An updated 18S rRNA phylogeny of tunicates based on mixture and secondary structure models

Background  

Tunicates have been recently revealed to be the closest living relatives of vertebrates. Yet, with more than 2500 described species, details of their evolutionary history are still obscure. From a molecular point of view, tunicate phylogenetic relationships have been mostly studied based on analyses of 18S rRNA sequences, which indicate several major clades at odds with the traditional class-level arrangements. Nonetheless, substantial uncertainty remains about the phylogenetic relationships and taxonomic status of key groups such as the Aplousobranchia, Appendicularia, and Thaliacea.     

Analysis of the life cycle of the soil saprophyte Bacillus cereus in liquid soil extract and in soil

Bacillus is commonly isolated from soils, with organisms of Bacillus cereus sensu lato being prevalent. Knowledge of the ecology of B. cereus and other Bacillus species in soil is far from complete. While the older literature favors a model of growth on soil-associated organic matter, the current paradigm is that B. cereus sensu lato germinates and grows in association with animals or plants, resulting in either symbiotic or pathogenic interactions. An in terra approach to study soil-associated bacteria is described, using filter-sterilized soil-extracted soluble organic matter (SESOM) and artificial soil microcosms (ASM) saturated with SESOM. B. cereus ATCC 14579 displayed a life cycle, with the ability to germinate, grow, and subsequently sporulate in both the liquid SESOM extract and in ASM inserted into wells in agar medium. Cells grew in liquid SESOM without separating, forming multicellular structures that coalesced to form clumps and encasing the ensuing spores in an extracellular matrix. Bacillus was able to translocate from the point of inoculation through soil microcosms as shown by the emergence of outgrowths on the surrounding agar surface. Microscopic inspection revealed bundles of parallel chains inside the soil. The motility inhibitor L-ethionine failed to suppress outgrowth, ruling out translocation by a flagellar-mediated mechanism such as swimming or swarming. Bacillus subtilis subsp. subtilis Marburg and four Bacillus isolates taken at random from soils also displayed a life cycle in SESOM and ASM and were all able to translocate through ASM, even in presence of L-ethionine. These data indicate that B. cereus is a saprophytic bacterium that is able to grow in soil and furthermore that it is adapted to translocate by employing a multicellular mode of growth.     

Identification of structural and functional O-linked N-acetylglucosamine-bearing proteins in Xenopus laevis oocyte

O-Linked N-acetylglucosaminylation (O-GlcNAcylation) (or O-linked N-acetylglucosamine (O-GlcNAc)) is an abundant and reversible glycosylation type found within the cytosolic and the nuclear compartments. We have described previously the sudden O-GlcNAcylation increase occurring during the Xenopus laevis oocyte G(2)/M transition, and we have demonstrated that the inhibition of O-GlcNAc-transferase (OGT) blocked this process, showing that the O-GlcNAcylation dynamism interferes with the cell cycle progression. In this work, we identified proteins that are O-GlcNAc-modified during the G(2)/M transition. Because of a low expression of O-GlcNAcylation in Xenopus oocyte, classical enrichment of O-GlcNAc-bearing proteins using O-GlcNAc-directed antibodies or wheat germ agglutinin lectin affinity were hard to apply, albeit these techniques allowed the identification of actin and erk2. Therefore, another strategy based on an in vitro enzymatic labeling of O-GlcNAc residues with azido-GalNAc followed by a chemical addition of a biotin alkyne probe and by enrichment of the tagged proteins on avidin beads was used. Bound proteins were analyzed by nano-LC-nano-ESI-MS/MS allowing for the identification of an average of 20 X. laevis oocyte O-GlcNAcylated proteins. In addition to actin and beta-tubulin, we identified metabolic/functional proteins such as PP2A, proliferating cell nuclear antigen, transitional endoplasmic reticulum ATPase, aldolase, lactate dehydrogenase, and ribosomal proteins. This labeling allowed for the mapping of a major O-GlcNAcylation site within the 318-324 region of beta-actin. Furthermore immunofluorescence microscopy enabled the direct visualization of O-GlcNAcylation and OGT on the meiotic spindle as well as the observation that chromosomally bound proteins were enriched in O-GlcNAc and OGT. The biological relevance of this post-translational modification both on microtubules and on chromosomes remains to be determined. However, the mapping of the O-GlcNAcylation sites will help to underline the function of this post-translational modification on each identified protein and will provide a better understanding of O-GlcNAcylation in the control of the cell cycle.     

