Página 7 dos resultados de 9995 itens digitais encontrados em 0.058 segundos

‣ Significance analysis of time course microarray experiments

Storey, John D.; Xiao, Wenzhong; Leek, Jeffrey T.; Tompkins, Ronald G.; Davis, Ronald W.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Characterizing the genome-wide dynamic regulation of gene expression is important and will be of much interest in the future. However, there is currently no established method for identifying differentially expressed genes in a time course study. Here we propose a significance method for analyzing time course microarray studies that can be applied to the typical types of comparisons and sampling schemes. This method is applied to two studies on humans. In one study, genes are identified that show differential expression over time in response to in vivo endotoxin administration. By using our method, 7,409 genes are called significant at a 1% false-discovery rate level, whereas several existing approaches fail to identify any genes. In another study, 417 genes are identified at a 10% false-discovery rate level that show expression changing with age in the kidney cortex. Here it is also shown that as many as 47% of the genes change with age in a manner more complex than simple exponential growth or decay. The methodology proposed here has been implemented in the freely distributed and open-source edge software package.

‣ Coupled protein domain motion in Taq polymerase revealed by neutron spin-echo spectroscopy

Bu, Zimei; Biehl, Ralf; Monkenbusch, Michael; Richter, Dieter; Callaway, David J. E.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Long-range conformational changes in proteins are ubiquitous in biology for the transmission and amplification of signals; such conformational changes can be triggered by small-amplitude, nanosecond protein domain motion. Understanding how conformational changes are initiated requires the characterization of protein domain motion on these timescales and on length scales comparable to protein dimensions. Using neutron spin-echo spectroscopy (NSE), normal mode analysis, and a statistical-mechanical framework, we reveal overdamped, coupled domain motion within DNA polymerase I from Thermus aquaticus (Taq polymerase). This protein utilizes correlated domain dynamics over 70 Å to coordinate nucleotide synthesis and cleavage during DNA synthesis and repair. We show that NSE spectroscopy can determine the domain mobility tensor, which determines the degree of dynamical coupling between domains. The mobility tensor defines the domain velocity response to a force applied to it or to another domain, just as the sails of a sailboat determine its velocity given the applied wind force. The NSE results provide insights into the nature of protein domain motion that are not appreciated by conventional biophysical techniques.

‣ Global asymptotic coherence in discrete dynamical systems

Earn, David J. D.; Levin, Simon A.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Spatial synchrony (coherence) in dynamical systems is of both theoretical and applied importance. We address this problem for a generalization of coupled map lattices (CMLs). In the systems we study, which we term “meta-CMLs,” the map at each lattice point may be multidimensional (corresponding, for example, to multispecies ecological systems in which all species have the same dispersal pattern). Most previous work on coherence of CMLs has focused on local stability. Here, we prove a global theorem that provides a useful sufficient condition guaranteeing decay of incoherence in meta-CMLs regardless of initial conditions and regardless of the nature of the attractors of the system. This result facilitates useful analyses of a variety of applied problems, including conservation of endangered species and eradication of pests or infectious diseases.

‣ Induction of tolerance in arthritogenic B cells with receptors of differing affinity for self-antigen

Huang, Haochu; Kearney, John F.; Grusby, Michael J.; Benoist, Christophe; Mathis, Diane
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Multiple mechanisms of tolerance induction limit autoimmunity, but their relative contribution for lymphocytes recognizing self-antigens of differing availability is incompletely understood. The mechanisms applied to arthritogenic B cells expressing antigen-specific B cell receptors (BCRs) with different affinities for glucose-6-phosphate isomerase (GPI) were examined in the corresponding Ig gene knock-in mice. This ubiquitously expressed and blood-borne enzyme is the target autoantigen in the K/BxN model of inflammatory arthritis and perhaps in some humans with arthritis. Negative selection of B cells expressing high-affinity anti-GPI specificities, whose surface receptors were occupied by GPI, operated mainly at the transitional B cell stages in the spleen, preventing their final differentiation and entry into follicular areas. Receptor editing contributed to the purging of cells displaying anti-GPI BCRs, and significant numbers of autoreactive cells escaped through expression of an additional Ig light (L) chain, accumulating gradually in lymphoid organs. In contrast, low-affinity anti-GPI B cells, whose surface receptors did not carry GPI, matured normally. The “escaped” dual-L-chain cells and the “ignored” low-affinity cells are the likely precursors of cells that produce pathogenic autoantibodies once T cell help is provided. These studies portray...

