Biodistribution of 137Cs in a mouse model of chronic contamination by ingestion and effects on the hematopoietic system

The aim of this work was to define the possible occurrence of hematological changes during the course of a chronic ingestion of ¹³⁷Cs. A mouse model was used, with ingestion through drinking water with a cesium concentration of 20 kBq l⁻¹. Ingestion started in parent animals before mating, and ¹³⁷Cs intake and its effect on the hematopoietic system was studied in offspring at various ages between birth and 20 weeks. ¹³⁷Cs content was measured in various organs, indicating that ¹³⁷Cs was distributed throughout the organism including lympho-hematopoietic organs, i.e., femurs, spleen and thymus. However, we did not observe any effect on the hematopoietic system, whatever the parameter used. In fact, blood cell counts, mononuclear cell counts and progenitor frequency in bone marrow and spleen, and Flt3-ligand, Erythropoietin, G-CSF and SDF-1 concentration in plasma remained unchanged when compared to control animals. Moreover, phenotypic analysis did not show any change in the proportions of bone marrow cell populations. These results indicate that, although ¹³⁷Cs was found in all organs implicated in the hematopoietic system, this did not induce any changes in bone marrow function.     

Association of specific expansins with growth in maize leaves is maintained under environmental, genetic, and developmental sources of variation

We aimed to evaluate whether changes in maize (Zea mays) leaf expansion rate in response to environmental stimuli or developmental gradients are mediated by common or specific expansins, a class of proteins known to enhance cell wall extensibility. Among the 33 maize expansin or putative expansin genes analyzed, 19 were preferentially expressed at some point of the leaf elongation zone and these expansins could be organized into three clusters related to cell division, maximal leaf expansion, and cell wall differentiation. Further analysis of the spatial distribution of expression was carried out for three expansins in leaves displaying a large range of expansion rates due to water deficit, genotype, and leaf developmental stage. With most sources of variation, the three genes showed similar changes in expression and consistent association with changes in leaf expansion. Moreover, our analysis also suggested preferential association of each expansin with elongation, widening, or both of these processes. Finally, using in situ hybridization, expression of two of these genes was increased in load-bearing tissues such as the epidermis and differentiating xylem. Together, these results suggest that some expansins may be preferentially related to elongation and widening after integrating several spatial, environmental, genetic, and developmental cues.     

Cyathane diterpenes from Sarcodon cyrneus and evaluation of their activities of neuritegenesis and nerve growth factor production

Two new cyathane diterpenes, cyrneine C (4) and D (5), were isolated from the mushroom Sarcodon cyrneus, along with previously isolated cyrneine A, B and glaucopine C. The structures of the novel diterpenoids were determined by the analysis of spectroscopic data. Effects of the cyrneines and glaucopine C on the NGF gene expression in 1321N1 cells and on neurite outgrowth on PC12 cells were evaluated.     

Synthesis, pharmacological evaluation and electrochemical studies of novel 6-nitro-3,4-methylenedioxyphenyl-N-acylhydrazone derivatives: Discovery of LASSBio-881, a new ligand of cannabinoid receptors

We describe herein the discovery of LASSBio-881 (3c) as a novel in vivo antinociceptive, anti-inflammatory, and in vitro antiproliferative and antioxidant compound, with a cannabinoid ligand profile. We observed that LASSBio-881 (3c) was able to bind to CB1 receptors (71% at 100microM) and also to inhibit T-cell proliferation (66% at 10microM) probably by binding to CB2 receptors, in a non-proapoptotic manner, different from anandamide (1). It was also demonstrated that LASSBio-881 (3c) had an important antioxidant profile toward free radicals (DPPH and hydroxyl), probably due to its particular redox behavior, which reflects the presence of both nitro and 3,5-di-tert-butyl-4-hydroxyphenyl sub-units, as demonstrated by cyclic voltammetry studies. In addition, we showed that these structural sub-units are essential for the observed pharmacological activity.     

The primate community of Cachoeira (Brazilian Amazonia): a model to decipher ecological partitioning among extinct species

Dental microwear analysis is conducted on a community of platyrrhine primates from South America. This analysis focuses on the primate community of Cachoeira Porteira (Para, Brazil), in which seven sympatric species occur: Alouatta seniculus, Ateles paniscus, Cebus apella, Chiropotes satanas, Pithecia Pithecia, Saguinus midas, and Saimiri sciureus. Shearing quotients are also calculated for each taxon of this primate community. Dental microwear results indicate significant differences between taxa, but are somewhat insufficient when it comes to discriminating between ecologically similar taxa. The primates of Cachoeira Porteira all incorporate a certain amount of fruit in their diet, entailing a definite amount of inter-specific competition as they must share food resources. Alouatta is the most folivorous taxon of this community, which is corroborated by dental microwear analysis. Ateles, although of a similar size to Alouatta, limits inter-specific competition by incorporating more fruit in its diet. Cebus has a very diverse omnivorous diet, which is highlighted in this study, as it compares to both fruit and leaf eating taxa. In some cases, microwear results need to be supplemented by other methods. For example, dental microwear seems insufficient to distinguish between Pithecia and Chiropotes, which eat foods with similar physical properties. However, other methods (i.e. shearing quotients and body mass) provide enough complimentary information to be able to highlight differences between the two taxa. On the other hand, dental microwear can highlight differences between primates which have similar diets, such as Saimiri and Saguinus. In this case, differences could be due to other exogenous factors.     

