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1H, 13C and 15N chemical shift assignment of the stem-loops 5b + c from the 5′-UTR of SARS-CoV-2
(2022)
The ongoing pandemic of the respiratory disease COVID-19 is caused by the SARS-CoV-2 (SCoV2) virus. SCoV2 is a member of the Betacoronavirus genus. The 30 kb positive sense, single stranded RNA genome of SCoV2 features 5′- and 3′-genomic ends that are highly conserved among Betacoronaviruses. These genomic ends contain structured cis-acting RNA elements, which are involved in the regulation of viral replication and translation. Structural information about these potential antiviral drug targets supports the development of novel classes of therapeutics against COVID-19. The highly conserved branched stem-loop 5 (SL5) found within the 5′-untranslated region (5′-UTR) consists of a basal stem and three stem-loops, namely SL5a, SL5b and SL5c. Both, SL5a and SL5b feature a 5′-UUUCGU-3′ hexaloop that is also found among Alphacoronaviruses. Here, we report the extensive 1H, 13C and 15N resonance assignment of the 37 nucleotides (nts) long sequence spanning SL5b and SL5c (SL5b + c), as basis for further in-depth structural studies by solution NMR spectroscopy.
The European Beech is the dominant climax tree in most regions of Central Europe and valued for its ecological versatility and hardwood timber. Even though a draft genome has been published recently, higher resolution is required for studying aspects of genome architecture and recombination. Here, we present a chromosome-level assembly of the more than 300 year-old reference individual, Bhaga, from the Kellerwald-Edersee National Park (Germany). Its nuclear genome of 541 Mb was resolved into 12 chromosomes varying in length between 28 and 73 Mb. Multiple nuclear insertions of parts of the chloroplast genome were observed, with one region on chromosome 11 spanning more than 2 Mb which fragments up to 54,784 bp long and covering the whole chloroplast genome were inserted randomly. Unlike in Arabidopsis thaliana, ribosomal cistrons are present in Fagus sylvatica only in four major regions, in line with FISH studies. On most assembled chromosomes, telomeric repeats were found at both ends, while centromeric repeats were found to be scattered throughout the genome apart from their main occurrence per chromosome. The genome-wide distribution of SNPs was evaluated using a second individual from Jamy Nature Reserve (Poland). SNPs, repeat elements and duplicated genes were unevenly distributed in the genomes, with one major anomaly on chromosome 4. The genome presented here adds to the available highly resolved plant genomes and we hope it will serve as a valuable basis for future research on genome architecture and for understanding the past and future of European Beech populations in a changing climate.
Species of the genus Blautia are typical inhabitants of the human gut and considered as beneficial gut microbes. However, their role in the gut microbiome and their metabolic features are poorly understood. Blautia schinkii was described as an acetogenic bacterium, characterized by a functional Wood–Ljungdahl pathway (WLP) of acetogenesis from H2 + CO2. Here we report that two relatives, Blautia luti and Blautia wexlerae do not grow on H2 + CO2. Inspection of the genome sequence revealed all genes of the WLP except genes encoding a formate dehydrogenase and an electron-bifurcating hydrogenase. Enzyme assays confirmed this prediction. Accordingly, resting cells neither converted H2 + CO2 nor H2 + HCOOH + CO2 to acetate. Carbon monoxide is an intermediate of the WLP and substrate for many acetogens. Blautia luti and B. wexlerae had an active CO dehydrogenase and resting cells performed acetogenesis from HCOOH + CO2 + CO, demonstrating a functional WLP. Bioinformatic analyses revealed that many Blautia strains as well as other gut acetogens lack formate dehydrogenases and hydrogenases. Thus, the use of formate instead of H2 + CO2 as an interspecies hydrogen and electron carrier seems to be more common in the gut microbiome.
