The distribution of elements in the tissues of Watanabe Heritable Hyperlipidemic rabbits

A deficiency or an excess of some elements in the diet is reported to modify the concentration of cholesterol in plasma, and, conversely, a reduction of cholesterol in the diet decreases zinc in plasma. We have studied the distribution of elements Na, K, Ca, Mg, Fe, Cu, Zn, S, P, and Mn in the tissues, plasma, heart, aorta, lung, liver, spleen, kidney, thymus, and brain of New Zealand White rabbits (NZW) and of Watanabe Heritable Hyperlipidemic rabbits (WHHL). The WHHL rabbits had a massive hypercholesterolemia (7.45 +/- 1.2 g/L) induced by a lack of liver low density lipoprotein receptors. The concentrations of elements in the tissues of the control NZW rabbits were very similar to those found in the normal rat. In WHHL, compared to NZW, besides the very important increase of total phosphorus in plasma explained by the augmentation of phospholipids, there was an increase of plasma copper (+44%) and zinc (+36%). The other noticeable changes were an increase of iron in heart (+19%), sulfur, and zinc in liver (+15% and +18%). The other changes observed in WHHL rabbits were, besides the increase of ceruloplasmin, the increase of vit E (+468%) and MDA (+62%). In conclusion, despite a massive increase of lipids in plasma, there was no major disturbance of element distribution in WHHL rabbits.     

Characterization of chloroplastic fructose 1,6-bisphosphate aldolases as lysine-methylated proteins in plants

In pea (Pisum sativum), the protein-lysine methyltransferase (PsLSMT) catalyzes the trimethylation of Lys-14 in the large subunit (LS) of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), the enzyme catalyzing the CO(2) fixation step during photosynthesis. Homologs of PsLSMT, herein referred to as LSMT-like enzymes, are found in all plant genomes, but methylation of LS Rubisco is not universal in the plant kingdom, suggesting a species-specific protein substrate specificity of the methyltransferase. In this study, we report the biochemical characterization of the LSMT-like enzyme from Arabidopsis thaliana (AtLSMT-L), with a focus on its substrate specificity. We show that, in Arabidopsis, LS Rubisco is not naturally methylated and that the physiological substrates of AtLSMT-L are chloroplastic fructose 1,6-bisphosphate aldolase isoforms. These enzymes, which are involved in the assimilation of CO(2) through the Calvin cycle and in chloroplastic glycolysis, are trimethylated at a conserved lysyl residue located close to the C terminus. Both AtLSMT-L and PsLSMT are able to methylate aldolases with similar kinetic parameters and product specificity. Thus, the divergent substrate specificity of LSMT-like enzymes from pea and Arabidopsis concerns only Rubisco. AtLSMT-L is able to interact with unmethylated Rubisco, but the complex is catalytically unproductive. Trimethylation does not modify the kinetic properties and tetrameric organization of aldolases in vitro. The identification of aldolases as methyl proteins in Arabidopsis and other species like pea suggests a role of protein lysine methylation in carbon metabolism in chloroplasts.     

Microbial diversity of eolian dust sources from saline lake sediments and biological soil crusts in arid Southern Australia

While microbial communities of aerosols have been examined, little is known about their sources. Nutrient composition and microbial communities of potential dust sources, saline lake sediments (SLS) and adjacent biological soil crusts (BSC), from Southern Australia were determined and compared with a previously analyzed dust sample. Multivariate analyses of fingerprinting profiles indicated that the bacterial communities of SLS and BSC were different, and these differences were mainly explained by salinity. Nutrient concentrations varied among the sites but could not explain the differences in microbial diversity patterns. Comparison of microbial communities with dust samples showed that deflation selects against filamentous cyanobacteria, such as the Nostocales group. This could be attributed to the firm attachment of cyanobacterial filaments to soil particles and/or because deflation occurs mainly in disturbed BSC, where cyanobacterial diversity is often low. Other bacterial groups, such as Actinobacteria and the spore-forming Firmicutes, were found in both dust and its sources. While Firmicutes-related sequences were mostly detected in the SLS bacterial communities (10% of total sequences), the actinobacterial sequences were retrieved from both (11-13%). In conclusion, the potential dust sources examined here show highly diverse bacterial communities and contain nutrients that can be transported with aerosols. The obtained fingerprinting and sequencing data may enable back tracking of dust plumes and their microorganisms.     

Differential regulation of motor control and response to dopaminergic drugs by D1R and D2R neurons in distinct dorsal striatum subregions

The dorsal striatum is critically involved in a variety of motor behaviours, including regulation of motor activity, motor skill learning and motor response to psychostimulant and neuroleptic drugs, but contribution of D(2)R-striatopallidal and D(1)R-striatonigral neurons in the dorsomedial (DMS, associative) and dorsolateral (DLS, sensorimotor) striatum to distinct functions remains elusive. To delineate cell type-specific motor functions of the DMS or the DLS, we selectively ablated D(2)R- and D(1)R-expressing striatal neurons with spatial resolution. We found that associative striatum exerts a population-selective control over locomotion and reactivity to novelty, striatopallidal and striatonigral neurons inhibiting and stimulating exploration, respectively. Further, DMS-striatopallidal neurons are involved only in early motor learning whereas gradual motor skill acquisition depends on striatonigral neurons in the sensorimotor striatum. Finally, associative striatum D(2)R neurons are required for the cataleptic effect of the typical neuroleptic drug haloperidol and for amphetamine motor response sensitization. Altogether, these data provide direct experimental evidence for cell-specific topographic functional organization of the dorsal striatum.     

