A conserved tetrameric interaction of cry toxin helix α3 suggests a functional role for toxin oligomerization

Crystal (Cry) toxins are widely used for insect control, but their mechanism of toxicity is still uncertain. These toxins can form lytic pores in vitro, and water soluble tetrameric pre-pore intermediates have been reported. Even the precise oligomeric state of the toxin in membranes, trimeric or tetrameric, is still a debated issue. Based on previous reports, we have assumed that interactions between toxin monomers in solution are at least partly mediated by domain I, and we have analyzed in silico the homo-oligomerization tendencies of the domain I α-helices individually. Using many homologous sequences for each α-helix, our strategy allows selection of evolutionarily conserved interactions. These interactions appeared only in helices α3 and α5, but only α3 produced a suitably oriented or α-helical sample in lipid bilayers, forming homotetramers in C14-betaine, and allowing determination of its rotational orientation in lipid bilayers using site-specific infrared dichroism (SSID). The determined orientation in the tetrameric model is in agreement with only one of the evolutionarily conserved models. In addition mutation R99E, which was found to inhibit oligomerization experimentally, greatly destabilized the tetramer in molecular dynamic simulations. In this model, helix 3 is able to form inter-monomer interactions without significant rearrangements of domain I, which is compatible with the available crystal structure of Cry toxins in solution. The model presented here at least partially explains the reported tetrameric oligomerization of Cry toxins in solution and the inhibition of this oligomerization by a synthetic α3 peptide.     

Compensatory Growth of Congenital Solitary Kidneys in Pigs Reflects Increased Nephron Numbers Rather Than Hypertrophy

Background

Patients with unilateral MultiCystic Kidney Dysplasia (MCKD) or unilateral renal agenesis (URA) have a congenital solitary functioning kidney (CSFK) that is compensatory enlarged. The question whether this enlargement is due to increased nephron numbers and/or to nephron hypertrophy is unresolved. This question is of utmost clinical importance, since hypertrophy is associated with a risk of developing hypertension and proteinuria later in life with consequent development of CKD and cardiovascular disease.

Methodology/Principal Findings

In a cohort of 32,000 slaughter pigs, 7 congenital solitary functioning kidneys and 7 control kidneys were identified and harvested. Cortex volume was measured and with a 3-dimensional stereologic technique the number and volume of glomeruli was determined and compared. The mean total cortex volume was increased by more than 80% and the mean number of glomeruli per kidney was 50% higher in CSFKs than in a single control kidney, equaling 75% of the total nephron number in both kidneys of control subjects. The mean total glomerular volume in the CSFKs was not increased relative to the controls.

Conclusions/Significance

Thus, in pigs, compensatory enlargement of a CSFK is based on increased nephron numbers. Extrapolation of these findings to the human situation suggests that patients with a CSFK might not be at increased risk for developing hyperfiltration-associated renal and cardiovascular disease in later life due to a lower nephron number.     

A Microscale Model for Combined CO2 Diffusion and Photosynthesis in Leaves

Transport of CO₂ in leaves was investigated by combining a 2-D, microscale CO₂ transport model with photosynthesis kinetics in wheat (Triticum aestivum L.) leaves. The biophysical microscale model for gas exchange featured an accurate geometric representation of the actual 2-D leaf tissue microstructure and accounted for diffusive mass exchange of CO_2. The resulting gas transport equations were coupled to the biochemical Farquhar-von Caemmerer-Berry model for photosynthesis. The combined model was evaluated using gas exchange and chlorophyll fluorescence measurements on wheat leaves. In general a good agreement between model predictions and measurements was obtained, but a discrepancy was observed for the mesophyll conductance at high CO₂ levels and low irradiance levels. This may indicate that some physiological processes related to photosynthesis are not incorporated in the model. The model provided detailed insight into the mechanisms of gas exchange and the effects of changes in ambient CO₂ concentration or photon flux density on stomatal and mesophyll conductance. It represents an important step forward to study CO₂ diffusion coupled to photosynthesis at the leaf tissue level, taking into account the leaf''s actual microstructure.     

Monosomy 9pter and trisomy 9q34.11qter in two sisters due to a maternal pericentric inversion

Pericentric inversions of chromosome 9 leading to unbalanced live-born offspring are relatively rare and so far only four cases have been reported. Here we present two sisters with an unbalanced recombinant chromosome 9 which resulted from a large maternal pericentric inversion inv(9)(p24.3q34.1). Further molecular characterisation of the aberrant chromosome 9 by 250k SNP array analysis showed a terminal 460 kb loss of 9p24.3 and a terminal 8.9 Mb gain of 9q34.11. We compared the clinical features of these two patients with the previous reported four cases as well as with patients with similar sized 9pter deletions or 9qter duplications. Based upon this study, we suggest that the recombinant chromosome 9 phenotype is mainly the result of duplication of a 3.4 Mb region of chromosome 9q34.11q34.13.     

