Shaping dots and lines: adding modularity into protein interaction networks using structural information

Determining protein interaction networks and generating models to simulate network changes in time and space are crucial for understanding a biological system and for predicting the effect of mutants found in diseases. In this review we discuss the great potential of using structural information together with computational tools towards reaching this goal: the prediction of new protein interactions, the estimation of affinities and kinetic rate constants between protein complexes, and finally the determination of which interactions are compatible with each other and which interactions are exclusive. The latter one will be important to reorganize large scale networks into functional modular networks.     

Specific regions within the embryonic midbrain and cerebellum require different levels of FGF signaling during development

Prospective midbrain and cerebellum formation are coordinated by FGF ligands produced by the isthmic organizer. Previous studies have suggested that midbrain and cerebellum development require different levels of FGF signaling. However, little is known about the extent to which specific regions within these two parts of the brain differ in their requirement for FGF signaling during embryogenesis. Here, we have explored the effects of inhibiting FGF signaling within the embryonic mouse midbrain (mesencephalon) and cerebellum (rhombomere 1) by misexpressing sprouty2 (Spry2) from an early stage. We show that such Spry2 misexpression moderately reduces FGF signaling, and that this reduction causes cell death in the anterior mesencephalon, the region furthest from the source of FGF ligands. Interestingly, the remaining mesencephalon cells develop into anterior midbrain, indicating that a low level of FGF signaling is sufficient to promote only anterior midbrain development. Spry2 misexpression also affects development of the vermis, the part of the cerebellum that spans the midline. We found that, whereas misexpression of Spry2 alone caused loss of the anterior vermis, reducing FGF signaling further, by decreasing Fgf8 gene dose, resulted in loss of the entire vermis. Our data suggest that cell death is not responsible for vermis loss, but rather that it fails to develop because reducing FGF signaling perturbs the balance between vermis and roof plate development in rhombomere 1. We suggest a molecular explanation for this phenomenon by providing evidence that FGF signaling functions to inhibit the BMP signaling that promotes roof plate development.     

Characterization of five novel Pseudomonas aeruginosa cell-surface signalling systems

Cell-surface signalling is a sophisticated regulatory mechanism used by Gram-negative bacteria to sense signals from outside the cell and transmit them into the cytoplasm. This regulatory system consists of an outer membrane-localized TonB-dependent receptor (TonB-dependent transducer), a cytoplasmic membrane-localized antisigma factor and an extracytoplasmic function (ECF) sigma factor. Pseudomonas aeruginosa contains 13 potential surface signalling systems of which only six have been studied in detail. In this work we have identified the regulons of five novel P. aeruginosa signalling systems. For that, the ECF sigmas PA0149, PA1912, PA2050, PA2093 and PA4896 have been overexpressed and their target gene candidates have been identified using DNA microarray, proteomic analysis, and/or lacZ reporter construct. All five ECF sigma factors control the production of one TonB-dependent transducer. Interestingly, two sigma factors, PA2050 and PA2093, regulate the synthesis of a second transducer. Furthermore, we show that although all these sigma factors seem to control putative (metal) transport systems, one of them also regulates the expression of P. aeruginosa pyocins. Finally, we also show that the PA1912-PA1911-PA1910 (designated FemI-FemR-FemA in this work) signalling system responds to the presence of the Mycobacterium siderophores mycobactin and carboxymycobactin and is involved in the utilization of these heterologous siderophores.     

Synergistic effect of green tea extract and probiotics on the pathogenic bacteria, <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> and <Emphasis Type="Italic">Streptococcus pyogenes</Emphasis>

The aim of this study was to assess the effect of a commercial green tea extract (TEAVIGO™) on the microbial growth of three probiotic strains (Lactobacillus and Bifidobacterium), as well as three pathogenic bacteria. MIC and co-culture studies were performed. The MICs of the green tea extract against Staphylococcus aureus and Streptococcus pyogenes (100 μg ml⁻¹) were considerably lower than those against the probiotic strains tested (>800 μg ml⁻¹) and Escherichia coli (800 μg ml⁻¹). In co-culture studies, a synergistic effect of the probiotic strains and the green tea extract was observed against both Staph. aureus and Strep. pyogenes. Green tea extract in combination with probiotics significantly reduced the viable count of both pathogens at 4 h and by 24 h had completely abolished the recovery of viable Staph. aureus and Strep. pyogenes. These reductions were more significant than the reductions induced by probiotics or green tea extracts used separately. These results demonstrate the potential for combined therapy using the green tea extract plus probiotics on microbial infections caused by Staph. aureus and Strep. pyogenes. As probiotics and the green tea extract are derived from natural products, treatment with these agents may represent important adjuncts to, or alternatives to, conventional antibiotic therapy.     