Although new advances in neuroscience allow the study of vocal communication in awake animals, substantial progress in the processing of vocalizations has been made from brains of anaesthetized preparations. Thus, understanding how anaesthetics affect neuronal responses is of paramount importance. Here, we used electrophysiological recordings and computational modelling to study how the auditory cortex of bats responds to vocalizations under anaesthesia and in wakefulness. We found that multifunctional neurons that process echolocation and communication sounds were affected by ketamine anaesthesia in a manner that could not be predicted by known anaesthetic effects. In wakefulness, acoustic contexts (preceding echolocation or communication sequences) led to stimulus-specific suppression of lagging sounds, accentuating neuronal responses to sound transitions. However, under anaesthesia, communication contexts (but not echolocation) led to a global suppression of responses to lagging sounds. Such asymmetric effect was dependent on the frequency composition of the contexts and not on their temporal patterns. We constructed a neuron model that could replicate the data obtained in vivo. In the model, anaesthesia modulates spiking activity in a channel-specific manner, decreasing responses of cortical inputs tuned to high-frequency sounds and increasing adaptation in the respective cortical synapses. Combined, our findings obtained in vivo and in silico reveal that ketamine anaesthesia does not reduce uniformly the neurons’ responsiveness to low and high frequency sounds. This effect depends on combined mechanisms that unbalance cortical inputs and ultimately affect how auditory cortex neurons respond to natural sounds in anaesthetized preparations.
Detailed information on species temperature preferences are needed to measure the effects of global warming on species and communities in European rivers. However, information currently available in the literature on taxon-specific temperature preferences or temperature tolerances is very heterogeneous and therefore not well suited for forecasting purposes. To close this gap, we derived so-called ’central temperature tendencies’ (CTTt values) for benthic invertebrate species. For this end, 547 species and temperature data from regional monitoring programmes in Germany collected at 4249 sites were analysed. Due to the vulnerability of species to high
temperatures, CTTt values were calculated for mean summer temperatures, following a robust approach of calculating a weighted average based on temperature classes. Derived CTTt values correspond well to species temperature preferences as reported in literature as long as the latter were homogeneous in terms of how they were derived and which temperature reference was at focus. Based on taxon-specific CTTt values, a community value, CTTCom, was calculated for each benthic invertebrate sample. CTTCom values were validated by correlation with mean summer water temperatures. As the slope a of the linear regression model between CTTCom values and measured summer temperatures was comparatively low (a = 0.49), a correction function was derived in order to optimise the relation between both. This was crucial, because it is assumed that although CTTt was derived solely from taxa abundances within summer temperature classes, CTTCom not only reflects the effect of (summer) water temperature itself, but also corresponds to a temperature equivalent value, which describes the overall quality of all respiration-relevant aquatic summer habitat conditions that determine the metabolism of respective benthic invertebrates. By comparing this equivalent value with water temperatures measured in the year previous of sampling, statements can be made about the influence of flow conditions and other factors determining oxygen availability.
Thus, CTTCom reflects the mean aerobic scope of the overall benthic invertebrate fauna: the better the respiration conditions for rheophilic species with high oxygen demand, the larger the aerobic scope and the lower CTTCom.
The approach taken in our study is promising and provides a tool to track and even project past, present, and future impacts of global warming on benthic invertebrates in rivers based on measured values of respiratory relevant environmental variables. We encourage all stakeholders in the field of freshwater ecology to test this
Plasmids are one of the most important genetic tools for basic research and biotechnology, as they enable rapid genetic manipulation. Here we present a novel pBBR1-based plasmid for Methylorubrum extorquens, a model methylotroph that is used for the development of C1-based microbial cell factories. To develop a vector with compatibility to the so far mainly used pCM plasmid system, we transferred the pBBR1-based plasmid pMiS1, which showed an extremely low transformation rate and caused a strong growth defect. Isolation of a suppressor mutant with improved growth led to the isolation of the variant pMis1_1B. Its higher transformation rate and less pronounced growth defect phenotype could be shown to be the result of a mutation in the promotor region of the rep gene. Moreover, cotransformation of pMis1_1B and pCM160 was possible, but the resulting transformants showed stronger growth defects in comparison with a single pMis1_1B transformant. Surprisingly, cotransformants carrying pCM160 and a pMis1_1B derivative containing a mCherry reporter construct showed higher fluorescence levels than strains containing only the pMis1_1B-based reporter plasmids or a corresponding pCM160 derivative. Relative plasmid copy number determination experiments confirmed our hypothesis of an increased copy number of pMis1_1B in the strain carrying both plasmids. Despite the slight metabolic burden caused by pMis1_1B, the plasmid strongly expands the genetic toolbox for M. extorquens.
