Priming in the Type I-F CRISPR-Cas system triggers strand-independent spacer acquisition,bi-directionally from the primed protospacer

Clustered regularly interspaced short palindromic repeats (CRISPR), in combination with CRISPR associated (cas) genes, constitute CRISPR-Cas bacterial adaptive immune systems. To generate immunity, these systems acquire short sequences of nucleic acids from foreign invaders and incorporate these into their CRISPR arrays as spacers. This adaptation process is the least characterized step in CRISPR-Cas immunity. Here, we used Pectobacterium atrosepticum to investigate adaptation in Type I-F CRISPR-Cas systems. Pre-existing spacers that matched plasmids stimulated hyperactive primed acquisition and resulted in the incorporation of up to nine new spacers across all three native CRISPR arrays. Endogenous expression of the cas genes was sufficient, yet required, for priming. The new spacers inhibited conjugation and transformation, and interference was enhanced with increasing numbers of new spacers. We analyzed ∼350 new spacers acquired in priming events and identified a 5′-protospacer-GG-3′ protospacer adjacent motif. In contrast to priming in Type I-E systems, new spacers matched either plasmid strand and a biased distribution, including clustering near the primed protospacer, suggested a bi-directional translocation model for the Cas1:Cas2–3 adaptation machinery. Taken together these results indicate priming adaptation occurs in different CRISPR-Cas systems, that it can be highly active in wild-type strains and that the underlying mechanisms vary.     

Neural inhibition sharpens auditory spatial selectivity of bat inferior collicular neurons

This study examines the role of neural inhibition in auditory spatial selectivity of inferior collicular neurons of the big brown bat, Eptesicus fuscus, using a two-tone inhibition paradigm. Two-tone inhibition decreases auditory spatial response areas but increases the slopes of directional sensitivity curves of inferior collicular neurons. Inferior collicular neurons have either directionally-selective or hemifield directional sensitivity curves. A directionally-selective curve always has a peak which is at least 50% larger than the minimum. A hemifield directional sensitivity curve rises from an ipsilateral angle by more than 50% and either reaches a plateau or declines by less than 50% over a range of contralateral angles. Two-tone inhibition does not change directionally-selective curves but changes most hemifield directional sensitivity curves into directionally-selective curves. Auditory spatial selectivity determined both with and without two-tone inhibition increases with increasing best-excitatory frequency. Sharpening of auditory spatial selectivity by two-tone inhibition is larger for neurons with smaller differences between excitatory and inhibitory best frequencies. The effect of two-tone inhibition on auditory spatial selectivity increases with increasing inhibitory tone intensity but decreases with increasing intertone interval. The implications of these findings in bat echolocation are discussed. Accepted: 18 January 2000     

Expression patterns and association analysis of the porcine DHX58 gene

RIG-1 signalling is responsible for the detection of cytoplasmic viral RNA molecules. DEXH (Asp-Glu-X-His) box polypeptide 58 (encoded by DHX58) is a negative regulator of the RIG-1 signalling pathway. In human, the DHX58 gene can be upregulated and can inhibit the RIG-1 signalling pathway during viral infection. In this study, porcine DHX58 gene expression patterns were studied. According to our results, the porcine DHX58 gene was upregulated not only by the stimulation of Poly I:C but also by the stimulation of 1ipopolysaccharides (LPS). One polymorphism (g.4919G>C), detected in the ninth intron, was significantly associated with some blood parameters including the red cell distribution width of 1-day-old pigs and white blood cell counts, lymphocyte absolute counts, and platelet distribution width of 17-day-old pigs (P < 0.05). Moreover, the individuals with the genotype GG have a significantly higher mean white blood cell count than individuals with genotype CC or GC (P < 0.05). Our study indicates that DHX58 is an important gene that is associated with the immune response in swine.     

Lysine-derived cross-links in the egg shell membrane

Egg shell membrane protein contains significant quantities of the lysine-derived aldehyde, allysine, and its aldol condensation product. NaB³H₄ reduction followed by alkaline hydrolysis of purified protein revealed that there were six residues/1000 of both allysine and the reduced aldol while only traces of desmosine and isodesmosine were detected. The amino acid composition of the membrane protein did not resemble that of mammalian elastin.     

A comparative analysis of photosynthetic characteristics of hulless barley at two altitudes on the Tibetan Plateau

To determine the photosynthetic characteristics of C₃ plants and their sensitivity to CO₂ at different altitudes on the Tibetan Plateau, hulless barley (Hordeum vulgare L. ssp. vulgare) was grown at altitudes of 4,333 m and 3,688 m. Using gas-exchange measurements, photosynthetic parameters were simulated, including the maximum net photosynthesis (P _max) and the apparent quantum efficiency (α). Plants growing at higher altitude had higher net photosynthetic rates (P _N), photosynthesis parameters (P _max and α) and sensitivities to CO₂ enhancement than plants growing at lower altitude on the Tibetan Plateau. The enhancements of P _N, P _max, and α for plants growing at higher altitude, corresponding with 10 μmol(CO₂) mol⁻¹ increments, were approximately 0.20∼0.45%, 0.05∼0.20% and 0.12∼0.36% greater, respectively, than for plants growing at lower altitude, respectively, where CO₂ levels rose from 10 to 170 μmol(CO₂) mol⁻¹. Therefore, on the Tibetan Plateau, the changes in the photosynthetic capacities and the photosynthetic sensitivities to CO₂ observed in the C₃ plants grown above 3,688 m are likely to increase with altitude despite the decreasing CO₂ partial pressure.     

Effect of propionate and pyruvate on citrulline synthesis and ATP content in rat liver mitochondria.

Propionate inhibits citrullinogenesis when succinate (plus rotenone) or glutamate are the oxidizable substrates used. Propionate decreases the intramitochondrial concentration of carbamylphosphate by decreasing the ATP content. When the energy supply for citrullinogenesis is provided by an influx of exogenous ATP, propionate is no longer an inhibitor. Pyruvate inhibits citrullinogenesis with glutamate but not with succinate (plus rotenone) as oxidizable substrates. Propionate and pyruvate deplete mitochondrial ATP but probably by different mechanisms.     

Free Edges in Epithelial Cell Sheets Stimulate Epidermal Growth Factor Receptor Signaling

The ability of epithelia to migrate and cover wounds is essential to maintaining their functions as physical barriers. Wounding induces many cues that may affect the transition to motility, including the immediate mechanical perturbation, release of material from broken cells, new interactions with adjacent extracellular matrix, and breakdown of physical separation of ligands from their receptors. Depending on the exact nature of wounds, some cues may be present only transiently or insignificantly. In many epithelia, activation of the epidermal growth factor receptor (EGFR) is a central event in induction of motility, and we find that its continuous activation is required for progression of healing of wounds in sheets of corneal epithelial cells. Here, we examine the hypothesis that edges, which are universally and continuously present in wounds, are a cue. Using a novel culture model we find that their presence is sufficient to cause activation of the EGFR and increased motility of cells in the absence of other cues. Edges that are bordered by agarose do not induce activation of the EGFR, indicating that activation is not due to loss of any specific type of cell–cell interaction but rather due to loss of physical constraints.