‣ Mechanisms of chromosome number reduction in Arabidopsis thaliana and related Brassicaceae species

Lysak, Martin A.; Berr, Alexandre; Pecinka, Ales; Schmidt, Renate; McBreen, Kim; Schubert, Ingo
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Evolution of chromosome complements can be resolved by genome sequencing, comparative genetic mapping, and comparative chromosome painting. Previously, comparison of genetic maps and gene-based phylogenies suggested that the karyotypes of Arabidopsis thaliana (n = 5) and of related species with six or seven chromosome pairs were derived from an ancestral karyotype with eight chromosome pairs. To test this hypothesis, we applied multicolor chromosome painting using contiguous bacterial artificial chromosome pools of A. thaliana arranged according to the genetic maps of Arabidopsis lyrata and Capsella rubella (both n = 8) to A. thaliana, A. lyrata, Neslia paniculata, Turritis glabra, and Hornungia alpina. This approach allowed us to map the A. lyrata centromeres as a prerequisite to defining a putative ancestral karyotype (n = 8) and to elucidate the evolutionary mechanisms that shaped the karyotype of A. thaliana and its relatives. We conclude that chromosome “fusions” in A. thaliana resulted from (i) generation of acrocentric chromosomes by pericentric inversions, (ii) reciprocal translocation between two chromosomes (one or both acrocentric), and (iii) elimination of a minichromosome that arose in addition to the “fusion chromosome.” Comparative chromosome painting applied to N. paniculata (n = 7)...

‣ Functional, fractal nonlinear response with application to rate processes with memory, allometry, and population genetics

Vlad, Marcel O.; Morán, Federico; Popa, Vlad T.; Szedlacsek, Stefan E.; Ross, John
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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We give a functional generalization of fractal scaling laws applied to response problems as well as to probability distributions. We consider excitations and responses, which are functions of a given state vector. Based on scaling arguments, we derive a general nonlinear response functional scaling law, which expresses the logarithm of a response at a given state as a superposition of the values of the logarithms of the excitations at different states. Such a functional response law may result from the balance of different growth processes, characterized by variable growth rates, and it is the first order approximation of a perturbation expansion similar to the phase expansion. Our response law is a generalization of the static fractal scaling law and can be applied to the study of various problems from physics, chemistry, and biology. We consider some applications to heterogeneous and disordered kinetics, organ growth (allometry), and population genetics. Kinetics on inhomogeneous reconstructing surfaces leads to rate equations described by our nonlinear scaling law. For systems with dynamic disorder with random energy barriers, the probability density functional of the rate coefficient is also given by our scaling law. The relative growth rates of different biological organs (allometry) can be described by a similar approach. Our scaling law also emerges by studying the variation of macroscopic phenotypic variables in terms of genotypic growth rates. We study the implications of the causality principle for our theory and derive a set of generalized Kramers–Kronig relationships for the fractal scaling exponents.

‣ Thermodynamic constraints on stochastic acceleration in compressional turbulence

Fisk, Lennard A.; Gloeckler, George
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Recent observations in the solar wind have revealed an important phenomenon. In circumstances where stochastic acceleration is expected, a suprathermal tail on the distribution function is formed with a common spectral shape: the spectrum is a power law in particle speed with a spectral index of −5. This common spectrum occurs in the quiet solar wind; in disturbed conditions downstream from shocks; and, in particular, throughout the heliosheath downstream from the termination shock of the solar wind currently being explored by Voyager 1. In this article, simple thermodynamic principles are applied to stochastic acceleration in compressional turbulence. The unique spectral index results when the entropy of the suprathermal tail has increased to the maximum allowable value. Relationships for the pressure in the suprathermal tail are also derived and found to be in agreement with observations. The results are shown to be consistent with the suprathermal tail arising from a cascade in energy, analogous to a turbulent cascade. The results may be applied broadly, because stochastic acceleration in compressional turbulence should be common in many astrophysical settings.

