The complete genome sequence of the gram-positive bacterium Bacillus subtilis.

Bacillus subtilis is the best-characterized member of the Gram-positive bacteria. Its genome of 4,214,810 base pairs comprises 4,100 protein-coding genes. Of these protein-coding genes, 53% are represented once, while a quarter of the genome corresponds to several gene families that have been greatly expanded by gene duplication, the largest family containing 77 putative ATP-binding transport proteins. In addition, a large proportion of the genetic capacity is devoted to the utilization of a variety of carbon sources, including many plant-derived molecules. The identification of five signal peptidase genes, as well as several genes for components of the secretion apparatus, is important given the capacity of Bacillus strains to secrete large amounts of industrially important enzymes. Many of the genes are involved in the synthesis of secondary metabolites, including antibiotics, that are more typically associated with Streptomyces species. The genome contains at least ten prophages or remnants of prophages, indicating that bacteriophage infection has played an important evolutionary role in horizontal gene transfer, in particular in the propagation of bacterial pathogenesis.

Étude des mécanismes de transfert des fibres en sciences forensiques

RESUME Les fibres textiles sont des produits de masse utilisés dans la fabrication de nombreux objets de notre quotidien. Le transfert de fibres lors d'une action délictueuse est dès lors extrêmement courant. Du fait de leur omniprésence dans notre environnement, il est capital que l'expert forensique évalue la valeur de l'indice fibres. L'interprétation de l'indice fibres passe par la connaissance d'un certain nombre de paramètres, comme la rareté des fibres, la probabilité de leur présence par hasard sur un certain support, ainsi que les mécanismes de transfert et de persistance des fibres. Les lacunes les plus importantes concernent les mécanismes de transfert des fibres. A ce jour, les nombreux auteurs qui se sont penchés sur le transfert de fibres ne sont pas parvenus à créer un modèle permettant de prédire le nombre de fibres que l'on s'attend à retrouver dans des circonstances de contact données, en fonction des différents paramètres caractérisant ce contact et les textiles mis en jeu. Le but principal de cette recherche est de démontrer que la création d'un modèle prédictif du nombre de fibres transférées lors d'un contact donné est possible. Dans le cadre de ce travail, le cas particulier du transfert de fibres d'un tricot en laine ou en acrylique d'un conducteur vers le dossier du siège de son véhicule a été étudié. Plusieurs caractéristiques des textiles mis en jeu lors de ces expériences ont été mesurées. Des outils statistiques (régression linéaire multiple) ont ensuite été utilisés sur ces données afin d'évaluer l'influence des caractéristiques des textiles donneurs sur le nombre de fibres transférées et d'élaborer un modèle permettant de prédire le nombre de fibres qui vont être transférées à l'aide des caractéristiques influençant significativement le transfert. Afin de faciliter la recherche et le comptage des fibres transférées lors des expériences de transfert, un appareil de recherche automatique des fibres (liber finder) a été utilisé dans le cadre de cette recherche. Les tests d'évaluation de l'efficacité de cet appareil pour la recherche de fibres montrent que la recherche automatique est globalement aussi efficace qu'une recherche visuelle pour les fibres fortement colorées. Par contre la recherche automatique perd de son efficacité pour les fibres très pâles ou très foncées. Une des caractéristiques des textiles donneurs à étudier est la longueur des fibres. Afin de pouvoir évaluer ce paramètre, une séquence d'algorithmes de traitement d'image a été implémentée. Cet outil permet la mesure de la longueur d'une fibre à partir de son image numérique à haute résolution (2'540 dpi). Les tests effectués montrent que les mesures ainsi obtenues présentent une erreur de l'ordre du dixième de millimètre, ce qui est largement suffisant pour son utilisation dans le cadre de cette recherche. Les résultats obtenus suite au traitement statistique des résultats des expériences de transfert ont permis d'aboutir à une modélisation du phénomène du transfert. Deux paramètres sont retenus dans le modèle: l'état de la surface du tissu donneur et la longueur des fibres composant le tissu donneur. L'état de la surface du tissu est un paramètre tenant compte de la quantité de fibres qui se sont détachées de la structure du tissu ou qui sont encore faiblement rattachées à celle-ci. En effet, ces fibres sont les premières à se transférer lors d'un contact, et plus la quantité de ces fibres par unité de surface est importante, plus le nombre de fibres transférées sera élevé. La longueur des fibres du tissu donneur est également un paramètre important : plus les fibres sont longues, mieux elles sont retenues dans la structure du tissu et moins elles se transféreront. SUMMARY Fibres are mass products used to produce numerous objects encountered everyday. The transfer of fibres during a criminal action is then very common. Because fibres are omnipresent in our environment, the forensic expert has to evaluate the value of the fibre evidence. To interpret fibre evidence, the expert has to know some parameters as frequency of fibres,' probability of finding extraneous fibres by chance on a given support, and transfer and persistence mechanisms. Fibre transfer is one of the most complex parameter. Many authors studied fibre transfer mechanisms but no model has been created to predict the number of fibres transferred expected in a given type of contact according to parameters as characteristics of the contact and characteristics of textiles. The main purpose of this research is to demonstrate that it is possible to create a model to predict the number of fibres transferred during a contact. In this work, the particular case of the transfer of fibres from a knitted textile in wool or in acrylic of a driver to the back of a carseat has been studied. Several characteristics of the textiles used for the experiments were measured. The data obtained were then treated with statistical tools (multiple linear regression) to evaluate the influence of the donor textile characteristics on the number of úbers transferred, and to create a model to predict this number of fibres transferred by an equation containing the characteristics having a significant influence on the transfer. To make easier the searching and the counting of fibres, an apparatus of automatic search. of fibers (fiber finder) was used. The tests realised to evaluate the efficiency of the fiber finder shows that the results obtained are generally as efficient as for visual search for well-coloured fibres. However, the efficiency of automatic search decreases for pales and dark fibres. One characteristic of the donor textile studied was the length of the fibres. To measure this parameter, a sequence of image processing algorithms was implemented. This tool allows to measure the length of a fibre from it high-resolution (2'540 dpi) numerical image. The tests done shows that the error of the measures obtained are about some tenths of millimetres. This precision is sufficient for this research. The statistical methods applied on the transfer experiment data allow to create a model of the transfer phenomenon. Two parameters are included in the model: the shedding capacity of the donor textile surface and the length of donor textile fibres. The shedding capacity of the donor textile surface is a parameter estimating the quantity of fibres that are not or slightly attached to the structure of the textile. These fibres are easily transferred during a contact, and the more this quantity of fibres is high, the more the number of fibres transferred during the contact is important. The length of fibres is also an important parameter: the more the fibres are long, the more they are attached in the structure of the textile and the less they are transferred during the contact.

