Proteome-wide measurement of non-canonical bacterial mistranslation by quantitative mass spectrometry of protein modifications.

The genetic code is virtually universal in biology and was likely established before the advent of cellular life. The extent to which mistranslation occurs is poorly understood and presents a fundamental question in basic research and production of recombinant proteins. Here we used shotgun proteomics combined with unbiased protein modification analysis to quantitatively analyze in vivo mistranslation in an E. coli strain with a defect in the editing mechanism of leucyl-tRNA synthetase. We detected the misincorporation of a non-proteinogenic amino acid norvaline on 10% of all measured leucine residues under microaerobic conditions and revealed preferential deployment of a tRNA(Leu)(CAG) isoacceptor during norvaline misincorporation. The strain with the norvalylated proteome demonstrated a substantial reduction in cell fitness under both prolonged aerobic and microaerobic cultivation. Unlike norvaline, isoleucine did not substitute for leucine even under harsh error-prone conditions. Our study introduces shotgun proteomics as a powerful tool in quantitative analysis of mistranslation

Impact of cooking method on phenolic composition and antioxidant potential of four varieties of Phaseolus vulgaris L. and Glycine max L.

The present study aimed to investigate the effects of different cooking conditions - atmospheric (100°C) and pressure cooking (115°C) - on the phenolic composition and antioxidant activity of methanolic extracts of four Phaseolus vulgaris varieties and soy (Glycine max). Contrary to soy, in P. vulgaris varieties both cooking methods increased drastically the total phenolic, flavonoid, and ortho-diphenol content, as well as antioxidant capacity. These results were corroborated by HPLC analysis, where an overall increase of phenolic acids and flavonoids was detected in processed samples. However, draining the cooking water significantly decreased phenolic acids, flavonoids and antioxidant activity in all P. vulgaris varieties and as well as soy. The hypothesis that cooking increases the compound accessibility and nutritional value through increased release of phytochemicals was verified in the present study for P. vulgaris varieties. Keeping the cooking water is crucial to the increased nutritional value of all Phaseolus varieties. Overall, compared with the tested varieties of Phaseolus, soy, to which many health benefits are attributed, is not the best legume source of antioxidants.The author Catarina I. Teixeira-Guedes acknowledges the financial support provided FCT-Portuguese Foundation for Science and Technology (SFRH/BD/52544/2014), under the Doctoral Program “Agricultural Production Chains – from fork to farm” (PD/00122/ 2012). This work was supported by the European Investment Funds FEDER/COMPETE/POCI– Operational Competitiveness and Internationalization Programme [POCI-01-0145-FEDER-006958] and Portuguese Foundation for Science and Technology (FCT) [UID/AGR/ 04033/2019]; Project I&D Interact - Integrative Research in Environment, Agro-Chain and Technology [NORTE-01-0145-FEDER000017], co-financed by European Regional Development Fund (FEDER) through NORTE 2020 (Programa Operacional Regional do Norte 2014/2020).info:eu-repo/semantics/publishedVersio

A Dynamic Contextual Change Management Application for Real Time Decision-Making Support

Decision making is a fundamental process within organizations for many reasons. It is indeed involved at all levels (new product decisions, management and marketing decisions, etc.) and has a direct impact on companies’ efficiency and effectiveness. Many researches are conducted to enhance the decision-making process by proposing decision support systems where the most frequent challenge is the change management. Indeed, all businesses operate within an environment that is subject to constant changes (like new customers’ needs and requirements, organisational and technological changes, changes in key information used to derive decisions, etc.). These changes have a major impact on the quality and accuracy of the proposed decision if they are not detected and propagated, at the right time, during the decision-making process. The present work attempts to resolve this challenge by proposing a dynamic change management technique that allows three tasks to be automatically performed. First, continuously detect changes and note them. Second, retrieve from the detected changes those that are related to the decision rules. Finally, propagate them by computing the new value of the decision rule. The proposal has been fully implemented and tested in the supervision process of gas network exploitation.projet FUI Gontran

Nonlinear mode coupling in rotating stars and the r-mode instability in neutron stars