‣ Microarray analysis of newly synthesized RNA in cells and animals

Kenzelmann, M.; Maertens, S.; Hergenhahn, M.; Kueffer, S.; Hotz-Wagenblatt, A.; Li, L.; Wang, S.; Ittrich, C.; Lemberger, T.; Arribas, R.; Jonnakuty, S.; Hollstein, M. C.; Schmid, W.; Gretz, N.; Gröne, H. J.; Schütz, G.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Current methods to analyze gene expression measure steady-state levels of mRNA. To specifically analyze mRNA transcription, we have developed a technique that can be applied in vivo in intact cells and animals. Our method makes use of the cellular pyrimidine salvage pathway and is based on affinity-chromatographic isolation of thiolated mRNA. When combined with data on mRNA steady-state levels, this method is able to assess the relative contributions of mRNA synthesis and degradation/stabilization. It overcomes limitations associated with currently available methods such as mechanistic intervention that disrupts cellular physiology, or the inability to apply the techniques in vivo. Our method was first tested in serum response of cultured fibroblast cells and then applied to the study of renal ischemia reperfusion injury, demonstrating its applicability for whole organs in vivo. Combined with data on mRNA steady-state levels, this method provided a detailed analysis of regulatory mechanisms of mRNA expression and the relative contributions of RNA synthesis and turnover within distinct pathways, and identification of genes expressed at low abundance at the transcriptional level.

‣ Genetic diversity of citrus bacterial canker pathogens preserved in herbarium specimens

Li, Wenbin; Song, Qijian; Brlansky, Ronald H.; Hartung, John S.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Citrus bacterial canker (CBC) caused by Xanthomonas axonopodis pv. citri (Xac) was first documented in India and Java in the mid 19th century. Since that time, the known distribution of the disease has steadily increased. Concurrent with the dispersion of the pathogen, the diversity of described strains continues to increase, with novel strains appearing in Saudi Arabia, Iran, and Florida in the last decade. Herbarium specimens of infected plants provide an historical record documenting both the geographic distribution and genetic diversity of the pathogen in the past. However, no method was available to assess the genetic diversity within these herbarium samples. We have developed a method, insertion event scanning (IES), and applied the method to characterize the diversity present within CBC populations documented as herbarium specimens over the past century. IES is based on the specific amplification of junction fragments that define insertion events. The potential for IES in current forensic applications is demonstrated by finding an exact match of pathogen genotypes preserved in herbarium specimens from Japan and Florida, demonstrating the source of the original outbreak of citrus canker in Florida in 1911. IES is a very sensitive technique for differentiating bacterial strains and can be applied to any of the several hundred bacteria for which full genomic sequence data are available.

‣ Pharmacological disruption of calcium channel trafficking by the α2δ ligand gabapentin

Hendrich, Jan; Van Minh, Alexandra Tran; Heblich, Fay; Nieto-Rostro, Manuela; Watschinger, Katrin; Striessnig, Jörg; Wratten, Jack; Davies, Anthony; Dolphin, Annette C.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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The mechanism of action of the antiepileptic and antinociceptive drugs of the gabapentinoid family has remained poorly understood. Gabapentin (GBP) binds to an exofacial epitope of the α2δ-1 and α2δ-2 auxiliary subunits of voltage-gated calcium channels, but acute inhibition of calcium currents by GBP is either very minor or absent. We formulated the hypothesis that GBP impairs the ability of α2δ subunits to enhance voltage-gated Ca2+channel plasma membrane density by means of an effect on trafficking. Our results conclusively demonstrate that GBP inhibits calcium currents, mimicking a lack of α2δ only when applied chronically, but not acutely, both in heterologous expression systems and in dorsal root-ganglion neurons. GBP acts primarily at an intracellular location, requiring uptake, because the effect of chronically applied GBP is blocked by an inhibitor of the system-L neutral amino acid transporters and enhanced by coexpression of a transporter. However, it is mediated by α2δ subunits, being prevented by mutations in either α2δ-1 or α2δ-2 that abolish GBP binding, and is not observed for α2δ-3, which does not bind GBP. Furthermore, the trafficking of α2δ-2 and CaV2 channels is disrupted both by GBP and by the mutation in α2δ-2...