STD and TRNOESY NMR studies for the epitope mapping of the phosphorylation motif of the oncogenic protein beta-catenin recognized by a selective monoclonal antibody

The interaction of the P-beta-Cat(19-44) peptide, a 26 amino acid peptide (K(19)AAVSHWQQQSYLDpSGIHpSGATTTAP(44)) that mimics the phosphorylated beta-Catenin antigen, has been studied with its monoclonal antibody BC-22, by transferred nuclear Overhauser effect NMR spectroscopy (TRNOESY) and saturation transfer difference NMR (STD NMR) spectroscopy. This antibody is specific to diphosphorylated beta-Catenin and does not react with the non-phosphorylated protein. Phosphorylation of beta-Catenin at sites Ser33 and Ser37 on the DSGXXS motif is required for the interaction of beta-Catenin with the ubiquitin ligase SCF(beta-TrCP). beta-TrCP is involved in the ubiquitination and proteasome targeting of the oncogenic protein beta-Catenin, the accumulation of which has been implicated in various human cancers. The three-dimensional structure of the P-beta-Cat(19-44) in the bound conformation was determined by TRNOESY NMR experiments; the peptide adopts a compact structure in the presence of mAb with formation of turns around Trp25 and Gln26, with a tight bend created by the DpS(33)GIHpS(37) motif; the peptide residues (D32-pS37) forming this bend are recognized by the antibody as demonstrated by STD NMR experiments. 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 bidentate association. These findings are in excellent agreement with a recently published NMR structure required for the interaction of beta-Catenin with the SCF(beta-TrCP) protein.     

The role of copper in cysteine oxidation: study of intra- and inter-molecular reactions in mass spectrometry

Cysteine-containing peptide oxidation was studied both by using an inert platinum electrode and a sacrificial electrode (copper or zinc) generating metallic ions in electrospray ionization mass spectrometry (ESI-MS). Using peptides containing one, two and three cysteines, we have compared the different chemical and electrochemical oxidation pathways of cysteine (RS(-II)H) to cystine (RS(-I)S(-I)R) and to sulfenic, sulfinic and sulfonic acid (RS(0)OH, RS(II)O(2)H and RS(IV)O(3)H, respectively). In the absence of copper ions, intra-molecular reactions were the most abundant, whereas inter-molecular reactions were found to be enhanced by the presence of copper ions. These cations favor the formation of 2 : 1 (peptide : copper) complexes compared to 1 : 1 complexes, thus enhancing the formation of inter-molecular bridges. This study highlights the importance of the position of cysteine inside a peptide during disulfide bridge formation.     

Social and Emotional Values of Sounds Influence Human (Homo sapiens) and Non-Human Primate (Cercopithecus campbelli) Auditory Laterality

The last decades evidenced auditory laterality in vertebrates, offering new important insights for the understanding of the origin of human language. Factors such as the social (e.g. specificity, familiarity) and emotional value of sounds have been proved to influence hemispheric specialization. However, little is known about the crossed effect of these two factors in animals. In addition, human-animal comparative studies, using the same methodology, are rare. In our study, we adapted the head turn paradigm, a widely used non invasive method, on 8–9-year-old schoolgirls and on adult female Campbell''s monkeys, by focusing on head and/or eye orientations in response to sound playbacks. We broadcast communicative signals (monkeys: calls, humans: speech) emitted by familiar individuals presenting distinct degrees of social value (female monkeys: conspecific group members vs heterospecific neighbours, human girls: from the same vs different classroom) and emotional value (monkeys: contact vs threat calls; humans: friendly vs aggressive intonation). We evidenced a crossed-categorical effect of social and emotional values in both species since only “negative” voices from same class/group members elicited a significant auditory laterality (Wilcoxon tests: monkeys, T = 0 p = 0.03; girls: T = 4.5 p = 0.03). Moreover, we found differences between species as a left and right hemisphere preference was found respectively in humans and monkeys. Furthermore while monkeys almost exclusively responded by turning their head, girls sometimes also just moved their eyes. This study supports theories defending differential roles played by the two hemispheres in primates'' auditory laterality and evidenced that more systematic species comparisons are needed before raising evolutionary scenario. Moreover, the choice of sound stimuli and behavioural measures in such studies should be the focus of careful attention.     

Probing cysteine reactivity in proteins by mass spectrometric EC-tagging

The on-line electrochemical tagging (EC-tagging) of cysteine residues in proteins during mass spectrometry is studied to probe the cysteine environment. Benzoquinone probes electrogenerated at a microspray electrode react with the thiol functions of the proteins within a microchannel and the products are analyzed by mass spectrometry. The fundamentals of the technique are discussed, with a focus on the kinetic aspects. The EC-tagging efficiency of the cysteine residues in proteins is used to probe their environment. Experiments with unmodified proteins and their chemically reduced forms highlight the strong effect of the cysteine site reactivity on the tagging efficiencies. This study highlights relevant parameters for such on-line electrochemical derivatization/MS detection strategies.