G-protein-coupled receptor oligomers: two or more for what? Lessons from mGlu and GABAB receptors.

G-protein-coupled receptors (GPCRs) are key players in the precise tuning of intercellullar communication. In the brain, both major neurotransmitters, glutamate and GABA, act on specific GPCRs [the metabotropic glutamate (mGlu) and GABA(B) receptors] to modulate synaptic transmission. These receptors are encoded by the largest gene family, and have been found to associate into both homo- and hetero-oligomers, which increases the complexity of this cell communication system. Here we show that dimerization is required for mGlu and GABA(B) receptors to function, since the activation process requires a relative movement between the subunits to occur. We will also show that, in contrast to the mGlu receptors, which form strict dimers, the GABA(B) receptors assemble into larger complexes, both in transfected cells and in the brain, resulting in a decreased G-protein coupling efficacy. We propose that GABA(B) receptor oligomerization offers a way to increase the possibility of modulating receptor signalling and activity, allowing the same receptor protein to have specific properties in neurons at different locations.

A lock-in-based method to examine the thermal signatures of magnetic nanoparticles in the liquid, solid and aggregated states.

We propose a new methodology based on lock-in thermography to study and quantify the heating power of magnetic nanoparticles. Superparamagnetic iron oxide nanoparticles exposed to a modulated alternating magnetic field were used as model materials to demonstrate the potency of the system. Both quantitative and qualitative information on their respective heating power was extracted at high thermal resolutions under increasingly complex conditions, including nanoparticles in the liquid, solid and aggregated states. Compared to conventional techniques, this approach offers a fast, sensitive and non-intrusive alternative to investigate multiple and dilute specimens simultaneously, which is essential for optimizing and accelerating screening procedures and comparative studies.

Characterizing nanoparticles in complex biological media and physiological fluids with depolarized dynamic light scattering.

Light scattering is one of the few techniques available to adequately characterize suspended nanoparticles (NPs) in real time and in situ. However, when it comes to NPs in multicomponent and optically complex aqueous matrices - such as biological media and physiological fluids - light scattering suffers from lack of selectivity, as distinguishing the relevant optical signals from the irrelevant ones is very challenging. We meet this challenge by building on depolarized scattering: Unwanted signals from the matrix are completely suppressed. This approach yields information with an unprecedented signal-to-noise ratio in favour of the NPs and NP-biomolecule corona complexes, which in turn opens the frontier to scattering-based studies addressing the behaviour of NPs in complex physiological/biological fluids.

Hypothesis: hyperstructures regulate bacterial structure and the cell cycle.

A myriad different constituents or elements (genes, proteins, lipids, ions, small molecules etc.) participate in numerous physico-chemical processes to create bacteria that can adapt to their environments to survive, grow and, via the cell cycle, reproduce. We explore the possibility that it is too difficult to explain cell cycle progression in terms of these elements and that an intermediate level of explanation is needed. This level is that of hyperstructures. A hyperstructure is large, has usually one particular function, and contains many elements. Non-equilibrium, or even dissipative, hyperstructures that, for example, assemble to transport and metabolize nutrients may comprise membrane domains of transporters plus cytoplasmic metabolons plus the genes that encode the hyperstructure's enzymes. The processes involved in the putative formation of hyperstructures include: metabolite-induced changes to protein affinities that result in metabolon formation, lipid-organizing forces that result in lateral and transverse asymmetries, post-translational modifications, equilibration of water structures that may alter distributions of other molecules, transertion, ion currents, emission of electromagnetic radiation and long range mechanical vibrations. Equilibrium hyperstructures may also exist such as topological arrays of DNA in the form of cholesteric liquid crystals. We present here the beginning of a picture of the bacterial cell in which hyperstructures form to maximize efficiency and in which the properties of hyperstructures drive the cell cycle.

A novel gene from Myxococcus xanthus that facilitates membrane translocation of an extracellular endoglucanase in Escherichia coli?

Expression in Escherichia coli of the Myxococcus xanthus gene celA, which encodes an extracellular endoglucanase, resulted in CelA being distributed between cytoplasm, periplasm and membrane. The presence of an adjacent open reading frame downstream from the full celA gene, or the absence of a putative lipoprotein signal sequence, confined CelA distribution to the periplasm and membrane, or to the cytoplasm and periplasm, respectively.

Comparison of beta-galactosidase production by two inducible promoters in Myxococcus xanthus.