Energy-converting hydrogenases (Ech) are ancient, membrane-bound enzymes that use reduced ferredoxin (Fd) as an electron donor to reduce protons to molecular H2. Experiments with whole cells, membranes and vesicle-fractions suggest that proton reduction is coupled to proton translocation across the cytoplasmatic membrane, but this has never been demonstrated with a purified enzyme. To this end, we produced a His-tagged Ech complex in the thermophilic and anaerobic bacterium Thermoanaerobacter kivui. The enzyme could be purified by affinity chromatography from solubilized membranes with full retention of its eight subunits, as well as full retention of physiological activities, i.e., H2-dependent Fd reduction and Fd2--dependent H2 production. We found the purified enzyme contained 34.2 ± 12.2 mol of iron/mol of protein, in accordance with seven predicted [4Fe-4S]-clusters and one [Ni-Fe]-center. The pH and temperature optima were at 7 to 8 and 66 °C, respectively. Notably, we found that the enzymatic activity was inhibited by N,N′-dicyclohexylcarbodiimide, an agent known to bind ion-translocating glutamates or aspartates buried in the cytoplasmic membrane and thereby inhibiting ion transport. To demonstrate the function of the Ech complex in ion transport, we further established a procedure to incorporate the enzyme complex into liposomes in an active state. We show the enzyme did not require Na+ for activity and did not translocate 22Na+ into the proteoliposomal lumen. In contrast, Ech activity led to the generation of a pH gradient and membrane potential across the proteoliposomal membrane, demonstrating that the Ech complex of T. kivui is a H+-translocating, H+-reducing enzyme.
Orientation hypercolumns in the visual cortex are delimited by the repeating pinwheel patterns of orientation selective neurons. We design a generative model for visual cortex maps that reproduces such orientation hypercolumns as well as ocular dominance maps while preserving retinotopy. The model uses a neural placement method based on t–distributed stochastic neighbour embedding (t–SNE) to create maps that order common features in the connectivity matrix of the circuit. We find that, in our model, hypercolumns generally appear with fixed cell numbers independently of the overall network size. These results would suggest that existing differences in absolute pinwheel densities are a consequence of variations in neuronal density. Indeed, available measurements in the visual cortex indicate that pinwheels consist of a constant number of ∼30, 000 neurons. Our model is able to reproduce a large number of characteristic properties known for visual cortex maps. We provide the corresponding software in our MAPStoolbox for Matlab.
Bisphenols and phthalates, chemicals frequently used in plastic products, promote obesity in cell and animal models. However, these well-known metabolism-disrupting chemicals (MDCs) represent only a minute fraction of all compounds found in plastics. To gain a comprehensive understanding of plastics as a source of exposure to MDCs, we characterized the chemicals present in 34 everyday products using nontarget high-resolution mass spectrometry and analyzed their joint adipogenic activities by high-content imaging. We detected 55,300 chemical features and tentatively identified 629 unique compounds, including 11 known MDCs. Importantly, the chemicals extracted from one-third of the products caused murine 3T3-L1 preadipocytes to proliferate, and differentiate into adipocytes, which were larger and contained more triglycerides than those treated with the reference compound rosiglitazone. Because the majority of plastic extracts did not activate the peroxisome proliferator-activated receptor γ and the glucocorticoid receptor, the adipogenic effects are mediated via other mechanisms and, thus, likely to be caused by unknown MDCs. Our study demonstrates that daily-use plastics contain potent mixtures of MDCs and can, therefore, be a relevant yet underestimated environmental factor contributing to obesity.
Animals use the geomagnetic field and astronomical cues to obtain compass information. The magnetic compass is not a uniform mechanism, as several functional modes have been described in different animal groups. The Sun compass requires the internal clock to interpret the position of the Sun. For star compass orientation, night-migrating birds seem to use the star pattern as a whole, without involving the internal clock. Both the astronomical compass mechanisms are based on learning processes to adapt them to the geographic latitude where the animals live and, in long-living animals, to compensate for the seasonal changes. Several mechanisms are used to determine the compass course to a goal. Using information collected during the outward journey is mostly done by path integration: recording the direction with a compass and integrating its twists and turns. Migratory animals have innate programs to guide them to their still unknown goal. Highly mobile animals with large ranges develop a so-called navigational ‘map’, a mental representation of the spatial distribution of navigational factors within their home region and their migration route. The nature of the factors involved is not yet entirely clear; magnetic intensity and inclination are the ones best supported so far.