USF binding sequences from the HS4 insulator element impose early replication timing on a vertebrate replicator

The nuclear genomes of vertebrates show a highly organized program of DNA replication where GC-rich isochores are replicated early in S-phase, while AT-rich isochores are late replicating. GC-rich regions are gene dense and are enriched for active transcription, suggesting a connection between gene regulation and replication timing. Insulator elements can organize independent domains of gene transcription and are suitable candidates for being key regulators of replication timing. We have tested the impact of inserting a strong replication origin flanked by the β-globin HS4 insulator on the replication timing of naturally late replicating regions in two different avian cell types, DT40 (lymphoid) and 6C2 (erythroid). We find that the HS4 insulator has the capacity to impose a shift to earlier replication. This shift requires the presence of HS4 on both sides of the replication origin and results in an advance of replication timing of the target locus from the second half of S-phase to the first half when a transcribed gene is positioned nearby. Moreover, we find that the USF transcription factor binding site is the key cis-element inside the HS4 insulator that controls replication timing. Taken together, our data identify a combination of cis-elements that might constitute the basic unit of multi-replicon megabase-sized early domains of DNA replication

Dynamique et synchronisme de réplication de l'ADN dans des cellules vivantes -- Analyse de marqueurs fluorescents

International audienceThis communication deals with the analysis of cellular biology data, for the analysis of replication timing and synchonization between allels. Data are obtained via cytometric imaging. The analysis include the development of a statistical model, the estimation of the parameter of a mixture of distributions. Results are validated through statistical tests and quantified using a bootstrap technique. From the biological point of view, new mechanisms involved in replication are exhibited.Cette communication présente l'analyse de données de biologie cellulaire en vue de l'étude du \textit{timing} et de la synchronie de réplication des allèles. Les données disponibles sont acquises massivement par une technique d'imagerie cytométrique. L'analyse fait notamment intervenir la définition d'un modèle statistique et l'identification des paramètres d'un mélange de distributions. Les résultats sont validés par des tests statistiques et quantifiés par bootstrap. Du point de vue biologique, des nouveaux mécanismes intervenant dans la réplication sont exhibés

Apn1 AP-endonuclease is essential for the repair of oxidatively damaged DNA bases in yeast frataxin-deficient cells

International audienceFrataxin deficiency results in mitochondrial dysfunction and oxidative stress and it is the cause of the hereditary neurodegenerative disease Friedreich ataxia (FA). Here, we present evidence that one of the pleiotrop-ic effects of oxidative stress in frataxin-deficient yeast cells (Dyfh1 mutant) is damage to nuclear DNA and that repair requires the Apn1 AP-endonuclease of the base excision repair pathway. Major phenotypes of Dyfh1 cells are respiratory deficit, disturbed iron homeostasis and sensitivity to oxidants. These phenotypes are weak or absent under anaerobiosis. We show here that exposure of anaerobically grown Dyfh1 cells to oxygen leads to down-regulation of antioxidant defenses, increase in reactive oxygen species, delay in G1-and S-phases of the cell cycle and damage to mitochondrial and nuclear DNA. Nuclear DNA lesions in Dyfh1 cells are primarily caused by oxidized bases and single-strand breaks that can be detected 15-30 min after oxygen exposition. The Apn1 enzyme is essential for the repair of the DNA lesions in Dyfh1 cells. Compared with Dyfh1, the double Dyfh1Dapn1 mutant shows growth impairment, increased mu-tagenesis and extreme sensitivity to H 2 O 2. On the contrary, overexpression of the APN1 gene in Dyfh1 cells decreases spontaneous and induced mutagenesis. Our results show that frataxin deficiency in yeast cells leads to increased DNA base oxidation and requirement of Apn1 for repair, suggesting that DNA damage and repair could be important features in FA disease progression

Inter-species variation in the oligomeric states of the higher plant Calvin cycle enzymes glyceraldehyde-3-phosphate dehydrogenase and phosphoribulokinase

In darkened leaves the Calvin cycle enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK) form a regulatory multi-enzyme complex with the small chloroplast protein CP12. GAPDH also forms a high molecular weight regulatory mono-enzyme complex. Given that there are different reports as to the number and subunit composition of these complexes and that enzyme regulatory mechanisms are known to vary between species, it was reasoned that protein-protein interactions may also vary between species. Here, this variation is investigated. This study shows that two different tetramers of GAPDH (an A2B2 heterotetramer and an A4 homotetramer) have the capacity to form part of the PRK/GAPDH/CP12 complex. The role of the PRK/GAPDH/CP12 complex is not simply to regulate the 'non-regulatory' A4 GAPDH tetramer. This study also demonstrates that the abundance and nature of PRK/GAPDH/CP12 interactions are not equal in all species and that whilst NAD enhances complex formation in some species, this is not sufficient for complex formation in others. Furthermore, it is shown that the GAPDH mono-enzyme complex is more abundant as a 2(A2B2) complex, rather than the larger 4(A2B2) complex. This smaller complex is sensitive to cellular metabolites indicating that it is an important regulatory isoform of GAPDH. This comparative study has highlighted considerable heterogeneity in PRK and GAPDH protein interactions between closely related species and the possible underlying physiological basis for this is discussed. © 2011 The Author(s)

Polycyclic aromatic hydrocarbon components contribute to the mitochondria-antiapoptotic effect of fine particulate matter on human bronchial epithelial cells via the aryl hydrocarbon receptor

