Evaluation of Azido 3-Deoxy- d - Manno-oct-2-ulosonic Acid (Kdo) Analogues for Click Chemistry-Mediated Metabolic Labeling of Myxococcus xanthus DZ2 Lipopolysaccharide

[Image: see text] Metabolic labeling paired with click chemistry is a powerful approach for selectively imaging the surfaces of diverse bacteria. Herein, we explored the feasibility of labeling the lipopolysaccharide (LPS) of Myxococcus xanthus—a Gram-negative predatory social bacterium known to display complex outer membrane (OM) dynamics—via growth in the presence of distinct azido (-N(3)) analogues of 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo). Determination of the LPS carbohydrate structure from strain DZ2 revealed the presence of one Kdo sugar in the core oligosaccharide, modified with phosphoethanolamine. The production of 8-azido-8-deoxy-Kdo (8-N(3)-Kdo) was then greatly improved over previous reports via optimization of the synthesis of its 5-azido-5-deoxy-d-arabinose precursor to yield gram amounts. The novel analogue 7-azido-7-deoxy-Kdo (7-N(3)-Kdo) was also synthesized, with both analogues capable of undergoing in vitro strain-promoted azide–alkyne cycloaddition (SPAAC) “click” chemistry reactions. Slower and faster growth of M. xanthus was displayed in the presence of 8-N(3)-Kdo and 7-N(3)-Kdo (respectively) compared to untreated cells, with differences also seen for single-cell gliding motility and type IV pilus-dependent swarm community expansion. While the surfaces of 8-N(3)-Kdo-grown cells were fluorescently labeled following treatment with dibenzocyclooctyne-linked fluorophores, the surfaces of 7-N(3)-Kdo-grown cells could not undergo fluorescent tagging. Activity analysis of the KdsB enzyme required to activate Kdo prior to its integration into nascent LPS molecules revealed that while 8-N(3)-Kdo is indeed a substrate of the enzyme, 7-N(3)-Kdo is not. Though a lack of M. xanthus cell aggregation was shown to expedite growth in liquid culture, 7-N(3)-Kdo-grown cells did not manifest differences in intrinsic clumping relative to untreated cells, suggesting that 7-N(3)-Kdo may instead be catabolized by the cells. Ultimately, these data provide important insights into the synthesis and cellular processing of valuable metabolic labels and establish a basis for the elucidation of fundamental principles of OM dynamism in live bacterial cells

From Spiking Neuron Models to Linear-Nonlinear Models

Neurons transform time-varying inputs into action potentials emitted stochastically at a time dependent rate. The mapping from current input to output firing rate is often represented with the help of phenomenological models such as the linear-nonlinear (LN) cascade, in which the output firing rate is estimated by applying to the input successively a linear temporal filter and a static non-linear transformation. These simplified models leave out the biophysical details of action potential generation. It is not a priori clear to which extent the input-output mapping of biophysically more realistic, spiking neuron models can be reduced to a simple linear-nonlinear cascade. Here we investigate this question for the leaky integrate-and-fire (LIF), exponential integrate-and-fire (EIF) and conductance-based Wang-Buzsáki models in presence of background synaptic activity. We exploit available analytic results for these models to determine the corresponding linear filter and static non-linearity in a parameter-free form. We show that the obtained functions are identical to the linear filter and static non-linearity determined using standard reverse correlation analysis. We then quantitatively compare the output of the corresponding linear-nonlinear cascade with numerical simulations of spiking neurons, systematically varying the parameters of input signal and background noise. We find that the LN cascade provides accurate estimates of the firing rates of spiking neurons in most of parameter space. For the EIF and Wang-Buzsáki models, we show that the LN cascade can be reduced to a firing rate model, the timescale of which we determine analytically. Finally we introduce an adaptive timescale rate model in which the timescale of the linear filter depends on the instantaneous firing rate. This model leads to highly accurate estimates of instantaneous firing rates

Meiotic Recombination Hotspots of Fission Yeast Are Directed to Loci that Express Non-Coding RNA

