Genetic distinctions among clinical and environmental strains of Vibrio vulnificus

Vibrio vulnificus causes rare but frequently fatal septicemia associated with raw oyster consumption by persons with underlying hepatic or immune system dysfunction. The virulence potential of environmental reservoirs appears widely distributed, because most strains are virulent in animal models; however, several investigations recently demonstrated genetic divergence among strains from clinical versus environmental origin at independent genetic loci. The present study used PCR to screen DNA polymorphisms in strains from environmental (n = 35) or clinical (n = 33) sources, and genomic relationships were determined by repetitive extragenic palindromic DNA PCR (rep-PCR) typing. Significant (P < 0.01) association was observed for typical "clinical" or "environmental" polymorphism profiles based on strain origin. Most oyster isolates (88%), including all of those with the "environmental" profile, also formed a single rep-PCR genogroup. Clinical isolates within this group did not have the typical "clinical" profile. On the other hand, clinical isolates with the typical polymorphism profile were distributed among multiple rep-PCR genogroups, demonstrating greater genetic diversity than was evident by profiling genetic polymorphisms. Wound isolates were genetically distinct from typical blood isolates by all assays. Strains from an outbreak of wound infections in Israel (biotype 3) were closely related to several U.S. strains by rep-PCR, indicating potential reservoirs of emerging disease. Strains genetically related to blood isolates appeared to be relatively rare in oysters, as only one had the "clinical" polymorphism profile or clustered by rep-PCR. However, this study was not an extensive survey, and more sampling using rep-PCR for sensitive genetic discrimination is needed to determine the virulence potential of environmental reservoirs.     

Direct evidence that receptor site-4 of sodium channel gating modifiers is not dipped in the phospholipid bilayer of neuronal membranes

In a recent note to Nature, R. MacKinnon has raised the possibility that potassium channel gating modifiers are able to partition in the phospholipid bilayer of neuronal membranes and that by increasing their partial concentration adjacent to their receptor, they affect channel function with apparent high affinity (Lee and MacKinnon (2004) Nature 430, 232-235). This suggestion was adopted by Smith et al. (Smith, J. J., Alphy, S., Seibert, A. L., and Blumenthal, K. M. (2005) J. Biol. Chem. 280, 11127-11133), who analyzed the partitioning of sodium channel modifiers in liposomes. They found that certain modifiers were able to partition in these artificial membranes, and on this basis, they have extrapolated that scorpion beta-toxins interact with their channel receptor in a similar mechanism as that proposed by MacKinnon. Since this hypothesis has actually raised a new conception, we examined it in binding assays using a number of pharmacologically distinct scorpion beta-toxins and insect and mammalian neuronal membrane preparations, as well as by analyzing the rate by which the toxin effect on gating of Drosophila DmNa(v)1 and rat brain rNa(v)1.2a develops. We show that in general, scorpion beta-toxins do not partition in neuronal membranes and that in the case in which a depressant beta-toxin partitions in insect neuronal membranes, this partitioning is unrelated to its interaction with the receptor site and the effect on the gating properties of the sodium channel. These results negate the hypothesis that the high affinity of beta-toxins for sodium channels is gained by their ability to partition in the phospholipid bilayer and clearly indicate that the receptor site for scorpion beta-toxins is accessible to the extracellular solvent.     

Mechanics in embryogenesis and embryonics: prime mover or epiphenomenon?

Mechanics is shown to be an important, perhaps central component to the differentiation and development of embryos. Mechanics of the nucleus may also be involved in determining which genes are expressed in a given cell. There are two major approaches at present to the mechanics of differentiation in embryos: morphomechanics and differentiation waves. These are compared in detail, to provide a starting point for future experimental work to bring them into one conceptual framework. This may rationalize the present cookbookery of stem cell production by placing it in the context of differentiation waves and the differentiation code. Embryonics, the realization of concepts from embryology in computer hardware and software, might be considerably enhanced by incorporating mechanical concepts of embryogenesis. Segmented robots, modular robotics, cellular microrobotics, flexible electronics, wearable computers, diatom nanotechnology and waves in active media point to a synthesis that we could call embryonic robotics.     