<p>Abstract</p> <p>Background</p> <p>Nowadays, effects of fine particulate matter (PM_2.5) are well-documented and related to oxidative stress and pro-inflammatory response. Nevertheless, epidemiological studies show that PM_2.5exposure is correlated with an increase of pulmonary cancers and the remodeling of the airway epithelium involving the regulation of cell death processes. Here, we investigated the components of Parisian PM_2.5involved in either the induction or the inhibition of cell death quantified by different parameters of apoptosis and delineated the mechanism underlying this effect.</p> <p>Results</p> <p>In this study, we showed that low levels of Parisian PM_2.5are not cytotoxic for three different cell lines and primary cultures of human bronchial epithelial cells. Conversely, a 4 hour-pretreatment with PM_2.5prevent mitochondria-driven apoptosis triggered by broad spectrum inducers (A23187, staurosporine and oligomycin) by reducing the mitochondrial transmembrane potential loss, the subsequent ROS production, phosphatidylserine externalization, plasma membrane permeabilization and typical morphological outcomes (cell size decrease, massive chromatin and nuclear condensation, formation of apoptotic bodies). The use of recombinant EGF and specific inhibitor led us to rule out the involvement of the classical EGFR signaling pathway as well as the proinflammatory cytokines secretion. Experiments performed with different compounds of PM_2.5suggest that endotoxins as well as carbon black do not participate to the antiapoptotic effect of PM_2.5. Instead, the water-soluble fraction, washed particles and organic compounds such as polycyclic aromatic hydrocarbons (PAH) could mimic this antiapoptotic activity. Finally, the activation or silencing of the aryl hydrocarbon receptor (AhR) showed that it is involved into the molecular mechanism of the antiapoptotic effect of PM_2.5at the mitochondrial checkpoint of apoptosis.</p> <p>Conclusions</p> <p>The PM_2.5-antiapoptotic effect in addition to the well-documented inflammatory response might explain the maintenance of a prolonged inflammation state induced after pollution exposure and might delay repair processes of injured tissues.</p

Secuelas por Accidente Cerebrovascular Isquémico en pacientes de 40-90 años, del servicio de Medicina Interna, Hospital Roberto Calderón Gutiérrez, de enero 2011 a diciembre 2014

Las secuelas neurológicas después del accidente cerebrovascular isquémico pueden impactar negativamente en la calidad de vida de las personas y ser factor determinante en la mortalidad de estas, existiendo datos limitados y variables en cuanto a la frecuencia de su desarrollo, siendo preponderante la investigación de este tópico. El presente estudio es de tipo descriptivo, retrospectivo y de corte transversal, en el cual se abordaron las Secuelas por Accidente Cerebrovascular Isquémico en pacientes de 40 – 90 años, del servicio de Medicina Interna, del Hospital Roberto Calderón Gutiérrez, de enero 2011 a diciembre 2014, que persigue describir las secuelas por esta patología en el grupo de estudio definido. El universo se conformó por 138 expedientes de pacientes con la patología, siendo la muestra de 103 expedientes, la fuente fue secundaria, conformada por la revisión de expedientes clínico, recopilando los datos por medio de la Ficha de recolección elaborada en base a los objetivos propuestos en el estudio. Los principales resultados reflejaron que el sexo predominante fue el femenino, entre el grupo etario de 71 a 80 años. La Hipertensión Arterial representó el antecedente patológico más frecuente, siendo la arteria cerebral media la más afectada. La parálisis / paresia de las extremidades contralaterales fue la secuela predominante. Por tanto, se recomienda hacer insistencia en la atención integral en salud brindada a los usuarios, logrando reconocer factores de riesgo patológico y no patológico incidiendo así en su control o eliminación y de esta forma mitigando el desarrollo de esta enfermedad. Palabras Claves: Secuelas, accidente cerebrovascular, isquemia

Interactions between Magnetic Nanowires and Living Cells : Uptake, Toxicity and Degradation

We report on the uptake, toxicity and degradation of magnetic nanowires by NIH/3T3 mouse fibroblasts. Magnetic nanowires of diameters 200 nm and lengths comprised between 1 {\mu}m and 40 {\mu}m are fabricated by controlled assembly of iron oxide ({\gamma}-Fe2O3) nanoparticles. Using optical and electron microscopy, we show that after 24 h incubation the wires are internalized by the cells and located either in membrane-bound compartments or dispersed in the cytosol. Using fluorescence microscopy, the membrane-bound compartments were identified as late endosomal/lysosomal endosomes labeled with lysosomal associated membrane protein (Lamp1). Toxicity assays evaluating the mitochondrial activity, cell proliferation and production of reactive oxygen species show that the wires do not display acute short-term (< 100 h) toxicity towards the cells. Interestingly, the cells are able to degrade the wires and to transform them into smaller aggregates, even in short time periods (days). This degradation is likely to occur as a consequence of the internal structure of the wires, which is that of a non-covalently bound aggregate. We anticipate that this degradation should prevent long-term asbestos-like toxicity effects related to high aspect ratio morphologies and that these wires represent a promising class of nanomaterials for cell manipulation and microrheology.Comment: 21 pages 12 figure

Structural features and kinetic characterization of alanine racemase from Staphylococcus aureus (Mu50)

The tertiary structure and kinetic properties of alanine racemase from Staphylococcus aureus are described and compared to other related alanine racemase structures

A Novel Role for Dbx1-Derived Cajal-Retzius Cells in Early Regionalization of the Cerebral Cortical Neuroepithelium

Patterning of the cerebral cortex during embryogenesis depends not only on passive diffusion of morphogens but also on signal delivery by Cajal-Retzius neurons that migrate over long distances

Osseointegrated screw-shaped titanium implant as orthodontic anchorage: a clinical and histological animal study

PLEASE NOTE: This work is protected by copyright. Downloading is restricted to the BU community: please click Download and log in with a valid BU account to access. If you are the author of this work and would like to make it publicly available, please contact [email protected] (M.Sc.D.)--Boston University, Henry M. Goldman School of Graduate Dentistry, 1990 (Orthodontics).Bibliography: leaves 179-208