Influence of Tertiary paleoenvironmental changes on the diversification of South American mammals: a relaxed molecular clock study within xenarthrans

Background  

Comparative genomic data among organisms allow the reconstruction of their phylogenies and evolutionary time scales. Molecular timings have been recently used to suggest that environmental global change have shaped the evolutionary history of diverse terrestrial organisms. Living xenarthrans (armadillos, anteaters and sloths) constitute an ideal model for studying the influence of past environmental changes on species diversification. Indeed, extant xenarthran species are relicts from an evolutionary radiation enhanced by their isolation in South America during the Tertiary era, a period for which major climate variations and tectonic events are relatively well documented.     

Proliferative activity in the cerebellar external granular layer evaluated by bromodeoxyuridine labeling

We have optimised an indirect immunoperoxidase technique demonstrating bromodeoxyuridine (BrdU) incorporation into dividing cells for cerebellar tissue sections of four-day-old rats injected with this marker. This permits confident identification of granule-cell precursors engaged in DNA synthesis in the external granular layer of the developing cerebellum. Preservation of BrdU immunoreactivity is attained using methanol/acetic acid fixation and different pretreatments before immunostaining, while unlabeled nuclei can be recognized clearly after Feulgen or hematoxylin counterstaining. We established conditions to ensure satisfactory BrdU uptake without affecting cell-cycle progression during the postlabeling time period. The dose of BrdU employed provides saturation S-phase labeling from at least 1 h after BrdU delivery. Various kinetic parameters and phase durations have been determined in experiments involving a single injection or cumulative labeling sequences, and the cycle time was calculated based on two models of generative behavior: steady-state and exponential growth. The working hypothesis of steadystate kinetics can be adopted successfully if the existence of neuroblasts with different proliferation rates is taken into account.     

GMDD: a database of GMO detection methods

Background  

Since more than one hundred events of genetically modified organisms (GMOs) have been developed and approved for commercialization in global area, the GMO analysis methods are essential for the enforcement of GMO labelling regulations. Protein and nucleic acid-based detection techniques have been developed and utilized for GMOs identification and quantification. However, the information for harmonization and standardization of GMO analysis methods at global level is needed.     

Interference of the PAF-acether antagonist BN 52021 with passive anaphylaxis in the guinea-pig

PAF-acether may be involved in anaphylaxis and asthma. We tested the new PAF-acether antagonist BN 52021 against the effects of antigen in passively sensitized guinea-pigs. Bronchoconstriction by ovalbumin administered i.v. (1 mg/kg) or by aerosol (1 or 10 mg/ml for a period of 1 min) was significantly reduced by BN 52021 (1-10 mg/kg), which did not inhibit drop of leukocyte counts after the i.v. challenge. In both cases, when the guinea-pigs were pretreated by propranolol, high amounts of BN 52021 became ineffective against shock. The reduction of the anaphylactic bronchoconstriction, induced by the combination of mepyramine, aspirin and FPL 55712 was not improved by BN 52021. Tested on isolated lung strips from passively sensitized guinea-pig, BN 52021, at a concentration which inhibits PAF-induced contraction (0.1 mM), did not inhibit the anaphylactic contraction triggered by the administration of ovalbumin (10 micrograms/ml) nor the accompanying release of histamine and thromboxane. In contrast, BN 52021 (30 microM) significantly reduced the anaphylactic release of histamine and of thromboxane from perfused lungs of passively sensitized guinea-pigs. The results with the isolated lung strips and the propranolol-treated guinea-pigs in vivo suggest a dissociation between the anti-anaphylactic and the anti-PAF-acether properties of BN 52021.     

Biofilm proteome: homogeneity or versatility?

The problems associated with biofilm infections in humans result from the distinct characteristics of biofilms, in particular their high level of resistance to antibiotics. One of the hypotheses that have been advanced to explain this resistance to antimicrobials is the phenotypic differentiation of biofilm cells. Although many studies on biofilms have highlighted physiological alterations following the attachment of bacteria to a surface, no studies have explicitly demonstrated a "biofilm" physiology. To contribute to this topical debate, we used principal component analysis to interpret spot quantity variations observed on electropherograms obtained by two-dimensional gel electrophoresis of crude protein extracts from planktonic and sessile Pseudomonas aeruginosa cells. These analyses showed that the proteome of attached P. aeruginosa cells differs from that of their planktonic counterparts. Furthermore, we found that the proteome of sessile P. aeruginosa is strongly dependent on the nature of the biofilm substratum.