Stable Vertical Takeoff of an Insect-Mimicking Flapping-Wing System Without Guide Implementing Inherent Pitching Stability

We briefly summarized how to design and fabricate an insect-mimicking flapping-wing system and demonstrate how to implement inherent pitching stability for stable vertical takeoff.The effect of relative locations of the Center of Gravity (CG) and the mean Aerodynamic Center (AC) on vertical flight was theoretically examined through static force balance consideration.We conducted a series of vertical takeoff tests in which the location of the mean AC was determined using an unsteady Blade Element Theory (BET) previously developed by the authors.Sequential images were captured during the takeoff tests using a high-speed camera.The results demonstrated that inherent pitching stability for vertical takeoff can be achieved by controlling the relative position between the CG and the mean AC of the flapping system.     

Blocking granule-mediated death by primary human NK cells requires both protection of mitochondria and inhibition of caspase activity

Human GraB (hGraB) preferentially induces apoptosis via Bcl-2-regulated mitochondrial damage but can also directly cleave caspases and caspase substrates in cell-free systems. How hGraB kills cells when it is delivered by cytotoxic lymphocytes (CL) and the contribution of hGraB to CL-induced death is still not clear. We show that primary human natural killer (hNK) cells, which specifically used hGraB to induce target cell death, were able to induce apoptosis of cells whose mitochondria were protected by Bcl-2. Purified hGraB also induced apoptosis of Bcl-2-overexpressing targets but only when delivered at 5- to 10-fold the concentration required to kill cells expressing endogenous Bcl-2. Caspases were critical in this process as inhibition of caspase activity permitted clonogenic survival of Bcl-2-overexpressing cells treated with hGraB or hNK cells but did not protect cells that only expressed endogenous Bcl-2. Our data therefore show that hGraB triggers caspase activation via mitochondria-dependent and mitochondria-independent mechanisms that are activated in a hierarchical manner, and that the combined effects of Bcl-2 and direct caspase inhibition can block cell death induced by hGraB and primary hNK cells.     

Analysis of the genome sequence of Lactobacillus gasseri ATCC 33323 reveals the molecular basis of an autochthonous intestinal organism

This study presents the complete genome sequence of Lactobacillus gasseri ATCC 33323, a neotype strain of human origin and a native species found commonly in the gastrointestinal tracts of neonates and adults. The plasmid-free genome was 1,894,360 bp in size and predicted to encode 1,810 genes. The GC content was 35.3%, similar to the GC content of its closest relatives, L. johnsonii NCC 533 (34%) and L. acidophilus NCFM (34%). Two identical copies of the prophage LgaI (40,086 bp), of the Sfi11-like Siphoviridae phage family, were integrated tandomly in the chromosome. A number of unique features were identified in the genome of L. gasseri that were likely acquired by horizontal gene transfer and may contribute to the survival of this bacterium in its ecological niche. L. gasseri encodes two restriction and modification systems, which may limit bacteriophage infection. L. gasseri also encodes an operon for production of heteropolysaccharides of high complexity. A unique alternative sigma factor was present similar to that of B. caccae ATCC 43185, a bacterial species isolated from human feces. In addition, L. gasseri encoded the highest number of putative mucus-binding proteins (14) among lactobacilli sequenced to date. Selected phenotypic characteristics that were compared between ATCC 33323 and other human L. gasseri strains included carbohydrate fermentation patterns, growth and survival in bile, oxalate degradation, and adhesion to intestinal epithelial cells, in vitro. The results from this study indicated high intraspecies variability from a genome encoding traits important for survival and retention in the gastrointestinal tract.     

Abnormal chloroplast development and growth inhibition in rice thioredoxin m knock-down plants

Plant cells contain several thioredoxin isoforms that are characterized by subcellular localization and substrate specificity. Here, we describe the functional characterization of a rice (Oryza sativa) thioredoxin m isoform (Ostrxm) using a reverse genetics technique. Ostrxm showed green tissue-specific and light-responsive mRNA expression. Ostrxm was localized in chloroplasts of rice mesophyll cells, and the recombinant protein showed dithiothreitol-dependent insulin β-chain reduction activity in vitro. RNA interference (RNAi) of Ostrxm resulted in rice plants with developmental defects, including semidwarfism, pale-green leaves, abnormal chloroplast structure, and reduced carotenoid and chlorophyll content. Ostrxm RNAi plants showed remarkably decreased F_v/F_m values under high irradiance conditions (1,000 μmol m⁻² s⁻¹) with delayed recovery. Two-dimensional electrophoresis and matrix-assisted laser-desorption/ionization time-of-flight analysis showed that the levels of several chloroplast proteins critical for photosynthesis and biogenesis were significantly decreased in Ostrxm RNAi plants. Furthermore, 2-Cys peroxiredoxin, a known target of thioredoxin, was present in oxidized forms, and hydrogen peroxide levels were increased in Ostrxm RNAi plants. The pleiotropic effects of Ostrxm RNAi suggest that Ostrxm plays an important role in the redox regulation of chloroplast target proteins involved in diverse physiological functions.     

A CASSCF/CASPT2 investigation on electron detachments from ScSi n − (n = 4–6) clusters

Journal of Molecular Modeling - The mechanism for the competitive adsorption of CH4 and CO2 on coal vitrinite (DV-8, maximum vitrinite reflectance R o,max = 0.58%) was revealed...