A substrate-dependent biological containment system for Pseudomonas putida based on the Escherichia coli gef gene.

A model substrate-dependent suicide system to biologically contain Pseudomonas putida KT2440 is reported. The system consists of two elements. One element carries a fusion between a synthetic lac promoter (PA1-04/03) and the gef gene, which encodes a killing function. This element is contained within a transposaseless mini-Tn5 transposon so that it can be integrated at random locations on the Pseudomonas chromosome. The second element, harbored by plasmid pCC102, is designed to control the first and bears a fusion between the promoter of the P. putida TOL plasmid-encoded meta-cleavage pathway operon (Pm) and the lacI gene, encoding the Lac repressor, plus xylS2, coding for a positive regulator of Pm. In liquid culture under optimal growth conditions and in sterile and nonsterile soil microcosms, P. putida KT2440 (pWWO) bearing the containment system behaves as designed. In the presence of a XylS effector, such as m-methylbenzoate, the LacI protein is synthesized, preventing the expression of the killing function. In the absence of effectors, expression of the PA1-04/03::gef cassette is no longer prevented and a high rate of cell killing is observed. Fluctuation test analyses revealed that mutants resistant to cell killing arise at a frequency of around 10(-5) to 10(-6) per cell per generation. Mutations are linked to the killing element rather than to the regulatory one. In bacteria bearing two copies of the killing cassette, the rate of appearance of mutants resistant to killing decreased to as low as 10(-8) per cell per generation.     

Left handed �� helix models for mammalian prion fibrils

We propose models for in vitro grown mammalian prion protein fibrils based upon left handed beta helices formed both from the N-terminal and C-terminal regions of the proteinase resistant infectious prion core. The C-terminal threading onto a β-helical structure is almost uniquely determined by fixing the cysteine disulfide bond on a helix corner. In comparison to known left handed helical peptides, the resulting model structures have similar stability attributes including relatively low root mean square deviations in all atom molecular dynamics, substantial side-chain-to-side-chain hydrogen bonding, good volume packing fraction, and low hydrophilic/hydrophobic frustration. For the N-terminus, we propose a new threading of slightly more than two turns, which improves upon the above characteristics relative to existing three turn β-helical models. The N-terminal and C-terminal beta helices can be assembled into eight candidate models for the fibril repeat units, held together by large hinge (order 30 residues) domain swapping, with three amenable to fibril promoting domain swapping via a small (five residue) hinge on the N-terminal side. Small concentrations of the metastable C-terminal β helix in vivo might play a significant role in templating the infectious conformation and in enhancing conversion kinetics for inherited forms of the disease and explain resistance (for canines) involving hypothesized coupling to the methionine 129 sulfur known to play a role in human disease.Key words: prion, amyloid fibril, domain swap, beta helix, computational biology     

transAlign: using amino acids to facilitate the multiple alignment of protein-coding DNA sequences

Background  

Alignments of homologous DNA sequences are crucial for comparative genomics and phylogenetic analysis. However, multiple alignment represents a computationally difficult problem. For protein-coding DNA sequences, it is more advantageous in terms of both speed and accuracy to align the amino-acid sequences specified by the DNA sequences rather than the DNA sequences themselves. Many implementations making use of this concept of "translated alignments" are incomplete in the sense that they require the user to manually translate the DNA sequences and to perform the amino-acid alignment. As such, they are not well suited to large-scale automated alignments of large and/or numerous DNA data sets.     

Progesterone transport across the wall of the rumen of sheep

Data on content of fiber, lignin and progesterone in reticulo-ruminal chyme and progesterone in blood serum and saliva of sheep throughout the reproductive cycle and at intramuscular injection of progesterone, are presented. A direct correlation between concentration of progesterone in blood and chyme was revealed. 1.5% of progesterone entered the reticulo-rumen with saliva. Transport of hormone through the wall of forestomach is the main way of progesterone entering the reticulo-rumen. The transport is carried out against the gradient of hormone concentration. Possible participation of progesterone adsorption on lignin in explanation of this transport, is discussed.     

