable DALYs from high BMI, high fasting plasma glucose, occupational carcinogens, and drug use. Environmental risks and childhood undernutrition declined steadily with SDI; low physical activity, high BMI, and high fasting plasma glucose increased with SDI. In 119 countries, metabolic risks, such as high BMI and fasting plasma glucose, contributed the most attributable DALYs in 2015. Regionally, smoking still ranked among the leading five risk factors for attributable DALYs in 109 countries; childhood underweight and unsafe sex remained primary drivers of early death and disability in much of sub-Saharan Africa. Interpretation Declines in some key environmental risks have contributed to declines in critical infectious diseases. Some risks appear to be invariant to SDI. Increasing risks, including high BMI, high fasting plasma glucose, drug use, and some occupational exposures, contribute to rising burden from some conditions, but also provide opportunities for intervention. Some highly preventable risks, such as smoking, remain major causes of attributable DALYs, even as exposure is declining. Public policy makers need to pay attention to the risks that are increasingly major contributors to global burden. Funding Bill & Melinda Gates Foundation

A review of elliptical and disc galaxy structure, and modern scaling laws

A century ago, in 1911 and 1913, Plummer and then Reynolds introduced their models to describe the radial distribution of stars in `nebulae'. This article reviews the progress since then, providing both an historical perspective and a contemporary review of the stellar structure of bulges, discs and elliptical galaxies. The quantification of galaxy nuclei, such as central mass deficits and excess nuclear light, plus the structure of dark matter halos and cD galaxy envelopes, are discussed. Issues pertaining to spiral galaxies including dust, bulge-to-disc ratios, bulgeless galaxies, bars and the identification of pseudobulges are also reviewed. An array of modern scaling relations involving sizes, luminosities, surface brightnesses and stellar concentrations are presented, many of which are shown to be curved. These 'redshift zero' relations not only quantify the behavior and nature of galaxies in the Universe today, but are the modern benchmark for evolutionary studies of galaxies, whether based on observations, N-body-simulations or semi-analytical modelling. For example, it is shown that some of the recently discovered compact elliptical galaxies at 1.5 < z < 2.5 may be the bulges of modern disc galaxies.Comment: Condensed version (due to Contract) of an invited review article to appear in "Planets, Stars and Stellar Systems"(www.springer.com/astronomy/book/978-90-481-8818-5). 500+ references incl. many somewhat forgotten, pioneer papers. Original submission to Springer: 07-June-201

No evidence for association between SLC11A1 and visceral leishmaniasis in India.

BACKGROUND: SLC11A1 has pleiotropic effects on macrophage function and remains a strong candidate for infectious disease susceptibility. 5' and/or 3' polymorphisms have been associated with tuberculosis, leprosy, and visceral leishmaniasis (VL). Most studies undertaken to date were under-powered, and none has been replicated within a population. Association with tuberculosis has replicated variably across populations. Here we investigate SLC11A1 and VL in India. METHODS: Nine polymorphisms (rs34448891, rs7573065, rs2276631, rs3731865, rs17221959, rs2279015, rs17235409, rs17235416, rs17229009) that tag linkage disequilibrium blocks across SLC11A1 were genotyped in primary family-based (313 cases; 176 families) and replication (941 cases; 992 controls) samples. Family- and population-based analyses were performed to look for association between SLC11A1 variants and VL. Quantitative RT/PCR was used to compare SLC11A1 expression in mRNA from paired splenic aspirates taken before and after treatment from 24 VL patients carrying different genotypes at the functional promoter GTn polymorphism (rs34448891). RESULTS: No associations were observed between VL and polymorphisms at SLC11A1 that were either robust to correction for multiple testing or replicated across primary and replication samples. No differences in expression of SLC11A1 were observed when comparing pre- and post-treatment samples, or between individuals carrying different genotypes at the GTn repeat. CONCLUSIONS: This is the first well-powered study of SLC11A1 as a candidate for VL, which we conclude does not have a major role in regulating VL susceptibility in India.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

The anti-sigma factor RsrA responds to oxidative stress by reburying its hydrophobic core