‣ A modified cysteinyl-labeling assay reveals reversible oxidation of protein tyrosine phosphatases in angiomyolipoma cells

Boivin, Benoit; Zhang, Sheng; Arbiser, Jack L.; Zhang, Zhong-Yin; Tonks, Nicholas K.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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The production of reactive oxygen species (ROS) exerts an additional tier of control over tyrosine phosphorylation-dependent signal transduction by transiently inhibiting the catalytic activity of specific protein tyrosine phosphatases (PTPs). Hence, the ability to detect reversible oxidation of PTPs in vivo is critical to understanding the complex biological role of ROS in the control of cellular signaling. Here, we describe an assay for identifying those PTPs that are reversibly oxidized in vivo, which utilizes the unique chemistry of the invariant catalytic Cys residue in labeling the active site with biotinylated small molecules under mildly acidic conditions. We have applied this cysteinyl-labeling assay to the study of platelet-derived growth factor (PDGF) receptor signaling in an angiomyolipoma cell model. Doing so has allowed us to detect reversible oxidation of several proteins in response to sustained PDGF stimulation. As in other cell systems, we have observed the reversible oxidation of the classical PTP SHP2 and the tumor suppressor phosphatase PTEN in response to PDGF stimulation. Furthermore, we detected reversible oxidation of members of two other subclasses of PTPs, the receptor PTP LAR and the dual-specificity phosphatase MKP1. These data demonstrate the broad selectivity of the assay...

‣ Giant Stark effect in quantum dots at liquid/liquid interfaces: A new option for tunable optical filters

Flatté, M. E.; Kornyshev, A. A.; Urbakh, M.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Control of the fundamental absorption edge of a quantum dot with an applied electric field has been limited by the breakdown fields of the solid-state material surrounding the dot. However, much larger fields can be applied at the interface of two immiscible electrolytic solutions (ITIES) in an electrochemical cell. These electric fields also localize the quantum dots at the ITIES. Our analysis shows that semiconductor nanocrystals localized at the ITIES should have electric-field-tunable optical properties across much of the visible spectrum. The transparency of the liquids in such cells indicates that this configuration would be well suited for electrically tunable optical filters with wide-angle acceptance.

‣ Transmission of allostery through the lectin domain in selectin-mediated cell adhesion

Waldron, Travis T.; Springer, Timothy A.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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The selectins are cell adhesion proteins that must resist applied forces to mediate leukocyte tethering and rolling along the endothelium and have 2 conformational states. Selectin–ligand bond dissociation increases only modestly with applied force, and exhibits catch bond behavior in a low-force regime where bond lifetimes counterintuitively increase with increasing force. Both allosteric and sliding–rebinding models have emerged to explain catch bonds. Here, we introduce a large residue into a cleft that opens within the lectin domain to stabilize the more extended, high-affinity selectin conformation. This mutation stabilizes the high-affinity state, but surprisingly makes rolling less stable. The position of the mutation in the lectin domain provides evidence for an allosteric pathway through the lectin domain, connecting changes at the lectin–EGF interface to the distal binding interface.

‣ A synthesized pheromone induces upstream movement in female sea lamprey and summons them into traps

Johnson, Nicholas S.; Yun, Sang-Seon; Thompson, Henry T.; Brant, Cory O.; Li, Weiming
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Female insect pheromone blends induce robust tracking responses in males and direct them into traps. In vertebrates, pheromones that induce strong and precise tracking responses in natural habitats have rarely been described. Here, we show in the sea lamprey (Petromyzon marinus), a vertebrate invader of the Laurential Great Lakes, that a synthesized component of the male mating pheromone, 7α, 12α, 24-trihydroxy-5α-cholan-3-one 24-sulfate (3kPZS), when released into a stream to reach concentrations of 10−14, 10−13, 10−12, 10−11, or 10−10 M, triggers robust upstream movement in ovulated females drawing ≈50% into baited traps. Experiments conducted in diverse stream segments demonstrate the level of behavioral response was not affected by habitat conditions and is effective over hundreds of meters. 3kPZS is equally effective at luring ovulated females as the whole pheromone blend released by males between 10−14 and 10−11 M. 3kPZS diverts ovulated females away from and disrupts orientation to male washings when applied at concentrations higher than washings. Indeed, a single pheromone compound is able to redirect female sea lampreys away from a natural pheromone source and lure them into traps, which should be more effective than targeting males when applied in population control. Our findings may spur the discovery of other potent and environmentally benign agents to combat biological invasion...