Intracellular targeting of GLUT4 in transfected insulinoma cells: evidence for association with constitutively recycling vesicles distinct from synaptophysin and insulin vesicles.

In adipocytes and muscle cells, the GLUT4 glucose transporter isoform is present in intracellular vesicles which continuously recycle between an intracytoplasmic location and the plasma membrane. It is not clear whether the GLUT4-vesicles represent a specific kind of vesicle or resemble typical secretory granules or synaptic-like microvesicles. To approach this question, we expressed GLUT4 in the beta cell line RINm5F and determined its intracellular localization by subcellular fractionation and by immunofluorescence and immunoelectron microscopy. GLUT4 was not found in insulin granules but was associated with a subpopulation of smooth-surface vesicles present in the trans-Golgi region and in vesicular structures adjacent to the plasma membrane. In the trans-Golgi region, GLUT4 did not colocalize with synaptophysin or TGN38. Incubation of the cells with horseradish peroxidase (HRP) led to colocalization of HRP and GLUT4 in some endosomal structures adjacent to the plasma membrane and in occasional trans-Golgi region vesicles. When cells were incubated in the presence of Bafilomycin A, analysis by confocal microscopy revealed GLUT4 in numerous large spots present throughout the cytoplasm, many of which costained for TGN38 and synaptophysin. By immunoelectron microscopy, numerous endosomes were observed which stained strongly for GLUT4. Together our data demonstrate that ectopic expression of GLUT4 in insulinoma cells reveals the presence of a subset of vesicular structures distinct from synaptic-like vesicles and insulin secretory granules. Furthermore, they indicate that GLUT4 constitutively recycles between the plasma membrane and its intracellular location by an endocytic route also taken by TGN38 and synaptophysin.

