Respiratory Phenomics across Multiple Models of Protein Hyperacylation in Cardiac Mitochondria Reveals a Marginal Impact on Bioenergetics

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A reference microsatellite kit to assess for genetic diversity of Sorghum bicolor (Poaceae)

? Premise of the study: Discrepancies in terms of genotyping data are frequently observed when comparing simple sequence repeat (SSR) data sets across genotyping technologies and laboratories. This technical concern introduces biases that hamper any synthetic studies or comparison of genetic diversity between collections. To prevent this for Sorghum bicolor, we developed a control kit of 48 SSR markers. ? Methods and Results: One hundred seventeen markers were selected along the genome to provide coverage across the length of all 10 sorghum linkage groups. They were tested for polymorphism and reproducibility across two laboratories (Centre de Cooperation Internationale en Recherche Agronomique pour le Developpement [CIRAD], France, and International Crops Research Institute for the Semi-Arid Tropics [ICRISAT], India) using two commonly used genotyping technologies (polyacrylamide gel-based technology with LI-COR sequencing machines and capillary systems with ABI sequencing apparatus) with DNA samples from a diverse set of 48 S. bicolor accessions. ? Conclusions: A kit for diversity analysis (http://sat.cirad.fr/sat/sorghum_SSR_kit/) was developed. It contains information on 48 technically robust sorghum microsatellite markers and 10 DNA controls. It can further be used to calibrate sorghum SSR genotyping data acquired with different technologies and compare those to genetic diversity references.     

Long-distance seed dispersal, clone longevity and lack of phylogeographical structure in the European distributional range of the coastal Calystegia soldanella (L.) R. Br. (Convolvulaceae)

Aim  To explore the relative effects of Quaternary climatic history vs. species-specific biological properties (high seed dispersability, high seed longevity, clonal growth) on phylogeographical structure in European Mediterranean and Atlantic Ocean and Black Sea material of the coastal dune plant Calystegia soldanella (L.) R. Br.
Location  Black Sea and European Mediterranean and Atlantic Ocean coasts.
Methods  Variation in amplified fragment length polymorphism was analysed at two different sampling levels. First, an entire-range sample from the Black Sea to the North Sea, including single individuals from sites evenly spread along this entire coast was analysed. Second, in a population-level sample, seven populations from the European Mediterranean and Atlantic Ocean coasts were analysed.
Results  Neither the entire-range nor the population-level sampling resulted in clear phylogeographical patterns. Instead, individuals from geographically distant areas were often genetically more similar to each other than individuals from the same area. Non-significant isolation-by-distance was found for both sampling approaches, and comparatively low levels of intrapopulational genetic variation were observed.
Main conclusions  The lack of phylogeographical structure in C. soldanella , in comparison with the clear phylogeographical patterns observed in other coastal plant species analysed previously, is postulated to be the result of the specific biology of C. soldanella . The combination of high seed longevity, high dispersability of seeds by sea water and clonal growth and probable high clone age are likely to be responsible for the observed absence of phylogeographical structure. This implies that extreme biological properties such as those shown by C. soldanella can either erase or prevent the formation of historical patterns of genetic variation.     

Characterization of xanthans from selected Xanthomonas strains cultivated under constant dissolved oxygen

Seventeen wild-type Xanthomonas isolates were screened in terms of broth viscosity in shake-flasks. As culture conditions affect polymer characteristics, a fair comparison among isolates required their cultivation in a fermenter under controlled dissolved oxygen tension. Three isolates and a reference strain were studied. The mean molecular weights and molecular weight distributions of their xanthans were determined. Products showed different pyruvate (0.2–7%), acetate (5–10%) and proteinaceous nitrogen (1–3%) contents. The selected isolates exhibit properties which could improve xanthan gum production and some could be used to produce polymers with specific characteristics.     

Proteome wide evaluation of the separation ability of hydrophobic interaction chromatography by fluorescent dye binding analysis

Hydrophobic interaction chromatography (HIC) is an important tool in the industrial purification of proteins from various sources. The HIC separation behavior of individual (or model) proteins has been widely researched by others. On the contrary, this study focused on the fractionation ability of HIC when it is challenged with whole proteomes. The impact of the nature of three different proteomes, that is, yeast, soybean, and Chinese hamster ovary cells, on HIC separation was investigated. In doing so, chromatography fractions obtained under standardized conditions were evaluated in terms of their overall hydrophobicity—as measured by fluorescence dye binding. This technique allowed for the calculation of an average protein surface hydrophobicity (S₀) for each fraction; a unique correlation between S₀ and the observed chromatographic behavior was established in each case. Following a similar strategy, the effect of three different ligands (polypropylene glycol, phenyl, and butyl) and two adsorbent particle sizes (65 and 100 µm) on the chromatographic behavior of the yeast proteome was evaluated. As expected, the superficial hydrophobicity of the proteins eluted is correlated with the salt concentration of its corresponding elution step. The findings reveled how—and in which extent—the type of ligand and the size of the beads actually influenced the fractionation of the complex biological mixture. Summarizing, the approach presented here can be instrumental to the study of the performance of chromatography adsorbents under conditions close to industrial practice and to the development of downstream processing strategies. Copyright © 2013 John Wiley & Sons, Ltd.     

