Exponential Size Distribution of von Willebrand Factor

Von Willebrand Factor (VWF) is a multimeric protein crucial for hemostasis. Under shear flow, it acts as a mechanosensor responding with a size-dependent globule-stretch transition to increasing shear rates. Here, we quantify for the first time, to our knowledge, the size distribution of recombinant VWF and VWF-eGFP using a multilateral approach that involves quantitative gel analysis, fluorescence correlation spectroscopy, and total internal reflection fluorescence microscopy. We find an exponentially decaying size distribution of multimers for recombinant VWF as well as for VWF derived from blood samples in accordance with the notion of a step-growth polymerization process during VWF biosynthesis. The distribution is solely described by the extent of polymerization, which was found to be reduced in the case of the pathologically relevant mutant VWF-IIC. The VWF-specific protease ADAMTS13 systematically shifts the VWF size distribution toward smaller sizes. This dynamic evolution is monitored using fluorescence correlation spectroscopy and compared to a computer simulation of a random cleavage process relating ADAMTS13 concentration to the degree of VWF breakdown. Quantitative assessment of VWF size distribution in terms of an exponential might prove to be useful both as a valuable biophysical characterization and as a possible disease indicator for clinical applications.     

Archaeal diversity from hydrothermal systems of Deception Island, Antarctica

Antarctica is an extreme continent composed of cold environments but also of several geothermal sites, among them is Deception Island, an active stratovolcano located in the South Shetland archipelago. From this island, few microbiological studies have been performed, and the presence of archaea has not been reported. In order to investigate the archaeal diversity in hydrothermalism from Deception Island, different submarine samples were taken from the flooded caldera. Samples were analyzed by denaturing gradient gel electrophoresis (DGGE) of the 16S rRNA gene in conjunction with culture-dependent methods at hyperthermophilic temperatures. Analysis from DGGE band sequencing showed the presence of archaea belonging to the hyperthermophilic genus Thermococcus and different uncultured archaea closely related to environmental clones from hydrothermal vents. Archaea from the psychrotolerant genus Methanococcoides were also detected. Additionally, we have successfully isolated an anaerobic hyperthermophilic archaeon closely related to Thermococcus celericrescens. Cells were irregular cocci with a diameter between 0.6 and 2 μm and grew at 50–90 °C and at a NaCl concentration of 1–5 %. Here, we present, based on culture-dependent and culture-independent approaches, the first report on archaea from marine hydrothermal sites of Antarctica.     

Growth and nutrition of black spruce seedlings in response to disruption of Pleurozium and Sphagnum moss carpets in boreal forested peatlands

In boreal forested peatlands, we disturbed Sphagnum spp. and Pleurozium schreberi carpets to see how disturbance influenced substrate physico-chemistry, and growth and foliar nutrition of planted Picea mariana seedlings. Carpets were hand disturbed using gardening tools to a depth of approximately 25 cm. Carpet disturbance was aimed at disrupting only the organic layer and did not result in the mixing of organic matter with mineral soil. Disturbed carpets, whether P. schreberi or Sphagnum spp., were warmer than undisturbed carpets and had a lower cover of ericaceous shrubs. Pleurozium schreberi carpets had a higher decomposition index than Sphagnum spp. carpets, whereas disturbance had no effect on this variable. Pleurozium schreberi had higher N_tot and dissolved organic N concentrations (DON) than Sphagnum spp., whereas disturbance increased NH ₄ ⁺ availability in both substrates. Moss disruption increased seedling growth rates as well as their foliar N and P concentrations in both substrates and these variables remained higher in P. schreberi than in Sphagnum spp. within a given treatment. Seedling growth was positively correlated to substrate N_tot, NH ₄ ⁺ and DON concentrations, and to foliar N and P concentrations, and negatively to substrate C/N and ericaceous shrub cover. Disruption of the moss carpets without mineral soil mixing improved black spruce seedling growth and nutrition in both moss types but the superiority of P. schreberi compared to Sphagnum spp. as a growing substrate remained present.     