Polyadenylated, mRNA-like transcripts with no coding potential are abundant in eukaryotes, but the functions of these long non-coding RNAs (ncRNAs) are enigmatic. In meiosis, Rec12 (Spo11) catalyzes the formation of dsDNA breaks (DSBs) that initiate homologous recombination. Most meiotic recombination is positioned at hotspots, but knowledge of the mechanisms is nebulous. In the fission yeast genome DSBs are located within 194 prominent peaks separated on average by 65-kbp intervals of DNA that are largely free of DSBs.). Furthermore, we tested and rejected the hypothesis that the ncRNA loci and DSB peaks localize preferentially, but independently, to a third entity on the chromosomes.Meiotic DSB hotspots are directed to loci that express polyadenylated ncRNAs. This reveals an unexpected, possibly unitary mechanism for what directs meiotic recombination to hotspots. It also reveals a likely biological function for enigmatic ncRNAs. We propose specific mechanisms by which ncRNA molecules, or some aspect of RNA metabolism associated with ncRNA loci, help to position recombination protein complexes at DSB hotspots within chromosomes

Spike-Based Reinforcement Learning in Continuous State and Action Space: When Policy Gradient Methods Fail

Changes of synaptic connections between neurons are thought to be the physiological basis of learning. These changes can be gated by neuromodulators that encode the presence of reward. We study a family of reward-modulated synaptic learning rules for spiking neurons on a learning task in continuous space inspired by the Morris Water maze. The synaptic update rule modifies the release probability of synaptic transmission and depends on the timing of presynaptic spike arrival, postsynaptic action potentials, as well as the membrane potential of the postsynaptic neuron. The family of learning rules includes an optimal rule derived from policy gradient methods as well as reward modulated Hebbian learning. The synaptic update rule is implemented in a population of spiking neurons using a network architecture that combines feedforward input with lateral connections. Actions are represented by a population of hypothetical action cells with strong mexican-hat connectivity and are read out at theta frequency. We show that in this architecture, a standard policy gradient rule fails to solve the Morris watermaze task, whereas a variant with a Hebbian bias can learn the task within 20 trials, consistent with experiments. This result does not depend on implementation details such as the size of the neuronal populations. Our theoretical approach shows how learning new behaviors can be linked to reward-modulated plasticity at the level of single synapses and makes predictions about the voltage and spike-timing dependence of synaptic plasticity and the influence of neuromodulators such as dopamine. It is an important step towards connecting formal theories of reinforcement learning with neuronal and synaptic properties

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Collision kinematics in the western external Alps

International audienceThe kinematics of the collision in Western Alps are investigated through five balanced cross sections of the whole external domain from the Oisans to the Mont Blanc massif. These cross sections were built using published data for the Jura and subalpine fold-and-thrust belts and new structural and field analysis for the External Crystalline Massifs. Five units are defined: the sedimentary nappes from innermost parts of the external zone (e.g., ultra-Dauphinois/Helvetic), the crystalline units with their dysharmonically folded cover (e.g., Morcles nappe), sedimentary nappes over the frontal parts of the crystalline massifs (the Aravis-Granier unit), the subalpine belts (e.g., Vercors, Chartreuse, Bauges, and Bornes), and the Jura. Except for the ultra-Dauphinois nappes, the shortening, including the cover shortening, always corresponds to basement shortening. The total amount of shortening increases from south (28 km, 20%) to north (66 km, 27%). Moreover, the shortening is slightly older in the south than in the north; deepwater turbidites (flysch) and shallow marine to freshwater clastics (molasse) basins are more developed in the north; pressure and temperature conditions are higher in the north; the average uplift rates are about 3 times higher in the north and more localized in space. We propose that these differences are due to along-strike variations in the structure of the European continental margin inherited from Mesozoic times. We then build five palinspastic maps: one at Cretaceous times showing the inherited European Mesozoic margin structure and four from Priabonian to upper Miocene times showing the collision kinematics and the related rotation of Adria

EFIDIR : extraction et fusion d'informations pour la mesure de déplacements par imagerie radar

