Quantification of lentiviral vector copy numbers in individual hematopoietic colony-forming cells shows vector dose-dependent effects on the frequency and level of transduction

Lentiviral vectors are effective tools for gene transfer and integrate variable numbers of proviral DNA copies in variable proportions of cells. The levels of transduction of a cellular population may therefore depend upon experimental parameters affecting the frequency and/or the distribution of vector integration events in this population. Such analysis would require measuring vector copy numbers (VCN) in individual cells. To evaluate the transduction of hematopoietic progenitor cells at the single-cell level, we measured VCN in individual colony-forming cell (CFC) units, using an adapted quantitative PCR (Q-PCR) method. The feasibility, reproducibility and sensitivity of this approach were tested with characterized cell lines carrying known numbers of vector integration. The method was validated by correlating data in CFC with gene expression or with calculated values, and was found to slightly underestimate VCN. In spite of this, such Q-PCR on CFC was useful to compare transduction levels with different infection protocols and different vectors. Increasing the vector concentration and re-iterating the infection were two different strategies that improved transduction by increasing the frequency of transduced progenitor cells. Repeated infection also augmented the number of integrated copies and the magnitude of this effect seemed to depend on the vector preparation. Thus, the distribution of VCN in hematopoietic colonies may depend upon experimental conditions including features of vectors. This should be carefully evaluated in the context of ex vivo hematopoietic gene therapy studies

Deformation and Exhumation of the sub-Continental Mantle: Insigth from the Ronda Peridotite (Spain)

1 p.International audienc

Crop mixtures outperform rotations and landscape mosaics in regulation of two fungal wheat pathogens: a simulation study

International audienceContext Crop rotations, within-field mixtures, and landscape mosaics including susceptible and resistant crops are three commonly adopted crop diversification strategies that can limit crop epidemics. Typically, the effects of crop diversification at these three scales have been studied separately, on single pathogen species, and with low environmental variability. Objectives We aim to compare the disease-limitation effect of these three types of crop diversification on two highly damaging fungal pathogens of wheat Puccinia recondita (WLR) and Zymoseptoria tritici (STB) and under varying weather conditions (warmer or cooler climate for WLR, wetter or drier conditions for STB). Methods We built a dynamic mathematical model of epidemics at the field scale (based on classical Susceptible-Exposed-Infectious-Removed epidemiological models) embedded in a spatially explicit landscape grid framework. We use it to simulate an agricultural landscape in which diversification translates into different proportions of wheat and resistant crops in the landscape. Results In our simulations, for both pathogens and in all weather conditions, within-field crop mixtures had the greatest impact in limiting epidemics, crop rotations were second-best, while landscape mosaics were the least effective. We also found that the threshold above which further addition of resistant plants to crop mixtures would not cause further disease limitation to be dependent on weather conditions. The more favorable the weather is for pathogens the more resistant plants are required. Conclusions Our findings imply that interactions between spatial scale of crop diversification, pathogen characteristics and weather conditions should be considered in order to maximize benefits from disease-regulation properties of diversified cropping systems under climate change

Water pumping in mantle shear zones: From field observations to experimental evidence

International audienceWater plays an important role in geological processes. Providing constraints on what may influence the distribution of aqueous fluids is thus crucial to understanding how H2O impacts Earth's geodynamics. In a deep-seated environment, viscous shear zones have been identified as sites of massive fluid circulation, with many implications for ores deposits and rock rheology. However, although seismic pumping, fluid permeation and/or creep cavitation have been proposed as important processes, the source mechanism of such a fluid concentration remains unresolved. In this contribution based on both field and experimental data, we demonstrate that viscous flow exerts a dynamic control on H2O-rich fluid circulation in mantle shear zones. Using the distributions of amphibole and olivine dislocation slip-systems, we first highlight H2O accumulation around fine-grained shear zones in the Ronda peridotite massif (Spain). These observations give rise to a long-term and continuous process of fluid pumping during ductile deformation, which strongly suggest creep cavitation as the driving mechanism. Secondly, we used secondary ion mass spectrometry to document the H2O content of fine-grained olivine across an experimental shear zone. The latter developed with grain size reduction during a H2O-saturated shear experiment at 1.2 GPa and 900 °C. Through data interpolation, the olivine matrix reveals high H2O concentrations where shear strain is localized. These concentrations far exceed the predicted amount of H2O that grain boundaries can contain, excluding diffusive fluid permeation as a unique source of water storage. We also show that the H2O content increases per unit of grain boundary across the shear zone, highlighting an excess volume of H2O that depends on strain and/or strain rate. Based on tensile experiments in metals, we propose that a larger pore volume is produced with increasing strain rate due to competition between creep cavitation and "healing" processes, which include phase nucleation. Altogether, our findings therefore support creep cavitation to occur in mantle shear zones, providing a dynamic process for H2O to be infiltrated and stored in the deep lithosphere