BAFF binds to the tumor necrosis factor receptor-like molecule B cell maturation antigen and is important for maintaining the peripheral B cell population.

The tumor necrosis factor (TNF) family member B cell activating factor (BAFF) binds B cells and enhances B cell receptor-triggered proliferation. We find that B cell maturation antigen (BCMA), a predicted member of the TNF receptor family expressed primarily in mature B cells, is a receptor for BAFF. Although BCMA was previously localized to the Golgi apparatus, BCMA was found to be expressed on the surface of transfected cells and tonsillar B cells. A soluble form of BCMA, which inhibited the binding of BAFF to a B cell line, induced a dramatic decrease in the number of peripheral B cells when administered in vivo. Moreover, culturing splenic cells in the presence of BAFF increased survival of a percentage of the B cells. These results are consistent with a role for BAFF in maintaining homeostasis of the B cell population.

The systemic control of circadian gene expression.

The mammalian circadian timing system consists of a central pacemaker in the brain's suprachiasmatic nucleus (SCN) and subsidiary oscillators in nearly all body cells. The SCN clock, which is adjusted to geophysical time by the photoperiod, synchronizes peripheral clocks through a wide variety of systemic cues. The latter include signals depending on feeding cycles, glucocorticoid hormones, rhythmic blood-borne signals eliciting daily changes in actin dynamics and serum response factor (SRF) activity, and sensors of body temperature rhythms, such as heat shock transcription factors and the cold-inducible RNA-binding protein CIRP. To study these systemic signalling pathways, we designed and engineered a novel, highly photosensitive apparatus, dubbed RT-Biolumicorder. This device enables us to record circadian luciferase reporter gene expression in the liver and other organs of freely moving mice over months in real time. Owing to the multitude of systemic signalling pathway involved in the phase resetting of peripheral clocks the disruption of any particular one has only minor effects on the steady state phase of circadian gene expression in organs such as the liver. Nonetheless, the implication of specific pathways in the synchronization of clock gene expression can readily be assessed by monitoring the phase-shifting kinetics using the RT-Biolumicorder.

Interactome analysis reveals that FAM161A, deficient in recessive retinitis pigmentosa, is a component of the Golgi-centrosomal network.

Defects in FAM161A, a protein of unknown function localized at the cilium of retinal photoreceptor cells, cause retinitis pigmentosa, a form of hereditary blindness. By using different fragments of this protein as baits to screen cDNA libraries of human and bovine retinas, we defined a yeast two-hybrid-based FAM161A interactome, identifying 53 bona fide partners. In addition to statistically significant enrichment in ciliary proteins, as expected, this interactome revealed a substantial bias towards proteins from the Golgi apparatus, the centrosome and the microtubule network. Validation of interaction with key partners by co-immunoprecipitation and proximity ligation assay confirmed that FAM161A is a member of the recently recognized Golgi-centrosomal interactome, a network of proteins interconnecting Golgi maintenance, intracellular transport and centrosome organization. Notable FAM161A interactors included AKAP9, FIP3, GOLGA3, KIFC3, KLC2, PDE4DIP, NIN and TRIP11. Furthermore, analysis of FAM161A localization during the cell cycle revealed that this protein followed the centrosome during all stages of mitosis, likely reflecting a specific compartmentalization related to its role at the ciliary basal body during the G0 phase. Altogether, these findings suggest that FAM161A's activities are probably not limited to ciliary tasks but also extend to more general cellular functions, highlighting possible novel mechanisms for the molecular pathology of retinal disease.