‣ Normal forms for reduced stochastic climate models

Majda, Andrew J.; Franzke, Christian; Crommelin, Daan
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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The systematic development of reduced low-dimensional stochastic climate models from observations or comprehensive high-dimensional climate models is an important topic for atmospheric low-frequency variability, climate sensitivity, and improved extended range forecasting. Here techniques from applied mathematics are utilized to systematically derive normal forms for reduced stochastic climate models for low-frequency variables. The use of a few Empirical Orthogonal Functions (EOFs) (also known as Principal Component Analysis, Karhunen–Loéve and Proper Orthogonal Decomposition) depending on observational data to span the low-frequency subspace requires the assessment of dyad interactions besides the more familiar triads in the interaction between the low- and high-frequency subspaces of the dynamics. It is shown below that the dyad and multiplicative triad interactions combine with the climatological linear operator interactions to simultaneously produce both strong nonlinear dissipation and Correlated Additive and Multiplicative (CAM) stochastic noise. For a single low-frequency variable the dyad interactions and climatological linear operator alone produce a normal form with CAM noise from advection of the large scales by the small scales and simultaneously strong cubic damping. These normal forms should prove useful for developing systematic strategies for the estimation of stochastic models from climate data. As an illustrative example the one-dimensional normal form is applied below to low-frequency patterns such as the North Atlantic Oscillation (NAO) in a climate model. The results here also illustrate the short comings of a recent linear scalar CAM noise model proposed elsewhere for low-frequency variability.

‣ Population genomic inference of recombination rates and hotspots

Wang, Ying; Rannala, Bruce
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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As more human genomic data become available, fine-scale recombination rate variation can be inferred on a genome-wide scale. Current statistical methods to infer recombination rates that can be applied to moderate, or large, genomic regions are limited to approximated likelihoods. Here, we develop a Bayesian full-likelihood method using Markov Chain Monte Carlo (MCMC) to estimate background recombination rates and hotspots. The probability model is inspired by the observed patterns of recombination at several genomic regions analyzed in sperm-typing studies. Posterior probabilities and Bayes factors of recombination hotspots along chromosomes are inferred. For moderate-size genomic regions (e.g., with <100 SNPs), the full-likelihood method is used. Larger regions are split into subintervals (typically each having between 20 and 50 markers). The likelihood is approximated based on the genealogies for each subinterval. The background recombination rates, hotspots, and parameters are evaluated by using a parallel computing approach and assuming shared parameters across the subintervals. Simulation analyses show that our method can accurately estimate the variation in recombination rates across genomic regions. In particular, clusters of hotspots can be distinguished even though weaker hotspots are present. The method is applied to SNP data from the HLA region...

‣ A consensus genetic map of cowpea [Vigna unguiculata (L) Walp.] and synteny based on EST-derived SNPs

Muchero, Wellington; Diop, Ndeye N.; Bhat, Prasanna R.; Fenton, Raymond D.; Wanamaker, Steve; Pottorff, Marti; Hearne, Sarah; Cisse, Ndiaga; Fatokun, Christian; Ehlers, Jeffrey D.; Roberts, Philip A.; Close, Timothy J.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Consensus genetic linkage maps provide a genomic framework for quantitative trait loci identification, map-based cloning, assessment of genetic diversity, association mapping, and applied breeding in marker-assisted selection schemes. Among “orphan crops” with limited genomic resources such as cowpea [Vigna unguiculata (L.) Walp.] (2n = 2x = 22), the use of transcript-derived SNPs in genetic maps provides opportunities for automated genotyping and estimation of genome structure based on synteny analysis. Here, we report the development and validation of a high-throughput EST-derived SNP assay for cowpea, its application in consensus map building, and determination of synteny to reference genomes. SNP mining from 183,118 ESTs sequenced from 17 cDNA libraries yielded ≈10,000 high-confidence SNPs from which an Illumina 1,536-SNP GoldenGate genotyping array was developed and applied to 741 recombinant inbred lines from six mapping populations. Approximately 90% of the SNPs were technically successful, providing 1,375 dependable markers. Of these, 928 were incorporated into a consensus genetic map spanning 680 cM with 11 linkage groups and an average marker distance of 0.73 cM. Comparison of this cowpea genetic map to reference legumes...