Role of caspase 2 in apoptotic signaling in primate and murine germ cells

This study investigates the role of caspase 2 in apoptotic signaling of nonhuman primate male germ cells triggered by mild testicular hyperthermia, testosterone (T(e)) implants, or by combined interventions. Mean incidence of germ cell apoptosis increased significantly by Day 3 in the heat (H(e)) alone group and by Day 8 in the Te alone group but peaked at Day 3 in H(e) + T(e) group. We found activation of caspase 2 in both germ cells and Sertoli cells after induction of apoptosis. Most notably, active caspase 2 immunoreactivity was detected only in those germ cells susceptible to apoptosis compared with controls, where little or no such staining is detected. To further explore the role of caspase 2 in regulating male germ cell death, we next evaluated the efficacy of caspase 2 inhibition in preventing or attenuating heat-induced germ cell apoptosis in rats. Caspase 2 inhibition significantly (P < 0.05) prevented such heat-induced germ cell apoptosis. The protection offered by the caspase 2 inhibitor occurred upstream of mitochondria, involving suppression of mitogen-activated protein kinase (MAPK) 14 activation and inducible nitric oxide synthase (NOS2) induction and, in turn, suppression of cytochrome c-mediated death pathway. Together, our results show that caspase 2 is activated in male germ cells undergoing apoptosis in nonhuman primates after heat stress, hormonal deprivation, or after combined interventions. Blockade of caspase 2 activation prevents heat-induced germ cell apoptosis in rats by suppressing the MAPK14- and NO-mediated intrinsic pathway signaling.     

Seasonal activity of non-specific acid and alkaline phosphatases in beech (<Emphasis Type="Italic">Fagus sylvatica</Emphasis>) leaves

The activities of acid and alkaline phosphatases along with phosphorus content in leaves of European beech (Fagus sylvatica L.) were studied for a period from April to October. The phosphorus content of beech leaves was highest in April, at the beginning of the vegetation period; from May to October it was twofold lower than in April. Acid phosphatase activity (per unit fresh weight) in leaves collected from the middle part of the crown decreased significantly in May and July compared to the enzyme activity in April. In both the low and middle parts of the crown, the acid phosphatase activity had a peak in August, and thereafter decreased in September and October. No correlations between acid phosphatase activity and phosphorus concentrations were found. Alkaline phosphatase activity was very low and in some cases near the detection limit during the whole observation period.     

Windkesselness of coronary arteries hampers assessment of human coronary wave speed by single-point technique

A novel single-point technique to calculate local arterial wave speed (SPc) has recently been presented and applied in healthy human coronary arteries at baseline flow. We investigated its applicability for conditions commonly encountered in the catheterization laboratory. Intracoronary pressure (P(d)) and Doppler velocity (U) were recorded in 29 patients at rest and during adenosine-induced hyperemia in a distal segment of a normal reference vessel and downstream of a single stenosis before and after revascularization. Conduit vessel tone was minimized with nitroglycerin. Microvascular resistance (MR) and SPc were calculated from P(d) and U. In the reference vessel, SPc decreased from 21.5 m/s (SD 8.0) to 10.5 m/s (SD 4.1) after microvascular dilation (P < 0.0001). SPc was substantially higher in the presence of a proximal stenosis and decreased from 34.4 m/s (SD 18.2) at rest to 27.5 m/s (SD 13.4) during hyperemia (P < 0.0001), with a concomitant reduction in P(d) by 20 mmHg and MR by 55.4%. The stent placement further reduced hyperemic MR by 26% and increased P(d) by 26 mmHg but paradoxically decreased SPc to 13.1 m/s (SD 7.7) (P < 0.0001). Changes in SPc correlated strongly with changes in MR (P < 0.001) but were inversely related to changes in P(d) (P < 0.01). In conclusion, the single-point method yielded erroneous predictions of changes in coronary wave speed induced by a proximal stenosis and distal vasodilation and is therefore not appropriate for estimating local wave speed in coronary vessels. Our findings are well described by a lumped reservoir model reflecting the "windkesselness" of the coronary arteries.     