Imaging the time-integrated cerebral metabolic activity with subcellular resolution through nanometer-scale detection of biosynthetic products deriving from (13)C-glucose.

Glucose is the primary source of energy for the brain but also an important source of building blocks for proteins, lipids, and nucleic acids. Little is known about the use of glucose for biosynthesis in tissues at the cellular level. We demonstrate that local cerebral metabolic activity can be mapped in mouse brain tissue by quantitatively imaging the biosynthetic products deriving from [U-(13)C]glucose metabolism using a combination of in situ electron microscopy and secondary ion mass-spectroscopy (NanoSIMS). Images of the (13)C-label incorporated into cerebral ultrastructure with ca. 100nm resolution allowed us to determine the timescale on which the metabolic products of glucose are incorporated into different cells, their sub-compartments and organelles. These were mapped in astrocytes and neurons in the different layers of the motor cortex. We see evidence for high metabolic activity in neurons via the nucleus (13)C enrichment. We observe that in all the major cell compartments, such as e.g. nucleus and Golgi apparatus, neurons incorporate substantially higher concentrations of (13)C-label than astrocytes.

Surgical treatment for heart myxomas.

According to unselected autopsy data, primary cardiac tumours are a rare entity. About 80% of the tumours are benign and nearly half of these are myxomas. In clinical practice, when diagnosis of this pathological entity is ascertained, decision for surgical treatment is made in order to prevent thromboembolism and obstruction of the valvular apparatus. Surgical resection including total tumour removal is accompanied by low perioperative mortality. The recidive rate is low in sporadic cases. However, in familial syndrome groups, such as the Swiss-Carney syndrome, the recurrence rate is higher.

La prima redazione della «Orazione scritta a Carlo V» di Giovanni della Casa

Nel contributo è pubblicata la prima redazione conosciuta dell'Orazione con cui Giovanni della Casa nel 1549 chiese all'imperatore Carlo V la restituzione di Piacenza ai Farnese, loro sottratta dopo l'uccisione di Pier Luigi nel 1547. L'edizione documenta l'elaborazione d'autore con un apparato evolutivo del testo trasmesso dal manoscritto Vaticano Chigiano O vi 80. This article provides an edition of the first known version of Giovanni della Casa's request to the Emperor Charles V (1549) for the restitution of Piacenza to the Farnese family after Pier Luigi's murder in 1547. This edition documents the Author's working out with an evolutional apparatus of the text transmitted by the manuscript Chigiano O vi 80.

Dominant-Negative Effects of Adult-Onset Huntingtin Mutations Alter the Division of Human Embryonic Stem Cells-Derived Neural Cells.

Mutations of the huntingtin protein (HTT) gene underlie both adult-onset and juvenile forms of Huntington's disease (HD). HTT modulates mitotic spindle orientation and cell fate in mouse cortical progenitors from the ventricular zone. Using human embryonic stem cells (hESC) characterized as carrying mutations associated with adult-onset disease during pre-implantation genetic diagnosis, we investigated the influence of human HTT and of an adult-onset HD mutation on mitotic spindle orientation in human neural stem cells (NSCs) derived from hESCs. The RNAi-mediated silencing of both HTT alleles in neural stem cells derived from hESCs disrupted spindle orientation and led to the mislocalization of dynein, the p150Glued subunit of dynactin and the large nuclear mitotic apparatus (NuMA) protein. We also investigated the effect of the adult-onset HD mutation on the role of HTT during spindle orientation in NSCs derived from HD-hESCs. By combining SNP-targeting allele-specific silencing and gain-of-function approaches, we showed that a 46-glutamine expansion in human HTT was sufficient for a dominant-negative effect on spindle orientation and changes in the distribution within the spindle pole and the cell cortex of dynein, p150Glued and NuMA in neural cells. Thus, neural derivatives of disease-specific human pluripotent stem cells constitute a relevant biological resource for exploring the impact of adult-onset HD mutations of the HTT gene on the division of neural progenitors, with potential applications in HD drug discovery targeting HTT-dynein-p150Glued complex interactions.