Phylogenetic relationships between environmental and clinical isolates of Pseudomonas fluorescens and related species deduced from 16S rRNA gene and OprF protein sequences

The major surface protein of the genus Pseudomonas, OprF, is a non-specific porin that plays an important role in maintenance of cell shape, in growth in a low osmolarity environment, and in adhesion to various supports. The objectives of our study were (i) to carry out a comparative analysis of phylogenies obtained from the OprF protein and from the 16S rRNA gene in 41 isolates from various sources (water, soil, milk and the hospital) and (ii) to investigate the physiological characteristics correlated with the phylogeny of OprF. We report here an important incongruence between the phylogenies of the 16S rRNA gene and the OprF protein. Phylogenetic analysis of 16S rRNA genes grouped Pseudomonas fluorescens isolates into one cluster (termed fluorescens r-cluster) whilst the phylogeny of the OprF protein divided Pseudomonas fluorescens isolates into two quite distinct clusters (termed fluorescens 1 o-cluster and fluorescens 2 o-cluster) that may be related to the original habitat of the strain. The fluorescens 1 o-cluster contained the majority of non-rhizospheric soil isolates, while the fluorescens 2 o-cluster contained all our clinical isolates and most of the rhizospheric isolates (which are fixed to the roots). In order to check this correlation, we studied two physiological characteristics: the range of growth temperature and the capacity for non-specific adhesion to polystyrene. The temperature range study for strains did not explain the existence of the two o-clusters but it did confirm the capacity of certain P. fluorescens strains to grow at 37 degrees C. The adhesion capacities of the isolates in the two o-clusters seems to be correlated with ecological niche.     

A Model of the Structural and Functional Development of the Normal Human Fetal Left Ventricle Based on a Global Growth Law

The purpose of this research is to study the growth of the normal human left ventricle (LV) during the fetal period from 14 to 40 weeks of gestation. A new constitutive law for the active myocardium describing the mechanical properties of the active muscle during the whole cardiac cycle has been proposed. The LV model is a thick-walled, incompressible, hyperelastic cylinder, with families of helicoidal fibers running on cylindrical surfaces [1] . Based on the works of Lin and Taber [2] done on the embryonic chick heart, we use for the human fetal heart a growth law in which the growth rate depends on the wall stresses. The parameters of the growth law are adapted to agree with sizes and volumes inferred from two dimensional ultrasound measurements performed on 18 human fetuses. Then calculations are performed to extrapolate the cardiac performance during normal growth of the fetal LV. The results presented support the idea that a growth law in which the growth rate depends linearly on the mean wall stresses averaged through the space and during whole cardiac cycle, is adapted to the normal human fetal LV development.     

Chemical Composition and Antimicrobial Activity of the Essential Oils from New Caledonian Citrus macroptera and Citrus hystrix

The essential oils from the leaves of Citrus macroptera and C. hystrix, collected in New Caledonia, have been analyzed by gas chromatography/mass spectrometry (GC/MS) and evaluated for their antimicrobial activity. A total of 35 and 38 constituents were identified, representing 99.1 and 89.0% of the essential oils, respectively. Both essential oils were rich in monoterpenes (96.1 and 87.0%, resp.), with β‐pinene as major component (33.3 and 10.9%, resp.), and poor in limonene (2.4 and 4.7%, resp.). Other main components of C. macroptera oil were α‐pinene (25.3%), p‐cimene (17.6%), (E)‐β‐ocimene (6.7%), and sabinene (4.8%). The essential oil of C. hystrix was characterized by high contents of terpinen‐4‐ol (13.0%), α‐terpineol (7.6%), 1,8‐cineole (6.4%), and citronellol (6.0%). The antimicrobial activity was evaluated against five bacteria and five fungi strains. Both oils were inactive against bacteria. However, the C. macroptera leaf oil exhibited a pronounced activity against Trichophyton mentagrophytes var. interdigitale, with a minimal‐inhibitory concentration (MIC) of 12.5 μg/ml.     

Role of Water Molecules in Structure and Energetics of Pseudomonas aeruginosa Lectin I Interacting with Disaccharides

Calcium-dependent lectin I from Pseudomonas aeruginosa (PA-IL) binds specifically to oligosaccharides presenting an α-galactose residue at their nonreducing end, such as the disaccharides αGal1–2βGalOMe, αGal1–3βGalOMe, and αGal1–4βGalOMe. This provides a unique model for studying the effect of the glycosidic linkage of the ligands on structure and thermodynamics of the complexes by means of experimental and theoretical tools. The structural features of PA-IL in complex with the three disaccharides were established by docking and molecular dynamics simulations and compared with those observed in available crystal structures, including PA-IL·αGal1–2βGalOMe complex, which was solved at 2.4 Å resolution and reported herein. The role of a structural bridge water molecule in the binding site of PA-IL was also elucidated through molecular dynamics simulations and free energy calculations. This water molecule establishes three very stable hydrogen bonds with O6 of nonreducing galactose, oxygen from Pro-51 main chain, and nitrogen from Gln-53 main chain of the lectin binding site. Binding free energies for PA-IL in complex with the three disaccharides were investigated, and the results were compared with the experimental data determined by titration microcalorimetry. When the bridge water molecule was included in the free energy calculations, the simulations predicted the correct binding affinity trends with the 1–2-linked disaccharide presenting three times stronger affinity ligand than the other two. These results highlight the role of the water molecule in the binding site of PA-IL and indicate that it should be taken into account when designing glycoderivatives active against P. aeruginosa adhesion.     