Computational analysis of the influence of the microenvironment on carcinogenesis

The tumour microenvironment is known to play an important role in determining the sequence of acquired phenotypic traits that characterise cancer evolution. A more precise understanding of this role could have a major influence in the understanding of cancer growth and development, and potentially in the optimisation of innovative anti-cancer treatments delivery. However, to lead such an analysis in the basis of traditional biological experiments and observations is almost utopian given the complexity of the underlying biological processes and the typical time scales involved. In this context, computer models constitute a complementary exploratory tool.In this paper we introduce a two-dimensional cellular automaton that models key cancer cell capabilities. The model has been especially designed to mimic the behaviour of a cancer colony growing in vitro and to analyse the effect of different environmental conditions on the sequence of acquisition of phenotypic traits. Our results indicate that microenvironmental factors such as the local concentration of oxygen or nutrients and cell overcrowding may determine the expansion of the tumour colony. The results also show that tumour cells evolve and that their phenotypes adapt to the microenvironment so that environmental stress determines the dominance of particular phenotypical traits.     

Rigid-body ligand recognition drives cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor triggering

The inhibitory T-cell surface-expressed receptor, cytotoxic T lymphocyte-associated antigen-4 (CTLA-4), which belongs to the class of cell surface proteins phosphorylated by extrinsic tyrosine kinases that also includes antigen receptors, binds the related ligands, B7-1 and B7-2, expressed on antigen-presenting cells. Conformational changes are commonly invoked to explain ligand-induced “triggering” of this class of receptors. Crystal structures of ligand-bound CTLA-4 have been reported, but not the apo form, precluding analysis of the structural changes accompanying ligand binding. The 1.8-Å resolution structure of an apo human CTLA-4 homodimer emphasizes the shared evolutionary history of the CTLA-4/CD28 subgroup of the immunoglobulin superfamily and the antigen receptors. The ligand-bound and unbound forms of both CTLA-4 and B7-1 are remarkably similar, in marked contrast to B7-2, whose binding to CTLA-4 has elements of induced fit. Isothermal titration calorimetry reveals that ligand binding by CTLA-4 is enthalpically driven and accompanied by unfavorable entropic changes. The similarity of the thermodynamic parameters determined for the interactions of CTLA-4 with B7-1 and B7-2 suggests that the binding is not highly specific, but the conformational changes observed for B7-2 binding suggest some level of selectivity. The new structure establishes that rigid-body ligand interactions are capable of triggering CTLA-4 phosphorylation by extrinsic kinase(s).     

Evolutionary constraint in flanking regions of avian genes

An important comprehension from comparative genomic analysis is that sequence conservation beyond neutral expectations is frequently found outside protein-coding regions, indicating important functional roles of noncoding DNA. Understanding the causes of constraint on noncoding sequence evolution forms an important area of research, not least in light of the importance for understanding the evolution of gene expression. We aligned all orthologous genes of chicken and zebra finch together with 5 kb of their upstream and downstream noncoding sequences, to study the evolution of gene flanking sequences in the avian genome. Using ancestral repeats as a neutral reference, we detected significant evolutionary constraint in the 3' flanking region, highest directly after termination (60%) and then gradually decreasing to about 20% 5 kb downstream. Constraint was higher in annotated 3' untranslated regions (UTRs) than in non-UTRs at the same distance from the stop codon and higher in sequences annotated as microRNA (miRNA)-binding sites than in non-miRNA-binding sites within 3' UTRs. Constraint was also higher when estimated for a smaller data set of genes from more closely related songbird species, indicating turnover of functional elements during avian evolution. On the 5' flanking side constraint was readily seen within the first 125 bp immediately upstream of the start codon (34%) and was about 10% for remaining sequence within 5 kb upstream. Analysis of chicken polymorphism data gave further support for the highest constraint directly before and after the translated region. Finally, we found that genes evolving under the highest constraint measured by d(N)/d(S) also had the highest level of constraint in the 3' flanking region. This study broadens the insights into gene flanking sequence evolution by adding new findings from a vertebrate lineage other than mammals.