The inducibility of two promoter systems, one heterologous and one homologous, has been assessed in the Gram-negative bacterium Myxococcus xanthus. The heterologous system involved the hybrid tac promoter and the presence of lacIq, the lac repressor from Escherichia coli. This system is inducible in its natural host with isopropyl-beta-D-thiogalactopyranoside (IPTG). The homologous promoter system involves the light-inducible carQRS promoter, which is normally involved in the expression of the regulators of the light-inducible light-protective carotenoid synthesis regulon in M. xanthus. In each case, promoter activity and strength was assayed using the E. coli gene lacZ. In our constructs, which were present in a single copy in the M. xanthus chromosome, the carQRS promoter yielded at least a 47-fold increase in beta-galactosidase production upon light induction, whilst IPTG increased by 8-fold the amount of enzyme produced under the control of the ptac-lacIq system. Regulation by the latter was significantly higher than that obtained with the unmodified lacZ promoter.

Elevated (+)-maze and hole-board exploration in lurcher mutant mice.

The behavior of lurcher mice, a mutant with degeneration of cerebellar cells, was compared to that of normal mice for three days in two tests of exploration; an elevated (+)-maze and a 4 x 4 hole-board. In the elevated (+)-maze, lurcher mutants visited fewer closed arms than normal mice only on the first test day. Lurcher mutants were slower to emerge from the first closed arm but did not differ from normal mice for entry latencies into the first open arm. The time spent by the mutants in the open arms was higher than that of normal mice, an indication of decreased inhibition to open spaces. In the hole-board, lurcher mutants visited fewer holes than normal mice only on the first day of testing. In proportion to the total number of holes explored, lurcher mutants visited fewer center holes and fewer holes situated next to each other. These results may be due to a lesion-induced tendency to explore a more restricted region of a novel spatial environment and to explore it in a more haphazard fashion.

Spatial learning in a Z-maze by cerebellar mutant mice.

Two types of cerebellar mutant mice (staggerer and lurcher) were evaluated during 5-day acquisition of a spatial learning task in a Z-maze filled with water. Although the number of errors and escape latencies decreased in normal mice, the acquisition of the cerebellar mutants was impaired but not abolished. These results indicate that the cerebellum has a role in spatial learning. Mice with cerebellar dysfunction take a more indirect route toward a goal during the course of swimming, when ataxic symptoms are no longer in evidence.

In vitro dosimetry of agglomerates.

Agglomeration of nanoparticles in biological fluids is a pervasive phenomenon that leads to difficulty in the interpretation of results from in vitro exposure, primarily due to differing particokinetics of agglomerates to nanoparticles. Therefore, well-defined small agglomerates were designed that possessed different particokinetic profiles, and their cellular uptake was compared to a computational model of dosimetry. The approach used here paves the way for a better understanding of the impact of agglomeration on the nanoparticle-cell interaction.

Quality of care of patients with type 1 diabetes: population-based results in a French region.

AIM: Although the incidence of type 1 diabetes (T1D) has been increasing, little is known of its quality of care. Thus, our survey was designed to retrospectively evaluate this issue in French patients. METHODS: Patients with T1D living in northeastern France were identified thanks to the healthcare system (CPAM) database, and the resulting list reviewed by local diabetes specialists. All of the listed patients and their primary physicians were asked to fill in a questionnaire including clinical data, laboratory results and follow-up habits. The 'optimized results' included CPAM-based results plus any specialized care provided during hospitalizations in diabetes and non-diabetes units, according to questionnaire data. RESULTS: A total of 227 individuals, for whom CPAM data were available, were identified as having T1D. From these patients, 174 questionnaires were answered, and optimized results (having both CPAM data and a completely filled-in questionnaire) were available for 149 patients. Of the 169 patients who responded, 71.3% reported at least a yearly visit with a diabetologist. This number reached 77.9% when optimized results were considered. Patients who received specialized care were younger, underwent HbA(1c) tests more often and were more frequently on optimal treatment; however, there was no difference in HbA(1c) values or in the prevalence of complications. Eye examinations and kidney tests had been performed at least once over the 2-year period in more than 87% of the patients, whereas around 30%, 21% and 23% had an eye exam, creatinine test and urinary albumin excretion measurement, respectively, only once over the same time period. CONCLUSION: This is the first large-scale study of the quality of care in patients with T1DM in France, and it could serve as a preliminary survey for a national study. Although the follow-up was better than previously reported, there is still considerable room for improvement.

Identification of hematite particles in sealed glass containers for pharmaceutical uses by Raman microspectroscopy.