We develop the formalism required to study the nonlinear interaction of modes in rotating Newtonian stars in the weakly nonlinear regime. The formalism simplifies and extends previous treatments. At linear order, we elucidate and extend slightly a formalism due to Schutz, show how to decompose a general motion of a rotating star into a sum over modes, and obtain uncoupled equations of motion for the mode amplitudes under the influence of an external force. Nonlinear effects are added perturbatively via three-mode couplings. We describe a new, efficient way to compute the coupling coefficients, to zeroth order in the stellar rotation rate, using spin-weighted spherical harmonics. We apply this formalism to derive some properties of the coupling coefficients relevant to the nonlinear interactions of unstable r-modes in neutron stars, postponing numerical integrations of the coupled equations of motion to a later paper. From an astrophysical viewpoint, the most interesting result of this paper is that many couplings of r-modes to other rotational modes (modes with zero frequencies in the non-rotating limit) are small: either they vanish altogether because of various selection rules, or they vanish to lowest order in the angular velocity. In zero-buoyancy stars, the coupling of three r-modes is forbidden entirely and the coupling of two r-modes to one hybrid rotational mode vanishes to zeroth order in rotation frequency. In incompressible stars, the coupling of any three rotational modes vanishes to zeroth order in rotation frequency.Comment: 62 pages, no figures. Corrected error in computation of coupling coefficients, added new selection rule and an appendix on energy and angular momentum of mode

Photometric mode identification methods of nonradial pulsations in eclipsing binaries I. -- Dynamic Eclipse Mapping

We present the Dynamic Eclipse Mapping (DEM) method designed specifically to reconstruct the surface intensity patterns of non-radial stellar oscillations in eclipsing binaries. The method needs a geometric model of the binary, accepts the light curve and the detected pulsation frequencies on input, and on output yields estimates of the pulsation patterns, in form of images -- thus allowing a direct identification of the surface mode numbers$(\ell,m)$. Since it has minimal modelling requirements and can operate on photometric observations in arbitrary wavelength bands, DEM is well suited to analyze the wide-band time series collected by space observatories. The method was extensively tested on simulated data, in which almost all photometrically detectable modes with a latitudinal complexity $\ell-|m|\le 4$ were properly restored. Multimode pulsations can be also reconstructed in a natural manner, as well as pulsations on components with tilted rotation axis of known direction. It can also be used in principle to isolate the contribution of hidden modes from the light curve. Sensitivity tests show that moderate errors in the geometric parameters and the assumed limb darkening can be partially tolerated by the inversion, in the sense that the lower degree modes are still recoverable. Tidally induced or mutually resonant pulsations, however, are an obstacle that neither the eclipse mapping, nor any other inversion technique can ever surpass. We conclude that, with reasonable assumptions, Dynamic Eclipse Mapping could be a powerful tool for mode identification, especially in moderately close eclipsing binary systems, where the pulsating component is not seriously affected by tidal interactions so that the pulsations are intrinsic to them, and not a consequence of the binarity.Comment: Accepted for publication in MNRAS. 16 pages, 11 figures and 5 table

Collaborative database to track Mass Mortality Events in the Mediterranean Sea

Anthropogenic climate change, and global warming in particular, has strong and increasing impacts on marine ecosystems (Poloczanska et al., 2013; Halpern et al., 2015; Smale et al., 2019). The Mediterranean Sea is considered a marine biodiversity hotspot contributing to more than 7% of world\u2019s marine biodiversity including a high percentage of endemic species (Coll et al., 2010). The Mediterranean region is a climate change hotspot, where the respective impacts of warming are very pronounced and relatively well documented (Cramer et al., 2018). One of the major impacts of sea surface temperature rise in the marine coastal ecosystems is the occurrence of mass mortality events (MMEs). The first evidences of this phenomenon dated from the first half of \u201980 years affecting the Western Mediterranean and the Aegean Sea (Harmelin, 1984; Bavestrello and Boero, 1986; Gaino and Pronzato, 1989; Voultsiadou et al., 2011). The most impressive phenomenon happened in 1999 when an unprecedented large scale MME impacted populations of more than 30 species from different phyla along the French and Italian coasts (Cerrano et al., 2000; Perez et al., 2000). Following this event, several other large scale MMEs have been reported, along with numerous other minor ones, which are usually more restricted in geographic extend and/or number of affected species (Garrabou et al., 2009; Rivetti et al., 2014; Marb\ue0 et al., 2015; Rubio-Portillo et al., 2016, authors\u2019 personal observations). These events have generally been associated with strong and recurrent marine heat waves (Crisci et al., 2011; Kersting et al., 2013; Turicchia et al., 2018; Bensoussan et al., 2019) which are becoming more frequent globally (Smale et al., 2019). Both field observations and future projections using Regional Coupled Models (Adloff et al., 2015; Darmaraki et al., 2019) show the increase in Mediterranean sea surface temperature, with more frequent occurrence of extreme ocean warming events. As a result, new MMEs are expected during the coming years. To date, despite the efforts, neither updated nor comprehensive information can support scientific analysis of mortality events at a Mediterranean regional scale. Such information is vital to guide management and conservation strategies that can then inform adaptive management schemes that aim to face the impacts of climate change