Preface. Developmental Morphodynamics - bridging the gap between the genome and embryo physics.

We, the Guest Editors of this Special Issue of The International Journal of Developmental Biology, are two older embryologists, who are trying to bridge the current chasm between Entwicklungsmechanic, the developmental mechanics of our embryogenesis forefathers, and the modern movement of molecular developmental biology. Our rallying cry is that of Wilhelm His: "To think that heredity will build organic beings without mechanical means is a piece of unscientific mysticism" (His, 1888). Until recently, this claim appeared to us to fall on the somewhat deaf ears of molecular developmental biologists. Yet, the world is still one, and both physics and chemistry obviously have their place in embryogenesis. Indeed, at the molecular level, membrane proteins which are mechanoreceptors and motor molecules may begin to point the way. Here, we and our colleagues will make the case for a more equitable consideration of molecules and mechanics.     

Structural and dynamic characterization of a vinculin binding site in the talin rod

Talin is a key protein involved in linking integrins to the actin cytoskeleton. The long flexible talin rod domain contains a number of binding sites for vinculin, a cytoskeletal protein important in stabilizing integrin-mediated cell-matrix junctions. Here we report the solution structure of a talin rod polypeptide (residues 1843-1973) which contains a single vinculin binding site (VBS; residues 1944-1969). Like other talin rod polypeptides, it consists of a helical bundle, in this case a four-helix bundle with a right-handed topology. The residues in the VBS important for vinculin binding were identified by studying the binding of a series of VBS-related peptides to the vinculin Vd1 domain. The key binding determinants are buried in the interior of the helical bundle, suggesting that a substantial structural change in the talin polypeptide is required for vinculin binding. Direct evidence for this was obtained by NMR and EPR spectroscopy. [1H,15N]-HSQC spectra of the talin fragment indicate that vinculin binding caused approximately two-thirds of the protein to adopt a flexible random coil. For EPR spectroscopy, nitroxide spin labels were attached to the talin polypeptide via appropriately located cysteine residues. Measurements of inter-nitroxide distances in doubly spin-labeled protein showed clearly that the helical bundle is disrupted and the mobility of the helices, except for the VBS helix, is markedly increased. Binding of vinculin to talin is thus a clear example of the unusual phenomenon of protein unfolding being required for protein/protein interaction.     

Temperature and the respiratory properties of whole blood in two reptiles, Pogona barbata and Emydura signata

We investigated the capacity of two reptiles, an agamid lizard Pogona barbata and a chelid turtle Emydura signata, to compensate for the effects of temperature by making changes in their whole blood respiratory properties. This was accomplished by measuring the P50 (at 10, 20 and 30 degrees C), hematocrit (Hct), haemoglobin concentration ([Hb]) and mean cell haemoglobin concentration (MCHC) in field acclimatised and laboratory acclimated individuals. The acute effect of temperature on P50 in P. barbata, expressed as heat of oxygenation (deltaH), ranged from -16.8+/-1.84 to -28.5+/-2.73 kJ/mole. P50 of field acclimatised P. barbata increased significantly from early spring to summer at the test temperatures of 20 degrees C (43.1+/-1.2 to 48.8+/-2.1 mmHg) and 30 degrees C (54.7+/-1.2 to 65.2+/-2.3 mmHg), but showed no acclimation under laboratory conditions. For E. signata, deltaH ranged from -31.1+/-6.32 to -48.2+/-3.59 kJ/mole. Field acclimatisation and laboratory acclimation of P50 did not occur. However, in E. signata, there was a significant increase in [Hb] and MCHC from early spring to summer in turtles collected from the wild (1.0+/-0.1 to 1.7+/-0.2 mmol/L and 4.0+/-0.3 to 6.7+/-0.7 mmol/L, respectively).     

Assessing stability of gene selection in microarray data analysis

Background  

The number of genes declared differentially expressed is a random variable and its variability can be assessed by resampling techniques. Another important stability indicator is the frequency with which a given gene is selected across subsamples. We have conducted studies to assess stability and some other properties of several gene selection procedures with biological and simulated data.