Epitope mapping of the phosphorylation motif of the HIV-1 protein Vpu bound to the selective monoclonal antibody using TRNOESY and STD NMR spectroscopy

The conformational preferences of a 22-amino acid peptide (LIDRLIERAEDpSGNEpSEGEISA) that mimics the phosphorylated HIV-1-encoded virus protein U (Vpu) antigen have been investigated by NMR spectroscopy. Degradation of HIV receptor CD4 by the proteasome, mediated by the HIV-1 protein Vpu, is crucial for the release of fully infectious virions. Phosphorylation of Vpu at sites Ser52 and Ser56 on the DSGXXS motif is required for the interaction of Vpu with the ubiquitin ligase SCF(beta)(-TrCP) which triggers CD4 degradation by the proteasome. This motif is conserved in several signaling proteins known to be degraded by the proteasome. The interaction of the P-Vpu(41-62) peptide with its monoclonal antibody has been studied by transferred nuclear Overhauser effect NMR spectroscopy (TRNOESY) and saturation transfer difference NMR (STD NMR) spectroscopy. The peptide was found to adopt a bend conformation upon binding to the antibody; the peptide residues (Asp51-pSer56) forming this bend are recognized by the antibody as demonstrated by STD NMR experiments. The three-dimensional structure of P-Vpu(41-62) in the bound conformation was determined by TRNOESY spectra; the peptide adopts a compact structure in the presence of mAb with formation of several bends around Leu45 and Ile46 and around Ile60 and Ser61, with a tight bend created by the DpS(52)GNEpS(56) motif. STD NMR studies provide evidence for the existence of a conformational epitope containing tandem repeats of phosphoserine motifs. The peptide's epitope is predominantly located in the large bend and in the N-terminal segment, implicating bidentale association. These findings are in excellent agreement with a recently published NMR structure required for the interaction of Vpu with the SCF(beta)(-TrCP) protein.     

Molecular basis of inward rectification: polyamine interaction sites located by combined channel and ligand mutagenesis

Polyamines cause inward rectification of (Kir) K+ channels, but the mechanism is controversial. We employed scanning mutagenesis of Kir6.2, and a structural series of blocking diamines, to combinatorially examine the role of both channel and blocker charges. We find that introduced glutamates at any pore-facing residue in the inner cavity, up to and including the entrance to the selectivity filter, can confer strong rectification. As these negative charges are moved higher (toward the selectivity filter), or lower (toward the cytoplasm), they preferentially enhance the potency of block by shorter, or longer, diamines, respectively. MTSEA+ modification of engineered cysteines in the inner cavity reduces rectification, but modification below the inner cavity slows spermine entry and exit, without changing steady-state rectification. The data provide a coherent explanation of classical strong rectification as the result of polyamine block in the inner cavity and selectivity filter.     

NMR studies for identifying phosphopeptide ligands of the HIV-1 protein Vpu binding to the F-box protein beta-TrCP

The human immunodeficiency virus type 1 (HIV-1) Vpu enhances viral particle release and, its interaction with the ubiquitin ligase SCF-beta-TrCP triggers the HIV-1 receptor CD4 degradation by the proteasome. The interaction between beta-TrCP protein and ligands containing the phosphorylated DpSGXXpS motif plays a key role for the development of severe disease states, such as HIV or cancer. This study examines the binding and conformation of phosphopeptides (P1, LIERAEDpSG and P2, EDpSGNEpSE) from HIV protein Vpu to beta-TrCP with the objective of defining the minimum length of peptide needed for effective binding. The screening step can be analyzed by NMR spectroscopy, in particular, saturation transfer NMR methods clearly identify the residues in the peptide that make direct contact with beta-TrCP protein when bound. An analysis of saturation transfer difference (STD) spectra provided clear evidence that the two peptides efficiently bound beta-TrCP receptor protein. To better characterize the ligand-protein interaction, the bound conformation of the phosphorylated peptides was determined using transferred NOESY methods, which gave rise to a well-defined structure. P1 and P2 can fold in a bend arrangement for the DpSG motif, showing the protons identified by STD-NMR as exposed in close proximity at the molecule surface. Ser phosphorylation allows electrostatic interaction and hydrogen bond with the amino acids of the beta-TrCP binding pocket. The upstream LIER hydrophobic region was also essential in binding to a hydrophobic pocket of the beta-TrCP WD domain. These findings are in good agreement with a recently published X-ray structure of a shorter beta-Catenin fragment with the beta-TrCP complex.     

Hyperexpression of recombinant CFTR in heterologous cells alters its physiological properties

We investigated whether high levels of expression of the cysticfibrosis transmembrane conductance regulator (CFTR) would alter thefunctional properties of newly synthesized recombinant proteins. COS-7,CFPAC-1, and A549 cells were intranuclearly injected with a Simianvirus 40-driven pECE-CFTR plasmid and assayed for halide permeabilityusing the6-methoxy-N-(3-sulfopropyl)quinolinium fluorescent probe. With increasing numbers of microinjected pECE-CFTR copies, the baseline permeability to halide dose dependently increased, and the response to adenosine 3',5'-cyclic monophosphate(cAMP) stimulation decreased. In cells hyperexpressing CFTR, the high level of halide permeability was reduced when a cell metabolism poisoning cocktail was applied to decrease intracellular ATP and, inversely, was increased by orthovanadate. In CFPAC-1 cellsinvestigated with the patch-clamp technique, CFTR hyperexpression ledto a time-independent nonrectifying chloride current that was notsensitive to cAMP stimulation. CFPAC-1 cells hyperexpressing CFTRexhibited no outward rectifying chloride current nor inward rectifyingpotassium current either spontaneously or under cAMP stimulation. Weconclude that hyperexpression of recombinant CFTR proteins modifiestheir properties inasmuch as 1) CFTRchannels are permanently activated and not susceptible to cAMPregulation and 2) they lose their capacity to regulate heterologous ionic channels.