‣ Plant DNA barcodes and a community phylogeny of a tropical forest dynamics plot in Panama

Kress, W. John; Erickson, David L.; Jones, F. Andrew; Swenson, Nathan G.; Perez, Rolando; Sanjur, Oris; Bermingham, Eldredge
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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The assembly of DNA barcode libraries is particularly relevant within species-rich natural communities for which accurate species identifications will enable detailed ecological forensic studies. In addition, well-resolved molecular phylogenies derived from these DNA barcode sequences have the potential to improve investigations of the mechanisms underlying community assembly and functional trait evolution. To date, no studies have effectively applied DNA barcodes sensu strictu in this manner. In this report, we demonstrate that a three-locus DNA barcode when applied to 296 species of woody trees, shrubs, and palms found within the 50-ha Forest Dynamics Plot on Barro Colorado Island (BCI), Panama, resulted in >98% correct identifications. These DNA barcode sequences are also used to reconstruct a robust community phylogeny employing a supermatrix method for 281 of the 296 plant species in the plot. The three-locus barcode data were sufficient to reliably reconstruct evolutionary relationships among the plant taxa in the plot that are congruent with the broadly accepted phylogeny of flowering plants (APG II). Earlier work on the phylogenetic structure of the BCI forest dynamics plot employing less resolved phylogenies reveals significant differences in evolutionary and ecological inferences compared with our data and suggests that unresolved community phylogenies may have increased type I and type II errors. These results illustrate how highly resolved phylogenies based on DNA barcode sequence data will enhance research focused on the interface between community ecology and evolution.

‣ Constructing the equilibrium ensemble of folding pathways from short off-equilibrium simulations

Noé, Frank; Schütte, Christof; Vanden-Eijnden, Eric; Reich, Lothar; Weikl, Thomas R.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Characterizing the equilibrium ensemble of folding pathways, including their relative probability, is one of the major challenges in protein folding theory today. Although this information is in principle accessible via all-atom molecular dynamics simulations, it is difficult to compute in practice because protein folding is a rare event and the affordable simulation length is typically not sufficient to observe an appreciable number of folding events, unless very simplified protein models are used. Here we present an approach that allows for the reconstruction of the full ensemble of folding pathways from simulations that are much shorter than the folding time. This approach can be applied to all-atom protein simulations in explicit solvent. It does not use a predefined reaction coordinate but is based on partitioning the state space into small conformational states and constructing a Markov model between them. A theory is presented that allows for the extraction of the full ensemble of transition pathways from the unfolded to the folded configurations. The approach is applied to the folding of a PinWW domain in explicit solvent where the folding time is two orders of magnitude larger than the length of individual simulations. The results are in good agreement with kinetic experimental data and give detailed insights about the nature of the folding process which is shown to be surprisingly complex and parallel. The analysis reveals the existence of misfolded trap states outside the network of efficient folding intermediates that significantly reduce the folding speed.

‣ The selective antagonist EPPTB reveals TAAR1-mediated regulatory mechanisms in dopaminergic neurons of the mesolimbic system

Bradaia, Amyaouch; Trube, Gerhard; Stalder, Henri; Norcross, Roger D.; Ozmen, Laurence; Wettstein, Joseph G.; Pinard, Audrée; Buchy, Danièle; Gassmann, Martin; Hoener, Marius C.; Bettler, Bernhard
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
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Trace amine-associated receptor 1 (TAAR1) is a G protein-coupled receptor (GPCR) that is nonselectively activated by endogenous metabolites of amino acids. TAAR1 is considered a promising drug target for the treatment of psychiatric and neurodegenerative disorders. However, no selective ligand to identify TAAR1-specific signaling mechanisms is available yet. Here we report a selective TAAR1 antagonist, EPPTB, and characterize its physiological effects at dopamine (DA) neurons of the ventral tegmental area (VTA). We show that EPPTB prevents the reduction of the firing frequency of DA neurons induced by p-tyramine (p-tyr), a nonselective TAAR1 agonist. When applied alone, EPPTB increases the firing frequency of DA neurons, suggesting that TAAR1 either exhibits constitutive activity or is tonically activated by ambient levels of endogenous agonist(s). We further show that EPPTB blocks the TAAR1-mediated activation of an inwardly rectifying K+ current. When applied alone, EPPTB induces an apparent inward current, suggesting the closure of tonically activated K+ channels. Importantly, these EPPTB effects were absent in Taar1 knockout mice, ruling out off-target effects. We additionally found that both the acute application of EPPTB and the constitutive genetic lack of TAAR1 increase the potency of DA at D2 receptors in DA neurons. In summary...