Water pumping in mantle shear zones: From field observations to experimental evidence

International audienceWater plays an important role in geological processes. Providing constraints on what may influence the distribution of aqueous fluids is thus crucial to understanding how H2O impacts Earth's geodynamics. In a deep-seated environment, viscous shear zones have been identified as sites of massive fluid circulation, with many implications for ores deposits and rock rheology. However, although seismic pumping, fluid permeation and/or creep cavitation have been proposed as important processes, the source mechanism of such a fluid concentration remains unresolved. In this contribution based on both field and experimental data, we demonstrate that viscous flow exerts a dynamic control on H2O-rich fluid circulation in mantle shear zones. Using the distributions of amphibole and olivine dislocation slip-systems, we first highlight H2O accumulation around fine-grained shear zones in the Ronda peridotite massif (Spain). These observations give rise to a long-term and continuous process of fluid pumping during ductile deformation, which strongly suggest creep cavitation as the driving mechanism. Secondly, we used secondary ion mass spectrometry to document the H2O content of fine-grained olivine across an experimental shear zone. The latter developed with grain size reduction during a H2O-saturated shear experiment at 1.2 GPa and 900 °C. Through data interpolation, the olivine matrix reveals high H2O concentrations where shear strain is localized. These concentrations far exceed the predicted amount of H2O that grain boundaries can contain, excluding diffusive fluid permeation as a unique source of water storage. We also show that the H2O content increases per unit of grain boundary across the shear zone, highlighting an excess volume of H2O that depends on strain and/or strain rate. Based on tensile experiments in metals, we propose that a larger pore volume is produced with increasing strain rate due to competition between creep cavitation and "healing" processes, which include phase nucleation. Altogether, our findings therefore support creep cavitation to occur in mantle shear zones, providing a dynamic process for H2O to be infiltrated and stored in the deep lithosphere

Evolution of H2O content in deforming quartz aggregates: An experimental study

International audienceDeformation experiments were carried out on pure quartzite samples (>99% quartz) with a grain size of ~200 μm from Tana, Northern Norway. Deformation conditions were 900 • C, 0.1 wt% H 2 O added, strain rate ~1 × 10-6 s-1 at variable confining pressures from 600 to 2000 MPa. Detailed FTIR measurements of H 2 O indicate that the H 2 O content in the grain boundary region is higher than that inside quartz grains. Hydrostatic treatment and deformation at the chosen temperature and pressure conditions lead to further H 2 O loss from grain interiors and H 2 O increase in the grain boundary region. Varying the confining pressure does not have an observable effect on the H 2 O transfer from grains to the grain boundary region. The 3585 cm-1 absorption band increases systematically with increasing confining pressure. As this band is associated with OH in dislocations, the increase may indicate an increased dislocation density with increasing pressure. The triplet of 3317, 3375 and 3438 cm-1 associated with Al-content in quartz increases in the grain boundary region indicating an exchange of H + together with Al 3+ for Si 4+. The Al and H exchange suggest dissolution-precipitation processes in the grain boundary region facilitating the movement of the quartz grain boundaries (grain boundary migration). The H 2 O in the grain boundary region will be important for enhancing grain boundary migration and thus recrystallization processes during deformation

Actin scaffolding by clathrin heavy chain is required for skeletal muscle sarcomere organization

The ubiquitous clathrin heavy chain (CHC), the main component of clathrin-coated vesicles, is well characterized for its role in intracellular membrane traffic and endocytosis from the plasma membrane (PM). Here, we demonstrate that in skeletal muscle CH