Characterization of temperature-sensitive mutants of bacteriophage PM2: Membrane mutants

Summary In an effort to understand the genetic regulation of membrane morphogenesis, twenty-nine temperature-sensitive mutants of the membrane-containing bacteriophage PM2 were isolated. Characterization at restrictive temperature revealed groups showing no lysis (Groups I–IV), partial lysis (Groups V–VIII), and full lysis (Groups IX–XII) of the host Pseudomonas BAL-31. When the cell lysis data are considered in conjunction with data on stimulation of viral DNA synthesis, at least six mutant groups are defined. Analysis by gel electrophoresis of the pattern of viral proteins synthesized under restrictive conditions further divides the mutants into twelve groups. Temperature shift experiments delineate early, intermediate and late mutants. Complementation data support some of these groupings. The observed low levels of complementation and recombination are discussed in terms of gene product/genome restriction, bound to the membrane at the site of infection.It is of particular interest to membrane morphogenesis that under restrictive conditions late mutants in Groups II, III and IV make empty-appearing vesicles inside the cell that are the size of virus membranes as seen in thin sections of cells in the electron microscope. Mutants ts 1 (Group II) and ts 12 (Group III) show defects in their ability to incorporate into membranes viral structural proteins sp 13 and sp 6.6. The possibility is discussed that either of these proteins control the size and shape of the viral membrane.     

Hydrolysis of beta-D-glucopyranosyl fluoride to alpha-D-glucose catalyzed by Aspergillus niger alpha-D-glucosidase

Aspergillus niger alpha-D-glucosidase, crystallized and free of detectable activity for beta-D-glucosides, catalyzes the slow hydrolysis of beta-D-glucopyranosyl fluoride to form alpha-D-glucose. Maximal initial rates, V, for the hydrolysis of beta-D-glucosyl fluoride, p-nitrophenyl alpha-D-glucopyranoside, and alpha-D-glucopyranosyl fluoride are 0.27, 0.75, and 78.5 mumol.min-1.mg-1, respectively, with corresponding V/K constants of 0.0068, 1.44, and 41.3. Independent lines of evidence make clear that the reaction stems from beta-D-glucosyl fluoride and not from a contaminating trace of alpha-D-glucosyl fluoride, and is catalyzed by the alpha-D-glucosidase and not by an accompanying trace of beta-D-glucosidase or glucoamylase. Maltotriose competitively inhibits the hydrolysis, and beta-D-glucosyl fluoride in turn competitively inhibits the hydrolysis of p-nitrophenyl alpha-D-glucopyranoside, indicating that beta-D-glucosyl fluoride is bound at the same site as known substrates for the alpha-glucosidase. Present findings provide new evidence that alpha-glucosidases are not restricted to alpha-D-glucosylic substrates or to reactions providing retention of configuration. They strongly support the concept that product configuration in glycosylase-catalyzed reactions is primarily determined by enzyme structures controlling the direction of approach of acceptor molecules to the reaction center rather than by the anomeric configuration of the substrate.     

Magnesium isotope fractionation during microbially enhanced forsterite dissolution

Bacillus subtilis endospore‐mediated forsterite dissolution experiments were performed to assess the effects of cell surface reactivity on Mg isotope fractionation during chemical weathering. Endospores present a unique opportunity to study the isolated impact of cell surface reactivity because they exhibit extremely low metabolic activity. In abiotic control assays, ²⁴Mg was preferentially released into solution during forsterite dissolution, producing an isotopically light liquid phase (δ²⁶Mg = ?0.39 ± 0.06 to ?0.26 ± 0.09‰) relative to the initial mineral composition (δ²⁶Mg = ?0.24 ± 0.03‰). The presence of endospores did not have an apparent effect on Mg isotope fractionation associated with the release of Mg from the solid into the aqueous phase. However, the endospore surfaces preferentially adsorbed ²⁴Mg from the dissolution products, which resulted in relatively heavy aqueous Mg isotope compositions. These aqueous Mg isotope compositions increased proportional to the fraction of dissolved Mg that was adsorbed, with the highest measured δ²⁶Mg (?0.08 ± 0.07‰) corresponding to the highest degree of adsorption (~76%). The Mg isotope composition of the adsorbed fraction was correspondingly light, at an average δ²⁶Mg of ?0.49‰. Secondary mineral precipitation and Mg adsorption onto secondary minerals had a minimal effect on Mg isotopes at these experimental conditions. Results demonstrate the isolated effects of cell surface reactivity on Mg isotope fractionation separate from other common biological processes, such as metabolism and organic acid production. With further study, Mg isotopes could be used to elucidate the role of the biosphere on Mg cycling in the environment.     

Identification of Pyrus Single Nucleotide Polymorphisms (SNPs) and Evaluation for Genetic Mapping in European Pear and Interspecific Pyrus Hybrids

We have used new generation sequencing (NGS) technologies to identify single nucleotide polymorphism (SNP) markers from three European pear (Pyrus communis L.) cultivars and subsequently developed a subset of 1096 pear SNPs into high throughput markers by combining them with the set of 7692 apple SNPs on the IRSC apple Infinium® II 8K array. We then evaluated this apple and pear Infinium® II 9K SNP array for large-scale genotyping in pear across several species, using both pear and apple SNPs. The segregating populations employed for array validation included a segregating population of European pear (‘Old Home’בLouise Bon Jersey’) and four interspecific breeding families derived from Asian (P. pyrifolia Nakai and P. bretschneideri Rehd.) and European pear pedigrees. In total, we mapped 857 polymorphic pear markers to construct the first SNP-based genetic maps for pear, comprising 78% of the total pear SNPs included in the array. In addition, 1031 SNP markers derived from apple (13% of the total apple SNPs included in the array) were polymorphic and were mapped in one or more of the pear populations. These results are the first to demonstrate SNP transferability across the genera Malus and Pyrus. Our construction of high density SNP-based and gene-based genetic maps in pear represents an important step towards the identification of chromosomal regions associated with a range of horticultural characters, such as pest and disease resistance, orchard yield and fruit quality.     

Repeated oral administration of capsaicin increases anxiety-like behaviours with prolonged stress-response in rats

This study was conducted to examine the psycho-emotional effects of repeated oral exposure to capsaicin, the principal active component of chili peppers. Each rat received 1 mL of 0.02% capsaicin into its oral cavity daily, and was subjected to behavioural tests following 10 daily administrations of capsaicin. Stereotypy counts and rostral grooming were significantly increased, and caudal grooming decreased, in capsaicin-treated rats during the ambulatory activity test. In elevated plus maze test, not only the time spent in open arms but also the percent arm entry into open arms was reduced in capsaicin-treated rats compared with control rats. In forced swim test, although swimming duration was decreased, struggling increased in the capsaicin group, immobility duration did not differ between the groups. Repeated oral capsaicin did not affect the basal levels of plasma corticosterone; however, the stress-induced elevation of plasma corticosterone was prolonged in capsaicin treated rats. Oral capsaicin exposure significantly increased c-Fos expression not only in the nucleus tractus of solitarius but also in the paraventricular nucleus. Results suggest that repeated oral exposure to capsaicin increases anxiety-like behaviours in rats, and dysfunction of the hypothalamic-pituitary-adrenal axis may play a role in its pathophysiology.