Mouse heart valve structure and function: echocardiographic and morphometric analyses from the fetus through the aged adult

The purpose of this study is to provide standard echocardiographic and morphometric data for normal mouse valve structure and function from late fetal to aged adult stages. Cross-sectional, two-dimensional and Doppler transthoracic echocardiography was performed in C57BL6 mice anesthetized with 1% to 2% isoflurane at embryonic day 18.5 (late fetal), 10 days (neonate), 1 mo (juvenile), 2 mo (young adult), 9 mo (old adult), and 16 mo (aged adult). Normal annulus dimensions indexed to age or weight, and selected flow velocities, were established by echocardiography. After echocardiographic imaging, hearts were harvested and histological and morphometric analyses were performed. Morphometric analysis demonstrated a progressive valve thinning and elongation during the fetal and juvenile stages that plateaued during adult stages (ANOVA, P < 0.01); however, there was increased thickening of the hinge of the aortic valve with advanced age, reminiscent of human aortic valve sclerosis. There was no age-related calcification. The results of this study provide comprehensive echocardiographic and morphometric data for normal mouse valve structure and function from late fetal to aged adult stages and should prove useful as a reference standard for future studies using mouse models of progressive valve disease.     

Maintaining phase of the crustacean tri-phasic pyloric rhythm

We construct and analyze a model network of the pyloric rhythm of the crustacean stomatogastric ganglion consisting of an oscillator neuron that inhibits two reciprocally inhibitory follower neurons. We derive analytic expressions that determine the phase of firing of the follower neurons with respect to the oscillator. An important aspect of the model is the inclusion of synapses that exhibit short-term synaptic depression. We show that these type of synapses allow there to be a complicated relationship between the intrinsic properties of the neurons and the synapses between them in determining phase relationships. Our analysis reveals the circumstances and ranges of cycle periods under which these properties work in concert with or independently from one another. In particular, we show that phase maintenance over a range of oscillator periods can be enhanced through the interplay of the two follower neurons if the synapses between these neurons are depressing. Since our model represents the core of the oscillatory pyloric network, the results of our analysis can be compared to experimental data and used to make predictions about the biological network.     

Liver fat, visceral adiposity, and sleep disturbances contribute to the development of insulin resistance and glucose intolerance in nondiabetic dialysis patients

Hemodialysis patients exhibit insulin resistance (IR) in target organs such as liver, muscles, and adipose tissue. The aim of this study was to identify contributors to IR and to develop a model for predicting glucose intolerance in nondiabetic hemodialysis patients. After a 2-h, 75-g oral glucose tolerance test (OGTT), 34 hemodialysis patients were divided into groups with normal (NGT) and impaired glucose tolerance (IGT). Indices of insulin sensitivity were derived from OGTT data. Measurements included liver and muscle fat infiltration and central adiposity by computed tomography scans, body composition by dual energy X-ray absorptiometer, sleep quality by full polysomnography, and functional capacity and quality of life (QoL) by a battery of exercise tests and questionnaires. Cut-off points, as well as sensitivity and specificity calculations were based on IR (insulin sensitivity index by Matsuda) using a receiver operator characteristics (ROC) curve analysis. Fifteen patients were assigned to the IGT, and 19 subjects to the NGT group. Intrahepatic fat content and visceral adiposity were significantly higher in the IGT group. IR indices strongly correlated with sleep disturbances, visceral adiposity, functional capacity, and QoL. Visceral adiposity, O2 desaturation during sleep, intrahepatic fat content, and QoL score fitted into the model for predicting glucose intolerance. A ROC curve analysis identified an intrahepatic fat content of > 3.97% (sensitivity, 100; specificity, 35.7) as the best cutoff point for predicting IR. Visceral and intrahepatic fat content, as well as QoL and sleep seemed to be involved at some point in the development of glucose intolerance in hemodialysis patients. Means of reducing fat depots in the liver and splachnic area might prove promising in combating IR and cardiovascular risk in hemodialysis patients.