Basal activation of the P2X7 ATP receptor elevates mitochondrial calcium and potential, increases cellular ATP levels, and promotes serum-independent growth

P2X7 is a bifunctional receptor (P2X7R) for extracellular ATP that, depending on the level of activation, forms a cation-selective channel or a large conductance nonselective pore. The P2X7R has a strong proapoptotic activity but can also support growth. Here, we describe the mechanism involved in growth stimulation. Transfection of P2X7R increases resting mitochondrial potential (delta psi(mt)), basal mitochondrial Ca2+ ([Ca2+]mt), intracellular ATP content, and confers ability to grow in the absence of serum. These changes require a full pore-forming function, because they are abolished in cells transfected with a mutated P2X7R that retains channel activity but cannot form the nonselective pore, and depend on an autocrine/paracrine tonic stimulation by secreted ATP. On the other hand, sustained stimulation of P2X7R causes a delta psi(mt) drop, a large increase in [Ca2+]mt, mitochondrial fragmentation, and cell death. These findings reveal a hitherto undescribed mechanism for growth stimulation by a plasma membrane pore.     

MILVA: an interactive tool for the exploration of multidimensional microarray data

MOTIVATION: Clustering techniques such as k-means and hierarchical clustering are commonly used to analyze DNA microarray derived gene expression data. However, the interactions between processes underlying the cell activity suggest that the complexity of the microarray data structure may not be fully represented with discrete clustering methods. RESULTS: A newly developed software tool called MILVA (microarray latent visualization and analysis) is presented here to investigate microarray data without separating gene expression profiles into discrete classes. The underpinning of the MILVA software is the two-dimensional topographic representation of multidimensional microarray data. On this basis, the interactive MILVA functions allow a continuous exploration of microarray data driven by the direct supervision of the biologist in detecting activity patterns of co-regulated genes. AVAILABILITY: The MILVA software is freely available. The software and the related documentation can be downloaded from http://www.ncrg.aston.ac.uk/Projects/milva. User 'surrey' as username and '3245' as password to login. The software is currently available for Windows platform only.     

The orientations of cytochrome c in the highly dynamic complex with cytochrome b5 visualized by NMR and docking using HADDOCK

The interaction of bovine microsomal ferricytochrome b5 with yeast iso-1-ferri and ferrocytochrome c has been investigated using heteronuclear NMR techniques. Chemical-shift perturbations for 1H and 15N nuclei of both cytochromes, arising from the interactions with the unlabeled partner proteins, were used for mapping the interacting surfaces on both proteins. The similarity of the binding shifts observed for oxidized and reduced cytochrome c indicates that the complex formation is not influenced by the oxidation state of the cytochrome c. Protein-protein docking simulations have been performed for the binary cytochrome b5-cytochrome c and ternary (cytochrome b5)-(cytochrome c)2 complexes using a novel HADDOCK approach. The docking procedure, which makes use of the experimental data to drive the docking, identified a range of orientations assumed by the proteins in the complex. It is demonstrated that cytochrome c uses a confined surface patch for interaction with a much more extensive surface area of cytochrome b5. Taken together, the experimental data suggest the presence of a dynamic ensemble of conformations assumed by the proteins in the complex.     

Aerobic cometabolism of chloroform by butane-grown microorganisms: long-term monitoring of depletion rates and isolation of a high-performing strain

The focus of this microcosm study was to monitor the performances of 17 butane-utilizing microcosms during a long-term (100–250 days) aerobic cometabolic depletion of chloroform (CF). The depletion of the contaminant began after a lag-time variable between 0 and 23 days. All microcosms quickly reached a pseudo steady-state condition, in terms of biomass concentration (with an average of 9.3 × 10⁶ CFU ml^–1), chloroform depletion rate (5 mol l^–1 d^–1) and butane utilization rate (730 mol l^–1 d^–1). After about 100 days of CF depletion, a sudden 5- to 7-fold increase of the chloroform rate was observed in two microcosms, where the highest amount of contaminant had been depleted. In one of these high-performing microcosms, an experiment of chloroform depletion in the absence of butane resulted in the depletion of a surprisingly high amount of contaminant (765 mol_CF kg_{dry soil}^–1 in 2 months) and in a marked selection of a single bacterial strain. Bioaugmentation assays conducted with the biomass selected in this microcosm and with a pure culture of the selected strain immediately resulted in very high chloroform depletion rates. Preliminary results of a study conducted with resting cells of the selected strain indicated that it can degrade chloroform concentrations up to 119 M (14.2 mg l^–1) without any sign of substrate toxicity, and that it is able to transform vinyl chloride and 1,1,2-trichloroethane.