National audienceL'imagerie radar satellitaire permet de mesurer le déplacement du sol avec une pré- cision centimétrique, voire millimétrique à l'aide de séries d'images SAR (Synthetic Aperture Radar). Ces précisions ne peuvent être atteintes qu'en exploitant la masse de données radar disponibles ou le potentiel de satellites lancés récemment : images haute résolution (métrique) ou polarimétriques. Cet article présente les principaux résultats obtenus par le projet ANR EFIDIR (extraction et fusion d'informations pour la mesure de déplacement en imagerie radar, 2008-2011) qui rassemble des spécialistes de l'imagerie radar, du traitement de l'in- formation et des géosciences. Un radar imageur de terrain a été mis au point et des méthodes de traitement spécifiques ont été développées (tomographie radar, suréchantillonnage d'ima- ges complexes...) afin d'améliorer la caractérisation de la diffusion électromagnétique et les prétraitements des données SAR. Les méthodes d'extraction d'informations de déplacement à partir de l'amplitude (texture tracking) ou de la phase (interférométrie multitemporelle) ont été approfondies et testées sur les grands déplacements de glaciers, ou de faibles déformations intersismiques. Une approche originale de fouille de données a également été développée pour extraire les motifs séquentiels fréquents et appliquée avec succès à la détection de motifs liés à la déformation du sol noyés dans des artéfacts atmosphériques. Enfin une approche nouvel- le de représentation des incertitudes par la théorie des possibilités a été introduite dans l'éta- pe d'inversion qui permet de reconstruire des champs de déplacement 3D (Est, Nord, vertical) ou d'estimer les paramètres de modèles géophysiques des phénomènes à l'origine des dépla- cements observés en surface. Ces travaux illustrent ainsi les grandes étapes des chaînes de traitement permettant d'aller de la formation des données SAR à la modélisation géophysique

Evaluation of Azido 3‑Deoxy‑d-<i>manno</i>-oct-2-ulosonic Acid (Kdo) Analogues for Click Chemistry-Mediated Metabolic Labeling of Myxococcus xanthus DZ2 Lipopolysaccharide

Metabolic labeling paired with click chemistry is a powerful approach for selectively imaging the surfaces of diverse bacteria. Herein, we explored the feasibility of labeling the lipopolysaccharide (LPS) of Myxococcus xanthusa Gram-negative predatory social bacterium known to display complex outer membrane (OM) dynamicsvia growth in the presence of distinct azido (-N3) analogues of 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo). Determination of the LPS carbohydrate structure from strain DZ2 revealed the presence of one Kdo sugar in the core oligosaccharide, modified with phosphoethanolamine. The production of 8-azido-8-deoxy-Kdo (8-N3-Kdo) was then greatly improved over previous reports via optimization of the synthesis of its 5-azido-5-deoxy-d-arabinose precursor to yield gram amounts. The novel analogue 7-azido-7-deoxy-Kdo (7-N3-Kdo) was also synthesized, with both analogues capable of undergoing in vitro strain-promoted azide–alkyne cycloaddition (SPAAC) “click” chemistry reactions. Slower and faster growth of M. xanthus was displayed in the presence of 8-N3-Kdo and 7-N3-Kdo (respectively) compared to untreated cells, with differences also seen for single-cell gliding motility and type IV pilus-dependent swarm community expansion. While the surfaces of 8-N3-Kdo-grown cells were fluorescently labeled following treatment with dibenzocyclooctyne-linked fluorophores, the surfaces of 7-N3-Kdo-grown cells could not undergo fluorescent tagging. Activity analysis of the KdsB enzyme required to activate Kdo prior to its integration into nascent LPS molecules revealed that while 8-N3-Kdo is indeed a substrate of the enzyme, 7-N3-Kdo is not. Though a lack of M. xanthus cell aggregation was shown to expedite growth in liquid culture, 7-N3-Kdo-grown cells did not manifest differences in intrinsic clumping relative to untreated cells, suggesting that 7-N3-Kdo may instead be catabolized by the cells. Ultimately, these data provide important insights into the synthesis and cellular processing of valuable metabolic labels and establish a basis for the elucidation of fundamental principles of OM dynamism in live bacterial cells