Dysferlin Forms a Dimer Mediated by the C2 Domains and the Transmembrane Domain In Vitro and in Living Cells

Dysferlin was previously identified as a key player in muscle membrane repair and its deficiency leads to the development of muscular dystrophy and cardiomyopathy. However, little is known about the oligomerization of this protein in the plasma membrane. Here we report for the first time that dysferlin forms a dimer in vitro and in living adult skeletal muscle fibers isolated from mice. Endogenous dysferlin from rabbit skeletal muscle exists primarily as a ∼460 kDa species in detergent-solubilized muscle homogenate, as shown by sucrose gradient fractionation, gel filtration and cross-linking assays. Fluorescent protein (YFP) labeled human dysferlin forms a dimer in vitro, as demonstrated by fluorescence correlation spectroscopy (FCS) and photon counting histogram (PCH) analyses. Dysferlin also dimerizes in living cells, as probed by fluorescence resonance energy transfer (FRET). Domain mapping FRET experiments showed that dysferlin dimerization is mediated by its transmembrane domain and by multiple C2 domains. However, C2A did not significantly contribute to dimerization; notably, this is the only C2 domain in dysferlin known to engage in a Ca-dependent interaction with cell membranes. Taken together, the data suggest that Ca-insensitive C2 domains mediate high affinity self-association of dysferlin in a parallel homodimer, leaving the Ca-sensitive C2A domain free to interact with membranes

Monocytes of patients with familial hypercholesterolemia show alterations in cholesterol metabolism

<p>Abstract</p> <p>Background</p> <p>Elevated plasma cholesterol promotes the formation of atherosclerotic lesions in which monocyte-derived lipid-laden macrophages are frequently found. To analyze, if circulating monocytes already show increased lipid content and differences in lipoprotein metabolism, we compared monocytes from patients with Familial Hypercholesterolemia (FH) with those from healthy individuals.</p> <p>Methods</p> <p>Cholesterol and oxidized cholesterol metabolite serum levels of FH and of healthy, gender/age matched control subjects were measured by combined gas chromatography – mass spectroscopy. Monocytes from patients with FH and from healthy subjects were isolated by antibody-assisted density centrifugation. Gene expression profiles of isolated monocytes were measured using Affymetrix HG-U 133 Plus 2.0 microarrays. We compared monocyte gene expression profiles from FH patients with healthy controls using a Welch T-test with correction for multiple testing (p < 0.05; Benjamini Hochberg correction, False Discovery Rate = 0.05). The differential expression of FH associated genes was validated at the mRNA level by qRT-PCR and/or at the protein level by Western Blot or flow cytometry. Functional validation of monocyte scavenger receptor activities were done by binding assays and dose/time dependent uptake analysis using native and oxidized LDL.</p> <p>Results</p> <p>Using microarray analysis we found in FH patients a significant up-regulation of 1,617 genes and a down-regulation of 701 genes compared to monocytes from healthy individuals. These include genes of proteins that are involved in the uptake, biosynthesis, disposition, and cellular efflux of cholesterol. In addition, plasma from FH patients contains elevated amounts of sterols and oxysterols. An increased uptake of oxidized as well as of native LDL by FH monocytes combined with a down-regulation of NPC1 and ABCA1 explains the lipid accumulation observed in these cells.</p> <p>Conclusion</p> <p>Our data demonstrate that circulating FH monocytes show differences in cell physiology that may contribute to the early onset of atherosclerosis in this disease.</p

Atomic Resolution Cryo-EM Structure Of A Nativelike CENP-A Nucleosome Aided By An Antibody Fragment

Genomic DNA in eukaryotes is organized into chromatin through association with core histones to form nucleosomes, each distinguished by their DNA sequences and histone variants. Here, we used a single-chain antibody fragment (scFv) derived from the anti-nucleosome antibody mAb PL2-6 to stabilize human CENP-A nucleosome containing a native α-satellite DNA and solved its structure by the cryo-electron microscopy (cryo-EM) to 2.6 Å resolution. In comparison, the corresponding cryo-EM structure of the free CENP-A nucleosome could only reach 3.4 Å resolution. We find that scFv binds to a conserved acidic patch on the histone H2A-H2B dimer without perturbing the nucleosome structure. Our results provide an atomic resolution cryo-EM structure of a nucleosome and insight into the structure and function of the CENP-A nucleosome. The scFv approach is applicable to the structural determination of other native-like nucleosomes with distinct DNA sequences