Initiation of embryonic cardiac pacemaker activity by inositol 1,4,5-trisphosphate-dependent calcium signaling

In the adult, the heart rate is driven by spontaneous and repetitive depolarizations of pacemaker cells to generate a firing of action potentials propagating along the conduction system and spreading into the ventricles. In the early embryo before E9.5, the pacemaker ionic channel responsible for the spontaneous depolarization of cells is not yet functional. Thus the mechanisms that initiate early heart rhythm during cardiogenesis are puzzling. In the absence of a functional pacemaker ionic channel, the oscillatory nature of inositol 1,4,5-trisphosphate (InsP3)-induced intracellular Ca2+ signaling could provide an alternative pacemaking mechanism. To test this hypothesis, we have engineered pacemaker cells from embryonic stem (ES) cells, a model that faithfully recapitulates early stages of heart development. We show that InsP3-dependent shuttle of free Ca2+ in and out of the endoplasmic reticulum is essential for a proper generation of pacemaker activity during early cardiogenesis and fetal life.     

The In Vivo Association of BiP with Newly Synthesized Proteins Is Dependent on the Rate and Stability of Folding and Not Simply on the Presence of Sequences That Can Bind to BiP

Immunoglobulin heavy chain-binding protein (BiP) is a member of the hsp70 family of chaperones and one of the most abundant proteins in the ER lumen. It is known to interact transiently with many nascent proteins as they enter the ER and more stably with protein subunits produced in stoichiometric excess or with mutant proteins. However, there also exists a large number of secretory pathway proteins that do not apparently interact with BiP. To begin to understand what controls the likelihood that a nascent protein entering the ER will associate with BiP, we have examined the in vivo folding of a murine λI immunoglobulin (Ig) light chain (LC). This LC is composed of two Ig domains that can fold independent of the other and that each possess multiple potential BiP-binding sequences. To detect BiP binding to the LC during folding, we used BiP ATPase mutants, which bind irreversibly to proteins, as “kinetic traps.” Although both the wild-type and mutant BiP clearly associated with the unoxidized variable region domain, we were unable to detect binding of either BiP protein to the constant region domain. A combination of in vivo and in vitro folding studies revealed that the constant domain folds rapidly and stably even in the absence of an intradomain disulfide bond. Thus, the simple presence of a BiP-binding site on a nascent chain does not ensure that BiP will bind and play a role in its folding. Instead, it appears that the rate and stability of protein folding determines whether or not a particular site is recognized, with BiP preferentially binding to proteins that fold slowly or somewhat unstably.     

Cardiac specification of embryonic stem cells

Over the past decade, cell transplantation has been recognized as a mean of repairing infarcted myocardium. Both adult stem cells and differentiated cells have yielded encouraging results with regard to engraftment into postinfarction scars. However, these cells now feature serious restrictions. Asan alternative, embryonic stem (ES) cells are particularly attractive, because of their plasticity and the subsequent possibility to drive them towards a cardiomyogenic phenotype after exposure to appropriate growth factors. An additional theoretical advantage of ES cells is their expected immune privilege. In this article, we summarize the findings obtained in cell therapy using ES cells and discuss the molecular mechanisms of cardiac specification of the cells.     

The VP6 protein of rotavirus interacts with a large fraction of human naive B cells via surface immunoglobulins

Immunity to human group A rotavirus (RV), a major cause of viral gastroenteritis in infants, involves B lymphocytes that provide RV-specific antibodies. Additionally, some arguments suggest that naive B cells could be implicated in the first steps of the immune response against RV. The aim of our study was to analyze the interaction of VP6 and VP7 RV capsid proteins with human B cells depending on the immune status of the individual, i.e., naive or RV experienced. For this purpose, a two-color virus-like particle flow cytometry assay was devised to evaluate the blood B-lymphocyte reactivity to VP6 and VP7 proteins from healthy RV-exposed adults, recently infected infants, and neonates at birth. Both VP6 and VP7 interactions with B cells were mediated by surface immunoglobulins and probably by their Fab portions. VP7-reactive B lymphocytes were mainly detected from RV-experienced patients and almost exclusively in the CD27-positive memory cell fraction. Conversely, VP6-reactive B lymphocytes were detected at similar and high frequencies in adult, infant, and neonate samples. In adult samples, VP6 reacted with about 2% of the CD27-negative (CD27(neg)) naive B cells. These results demonstrated that the VP6 RV protein interacted with a large fraction of naive B lymphocytes from both adults and neonates. We propose that naive B cell-VP6 interaction might influence the strength and quality of the acquired immune response and should be considered for elaborating RV vaccine strategies.