Modeling Thrombus Shell: Linking Adhesion Receptor Properties and Macroscopic Dynamics

Damage to arterial vessel walls leads to the formation of platelet aggregate, which acts as a physical obstacle for bleeding. An arterial thrombus is heterogeneous; it has a dense inner part (core) and an unstable outer part (shell). The thrombus shell is very dynamic, being composed of loosely connected discoid platelets. The mechanisms underlying the observed mobility of the shell and its (patho)physiological implications are unclear. To investigate arterial thrombus mechanics, we developed a novel, to our knowledge, two-dimensional particle-based computational model of microvessel thrombosis. The model considers two types of interplatelet interactions: primary reversible (glycoprotein Ib (GPIb)-mediated) and stronger integrin-mediated interaction, which intensifies with platelet activation. At high shear rates, the former interaction leads to adhesion, and the latter is primarily responsible for stable platelet aggregation. Using a stochastic model of GPIb-mediated interaction, we initially reproduced experimental curves that characterize individual platelet interactions with a von Willebrand factor-coated surface. The addition of the second stabilizing interaction results in thrombus formation. The comparison of thrombus dynamics with experimental data allowed us to estimate the magnitude of critical interplatelet forces in the thrombus shell and the characteristic time of platelet activation. The model predicts moderate dependence of maximal thrombus height on the injury size in the absence of thrombin activity. We demonstrate that the developed stochastic model reproduces the observed highly dynamic behavior of the thrombus shell. The presence of primary stochastic interaction between platelets leads to the properties of thrombus consistent with in vivo findings; it does not grow upstream of the injury site and covers the whole injury from the first seconds of the formation. А simplified model, in which GPIb-mediated interaction is deterministic, does not reproduce these features. Thus, the stochasticity of platelet interactions is critical for thrombus plasticity, suggesting that interaction via a small number of bonds drives the dynamics of arterial thrombus shell.     

Neuregulin 3 promotes excitatory synapse formation on hippocampal interneurons

Hippocampal GABAergic interneurons are crucial for cortical network function and have been implicated in psychiatric disorders. We show here that Neuregulin 3 (Nrg3), a relatively little investigated low‐affinity ligand, is a functionally dominant interaction partner of ErbB4 in parvalbumin‐positive (PV) interneurons. Nrg3 and ErbB4 are located pre‐ and postsynaptically, respectively, in excitatory synapses on PV interneurons in vivo. Additionally, we show that ablation of Nrg3 results in a similar phenotype as the one described for ErbB4 ablation, including reduced excitatory synapse numbers on PV interneurons, altered short‐term plasticity, and disinhibition of the hippocampal network. In culture, presynaptic Nrg3 increases excitatory synapse numbers on ErbB4⁺ interneurons and affects short‐term plasticity. Nrg3 mutant neurons are poor donors of presynaptic terminals in the presence of competing neurons that produce recombinant Nrg3, and this bias requires postsynaptic ErbB4 but not ErbB4 kinase activity. Furthermore, when presented by non‐neuronal cells, Nrg3 induces postsynaptic membrane specialization. Our data indicate that Nrg3 provides adhesive cues that facilitate excitatory neurons to synapse onto ErbB4⁺ interneurons.     

Effect of latitude on adolescents living in the north

The latitude factor was shown to have a physiological effect on a growing body of adolescents aged 14–15 years even at a 3-degree difference in the latitude of residence. In adolescents living at 65°N, compared to those living at 62°N, the tolerance of hypoxemia, recovery of hemodynamics after short-term exercise, and their physical health level were decreased, attention switching was slower, and hypoxic events were more pronounced. Sex-related differences in several parameters and body features typical for northerners were revealed in adolescents living in the north.     

Functional role of apolipoprotein E in lipid metabolism in residents of the European North

The content of apolipoprotein E (ApoE) in the blood serum of residents of the European North was studied. It was shown that the limits of variation in the level of serum ApoE in northerners shifts towards lower values. ApoE contents below the standard values were detected in nearly half of the examined persons. It was shown that the ApoE content did not depend on sex and increased in older groups of northerners. Hypertriglyceridemia in the residents of the European North develops at lower ApoE contents in blood serum than in normolipidemia.