Raman microspectroscopy has been shown to enable the identification of micro-particles inside sealed glass containers for pharmaceutical use without any sample preparation. Raman spectra were collected from unknown particles with a maximum size of 1mm, adsorbed on the inner surface of ampoules. The particles were clearly identified as primarily hematite with traces of magnetite by their characteristic Raman spectral bands. The presence of this deposit was attributed to the projection of iron oxides during the manufacturing process. These oxide particles were not detected by the quality control process of the glass manufacturer, showing that in-process quality controls failed to detect this problem. Particle identification by Raman microspectroscopy appears to be a selective, rapid and reliable analytical procedure for quality control and assurance in the pharmaceutical industry. Identification of the particles was also helpful for evaluating the nature of the contaminant and enables consequences for the toxicological aspects of final product quality to be managed.

Enzyme activities associated with an invertebrate iridovirus: protein kinase activity associated with iridescent virus type 6 (chilo iridescent virus).

Iridescent virus type 6 was found to contain an endogenous protein kinase activity which can phosphorylate some viral proteins and exogenous basic proteins. The enzyme required a divalent metal ion but was not stimulated by cyclic nucleotides. Procedures which are known to solubilize the viral envelope indicated that the protein kinase was an internal component of the virion. Conditions for protein kinase activity are described.

Enzyme activities associated with an invertebrate iridovirus: nucleotide phosphohydrolase activity associated with iridescent virus type 6 (CIV).

A nucleoside triphosphate phosphohydrolase activity is firmly associated with a purified invertebrate iridovirus, iridescent virus type 6. The enzyme activity hydrolyzes all the nucleoside triphosphates, but has a high preference for ATP. The products of the reaction are nucleoside diphosphates. Conditions for nucleoside triphosphate phosphohydrolase activity are described.

Mutations in two new loci that impair both extracellular protein production and development in Myxococcus xanthus.

Two transposon insertion mutants of Myxococcus xanthus altered in the secretion of protein as determined by the hydrolytic activities of several enzymes during vegetative growth were also unable to complete fruiting body formation and were severely impaired in sporulation. The insertions were located in the same part of the M. xanthus chromosome but were unlinked by transduction and therefore define two distinct loci, called excA and excB. Since both Exc +/- mutants were able to rescue development of an asgB mutation, they do not belong to the Asg- group, despite of the fact that asg mutants are also Exc +/-. Our results sustain the hypothesis of a possible relationship between protein secretion during vegetative growth and development or sporulation.

[Effect of the extrusion process on the availability of proteins]. / Influence du procédé de cuisson extrusion sur la disponibilité des protéines.

The extrusion process has grown rapidly as in the preparation of meat-extenders from vegetal proteins as in the fabrication of cocktail-snacks and food for breakfast. The nutritional modifications induced by the thermic shock during the extrusion affect: -the diminution of the content of nutrient: loss in vitamins, destruction and unavailability of amino-acids; -the structural modification of a few compounds: starch gelatinization; -destruction of antinutritional factors: essentially antitrypsic factor. We studied the effect of extrusion with a model-mixture on the availability of amino-acids and particularly lysin. We effected the systematic study of different factors which can interfer in the intensity of the Maillard reaction in relation to the composition of the mixture (presence and nature of sugar, etc.) and to the technological parameters (screw-section, moisture of the mixture, etc.) The model mixture composed of cereal flours shows a global loss of 32 p. 100 in lysin (destruction 18 p. 100, biological unavailability 16 p. 100) after extrusion. When this mixture is added up with 7,2 p. 100 of saccharose the total loss in lysin reaches 40 p. 100. If saccharose is substituted by a more reducing sugar (fructose) the lysin loss approaches 80 p. 100. The addition of 4 p. 100 water at the mixture containing saccharose (total moisture 14 p. 100) limits the lysin loss to 10 p. 100. The modification of other technological parameters does not improve significatively the level of available lysin. The moisture of the mixture before extrusion appears to be a preponderatting factor in the intensity of the Maillard reaction during extrusion.

Hypothesis: the meeting place model for prion disease.

Prions are responsible for spongiform diseases such as scrapie and bovine spongiform encephalopathy. It is now generally accepted that the disease mechanism involves the conversion from the normal form, PrPC, to the pathogenic form, PrPSc, and that this isoform is infectious. In the case of scrapie, 15 different forms of the disease have been described and some of these different phenotypes can be conferred by infectious prions that are themselves encoded by normal genes. We propose here that a prion with an altered structure has a correspondingly altered preference for lipids; this altered preference creates a proteolipid domain containing different lipids and other factors such as chaperonins and enzymes responsible for post-translational modifications. Normal prions associated with this abnormal domain adopt the conformation dictated by its lipidic composition (and by the other factors present) and so acquire the lipidic preference of the original pathogenic prions. These transformed prions could then create new proteolipid domains. This process may be considered as semi-conservative replication in which prion and lipids are analogous to the Watson and Crick strands and the proteolipid domain to the double helix itself.