Redox-regulated effector systems that counteract oxidative stress are essential for all forms of life. Here we uncover a new paradigm for sensing oxidative stress centred on the hydrophobic core of a sensor protein. RsrA is an archetypal zinc-binding anti-sigma factor that responds to disulfide stress in the cytoplasm of Actinobacteria. We show that RsrA utilizes its hydrophobic core to bind the sigma factor σ R preventing its association with RNA polymerase, and that zinc plays a central role in maintaining this high-affinity complex. Oxidation of RsrA is limited by the rate of zinc release, which weakens the RsrA-σ R complex by accelerating its dissociation. The subsequent trigger disulfide, formed between specific combinations of RsrA's three zinc-binding cysteines, precipitates structural collapse to a compact state where all σ R-binding residues are sequestered back into its hydrophobic core, releasing σ R to activate transcription of anti-oxidant genes

Classification and Regression Tree and Spatial Analyses Reveal Geographic Heterogeneity in Genome Wide Linkage Study of Indian Visceral Leishmaniasis

Genome wide linkage studies (GWLS) have provided evidence for loci controlling visceral leishmaniasis on Chromosomes 1p22, 6q27, 22q12 in Sudan and 6q27, 9p21, 17q11-q21 in Brazil. Genome wide studies from the major focus of disease in India have not previously been reported.We undertook a GWLS in India in which a primary ∼10 cM (515 microsatellites) scan was carried out in 58 multicase pedigrees (74 nuclear families; 176 affected, 353 total individuals) and replication sought in 79 pedigrees (102 nuclear families; 218 affected, 473 total individuals). The primary scan provided evidence (≥2 adjacent markers allele-sharing LOD≥0.59; nominal P≤0.05) for linkage on Chromosomes 2, 5, 6, 7, 8, 10, 11, 20 and X, with peaks at 6p25.3-p24.3 and 8p23.1-p21.3 contributed to largely by 31 Hindu families and at Xq21.1-q26.1 by 27 Muslim families. Refined mapping confirmed linkage across all primary scan families at 2q12.2-q14.1 and 11q13.2-q23.3, but only 11q13.2-q23.3 replicated (combined LOD = 1.59; P = 0.0034). Linkage at 6p25.3-p24.3 and 8p23.1-p21.3, and at Xq21.1-q26.1, was confirmed by refined mapping for primary Hindu and Muslim families, respectively, but only Xq21.1-q26.1 replicated across all Muslim families (combined LOD 1.49; P = 0.0045). STRUCTURE and SMARTPCA did not identify population genetic substructure related to religious group. Classification and regression tree, and spatial interpolation, analyses confirm geographical heterogeneity for linkages at 6p25.3-p24.3, 8p23.1-p21.3 and Xq21.1-q26.1, with specific clusters of families contributing LOD scores of 2.13 (P = 0.0009), 1.75 (P = 0.002) and 1.84 (P = 0.001), respectively.GWLS has identified novel loci that show geographical heterogeneity in their influence on susceptibility to VL in India

A Battle Lost? Report on Two Centuries of Invasion and Management of Lantana camara L. in Australia, India and South Africa

Recent discussion on invasive species has invigorated the debate on strategies to manage these species. Lantana camara L., a shrub native to the American tropics, has become one of the worst weeds in recorded history. In Australia, India and South Africa, Lantana has become very widespread occupying millions of hectares of land. Here, we examine historical records to reconstruct invasion and management of Lantana over two centuries and ask: Can we fight the spread of invasive species or do we need to develop strategies for their adaptive management? We carried out extensive research of historical records constituting over 75% of records on invasion and management of this species in the three countries. The records indicate that governments in Australia, India and South Africa have taken aggressive measures to eradicate Lantana over the last two centuries, but these efforts have been largely unsuccessful. We found that despite control measures, the invasion trajectory of Lantana has continued upwards and that post-war land-use change might have been a possible trigger for this spread. A large majority of studies on invasive species address timescales of less than one year; and even fewer address timescales of >10 years. An understanding of species invasions over long time-scales is of paramount importance. While archival records may give only a partial picture of the spread and management of invasive species, in the absence of any other long-term dataset on the ecology of Lantana, our study provides an important insight into its invasion, spread and management over two centuries and across three continents. While the established paradigm is to expend available resources on attempting to eradicate invasive species, our findings suggest that in the future, conservationists will need to develop strategies for their adaptive management rather than fighting a losing battle