Characterization of transgenic poplar with ectopic expression of pine cytosolic glutamine synthetase under conditions of varying nitrogen availability

The present study addresses the hypothesis that enhanced expression of glutamine synthetase (GS) in transgenic poplar, characterized by the ectopic expression of pine cytosolic GS, results in an enhanced efficiency of nitrogen (N) assimilation and enhanced growth. Transgenic and control poplar were supplied with low and high N levels and the role of ectopic expression of the pine GS in growth and N assimilation was assessed by using amino acid analysis, (15)N enrichment, biochemical analyses, and growth measurements. While leaves of transgenic poplar contained 85% less (P < 0.01) free ammonium than leaves of nontransgenic control plants, leaves of transgenics showed increases in the levels of free glutamine and total free amino acids. Transgenic poplar lines also displayed significant increases in growth parameters when compared with controls grown under both low (0.3 mm) and high (10 mm) nitrate conditions. Furthermore, (15)N-enrichment experiments showed that 27% more (P < 0.05) (15)N was incorporated into structural compounds in transgenic lines than in nontransgenic controls. Using the methods described here, we present direct evidence for increased N assimilation efficiency and growth in GS transgenic lines.     

A small group of sulfated benzofurans induces steady-state submaximal inhibition of thrombin

Despite the development of promising direct oral anticoagulants, which are all orthosteric inhibitors, a sizable number of patients suffer from bleeding complications. We have hypothesized that allosterism based on the heparin-binding exosites presents a major opportunity to induce sub-maximal inhibition of coagulation proteases, thereby avoiding/reducing bleeding risk. We present the design of a group of sulfated benzofuran dimers that display heparin-binding site-dependent partial allosteric inhibition of thrombin against fibrinogen (ΔY?=?55–75%), the first time that a small molecule (MW??<?800) has been found to thwart macromolecular cleavage by a monomeric protease in a controlled manner. The work leads to the promising concept that it should be possible to develop allosteric inhibitors that reduce clotting, but do not completely eliminate it, thereby avoiding major bleeding complications that beset anticoagulants today.     

A genome-wide scan of Ashkenazi Jewish Crohn's disease suggests novel susceptibility loci

Crohn''s disease (CD) is a complex disorder resulting from the interaction of intestinal microbiota with the host immune system in genetically susceptible individuals. The largest meta-analysis of genome-wide association to date identified 71 CD–susceptibility loci in individuals of European ancestry. An important epidemiological feature of CD is that it is 2–4 times more prevalent among individuals of Ashkenazi Jewish (AJ) descent compared to non-Jewish Europeans (NJ). To explore genetic variation associated with CD in AJs, we conducted a genome-wide association study (GWAS) by combining raw genotype data across 10 AJ cohorts consisting of 907 cases and 2,345 controls in the discovery stage, followed up by a replication study in 971 cases and 2,124 controls. We confirmed genome-wide significant associations of 9 known CD loci in AJs and replicated 3 additional loci with strong signal (p<5×10⁻⁶). Novel signals detected among AJs were mapped to chromosomes 5q21.1 (rs7705924, combined p = 2×10⁻⁸; combined odds ratio OR = 1.48), 2p15 (rs6545946, p = 7×10⁻⁹; OR = 1.16), 8q21.11 (rs12677663, p = 2×10⁻⁸; OR = 1.15), 10q26.3 (rs10734105, p = 3×10⁻⁸; OR = 1.27), and 11q12.1 (rs11229030, p = 8×10⁻⁹; OR = 1.15), implicating biologically plausible candidate genes, including RPL7, CPAMD8, PRG2, and PRG3. In all, the 16 replicated and newly discovered loci, in addition to the three coding NOD2 variants, accounted for 11.2% of the total genetic variance for CD risk in the AJ population. This study demonstrates the complementary value of genetic studies in the Ashkenazim.     

Depth-dependent analysis of the role of collagen fibrils, fixed charges and fluid in the pericellular matrix of articular cartilage on chondrocyte mechanics

The pericellular matrix of articular cartilage has been shown to regulate the mechanical environment of chondrocytes. However, little is known about the mechanical role of collagen fibrils in the pericellular matrix, and how fibrils might help modulate strains acting on chondrocytes when cartilage is loaded. The primary objective was to clarify the effect of pericellular collagen fibrils on cell volume changes and strains during cartilage loading. Secondary objectives were to investigate the effects of pericellular fixed charges and fluid on cell responses. A microstructural model of articular cartilage, in which chondrocytes and pericellular matrices were represented with depth-dependent structural and morphological properties, was created. The extracellular matrix and pericellular matrices were modeled as fibril-reinforced, biphasic materials with swelling capabilities, while chondrocytes were assumed to be isotropic and biphasic with swelling properties. Collagen fibrils in the extracellular matrix were represented with an arcade-like architecture, whereas pericellular fibrils were assumed to run tangential to the cell surface. In the early stages of a stress-relaxation test, pericellular fibrils were found to sensitively affect cell volume changes, even producing a reversal from increasing to decreasing cell volume with increasing fibril stiffness in the superficial zone. Consequently, steady-state volume of the superficial zone cell decreased with increasing pericellular fibril stiffness. Volume changes in the middle and deep zone chondrocytes were smaller and opposite to those observed in the superficial zone chondrocyte. An increase in the pericellular fixed charge density reduced cell volumes substantially in every zone. The sensitivity of cell volume changes to pericellular fibril stiffness suggests that pericellular fibrils play an important, and as of yet largely neglected, role in regulating the mechanical environment of chondrocytes, possibly affecting matrix synthesis during cartilage development and degeneration, and affecting biosynthetic responses associated with articular cartilage loading.