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Dive into the research topics where Sophie Cribier is active.

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Featured researches published by Sophie Cribier.


Biophysical Journal | 2000

Translational Diffusion of Globular Proteins in the Cytoplasm of Cultured Muscle Cells

Martine Arrio-Dupont; Georges Foucault; Monique Vacher; Philippe F. Devaux; Sophie Cribier

Modulated fringe pattern photobleaching (MFPP) was used to measure the translational diffusion of microinjected fluorescein isothiocyanate (FITC)-labeled proteins of different sizes in the cytoplasm of cultured muscle cells. This technique, which is an extension of the classical fluorescence recovery after photobleaching (FRAP) technique, allows the measurement of the translational diffusion of macromolecules over several microns. Proteins used had molecular masses between 21 and 540 kDa. The results clearly indicated that the diffusivity of the various proteins is a decreasing function of their hydrodynamic radius. This decrease is more rapid with globular proteins than with FITC-labeled dextrans (, Biophys. J. 70:2327-2332), most likely because, unlike globular proteins, dextrans are randomly coiled macromolecules with a flexible structure. These data do not exclude the possibility of a rapid diffusion over a short distance, unobservable with our experimental set-up, which would take place within the first milliseconds after bleaching and would correspond to the diffusion in restricted domains followed by impeded diffusion provoked by the network of microtubules, microfilaments, and intermediate filaments. Thus our results may complement rather than contradict those of Verkman and collaborators (, J. Cell Biol. 138:1-12). The biological consequence of the size-dependent restriction of the mobility of proteins in the cell cytoplasm is that the formation of intracellular complexes with other proteins considerably reduces their mobility.


Biophysical Journal | 1996

Diffusion of fluorescently labeled macromolecules in cultured muscle cells.

Martine Arrio-Dupont; Sophie Cribier; Georges Foucault; Philippe F. Devaux; A. d'Albis

Myotubes were obtained from culture of satellite cells. They had a sarcomeric organization similar to that of muscle. The diffusion in the direction perpendicular to the fibers of microinjected fluorescein isothiocyanate-dextrans of molecular weight ranging from 9500 to 150,000 was examined by modulated fringe pattern photobleaching. On the time scale of the observation, 10-30 S, all of the dextrans were completely mobile in the cytoplasm. The diffusion coefficients were compared to the values obtained in water. The ratio D(cytoplasm)/D(w) decreased with the hydrodynamic radius R(h) of the macromolecules. The mobility of inert molecules in muscle cells is hindered by both the crowding of the fluid phase of the cytoplasm and the screening effect due to myofilaments: D(cytoplasm)/D(w) = (D/D(w)) protein crowding x (D/D(w))(filament screening). The equation (D/D(w))filament screening = exp(-K(L)RCh) was used for the contribution of the filaments to the restriction of diffusion. A free protein concentration of 135 mg/ml, a solvent viscosity of cytoplasm near that of bulk water, and a calculated K(L) of 0.066 nm(-1), which takes into account the sarcomeric organization of filaments, accurately represent our data.


Biophysical Journal | 2008

A 20-nm Step toward the Cell Membrane Preceding Exocytosis May Correspond to Docking of Tethered Granules

Erdem Karatekin; Viet Samuel Tran; Sébastien Huet; Isabelle Fanget; Sophie Cribier; Jean-Pierre Henry

In endocrine cells, plasma membrane (PM)-bound secretory granules must undergo a number of maturation stages (i.e., priming) to become fusion-competent. Despite identification of several molecules involved in binding granules to the PM and priming them, the exact nature of events occurring at the PM still largely remains a mystery. In stimulated BON cells, we used evanescent wave microscopy to study trajectories of granules shortly before their exocytoses, which provided a physical description of vesicle-PM interactions at an unprecedented level of detail, and directly lead to an original mechanistic model. In these cells, tethered (T), nonfusogenic, vesicles are prevented from converting to fusogenic, docked (D) ones in resting conditions. Upon elevation of calcium, T-vesicles perform a 21-nm step toward the PM to become D, and fuse approximately 3 s thereafter. Our ability to directly visualize different modes of PM-attachment paves the way for clarifying the exact role of various molecules implicated in attachment and priming of granules in future studies.


Biochimica et Biophysica Acta | 1993

Quantitative comparison between aminophospholipid translocase activity in human erythrocytes and in K562 cells

Sophie Cribier; Josette Sainte-Marie; Philippe F. Devaux

Spin-labeled phospholipids were used to determine the transbilayer movement of phospholipids in human erythrocytes, in K562 cells and in human neonatal red cells. The erythroleukemia cell line, K562, as well as human neonatal red cells, which are rich in reticulocytes, were considered as representative of human erythrocyte precursor cells. In the nucleated cells, the difference between outside-inside movement of aminophospholipids and that of phosphatidylcholine or sphingomyelin analogues allowed us to discriminate between lipid internalization due to aminophospholipid translocase activity and to endocytosis. From the initial rates of aminophospholipid inward movement, we inferred that the activity of the aminophospholipid translocase is higher in the precursor cells than in mature erythrocytes.


Annals of the New York Academy of Sciences | 2004

Serotonin secretion by human carcinoid BON cells.

Viet Samuel Tran; Anne-Marie Marion-Audibert; Erdem Karatekin; Sébastien Huet; Sophie Cribier; Koulm Guillaumie; Catherine Chapuis; Claire Desnos; François Darchen; Jean-Pierre Henry

Abstract: BON cells are human carcinoid cells that secrete serotonin (5‐HT) and various peptides. Secretion of [3H]5‐HT by cell cultures was investigated. Acetylcholine (Ach) stimulated secretion through a somatostatin‐sensitive muscarinic pathway, whereas isoproterenol was inefficient. [3H]5‐HT secretion also was induced by Ca2+ in the presence of the ionophore A‐23187 or after digitonin permeabilization. These two processes were insensitive to stomatostatin. Ba2+ induced an efficient somatostatin‐sensitive [3H]5‐HT secretory response. Secretion also was analyzed at the single‐cell level, using carbon fiber amperometry and evanescent‐field fluorescence microscopy, after labeling the secretory vesicles by transfection of the cells with a NPY‐GFP construct. Both techniques revealed slow kinetics of secretory responses, suggesting that ready‐to‐fuse vesicles do not accumulate in these cells. Single secretory vesicles were imaged either in resting conditions or after addition of Ca2+ ions to digitonin‐permeabilized cells. The three‐dimensional movements of the vesicles before exocytosis were analyzed. The mean velocity of vesicles that released their content was lower than that of silent ones. Even in the case of mobile vesicles, exocytosis often was preceded by a period of arrest lasting at least 15 seconds, consistent with a docking/priming step.


Angewandte Chemie | 2012

Photocontrol of the Translocation of Molecules, Peptides, and Quantum Dots through Cell and Lipid Membranes Doped with Azobenzene Copolymers

S. Sebai; Dimitra Milioni; Astrid Walrant; Isabel D. Alves; Sandrine Sagan; Cécile Huin; Loïc Auvray; Dominique Massotte; Sophie Cribier; Christophe Tribet

Light opens: Photocontrolled transmembrane passage of soluble dyes and delivery of small peptides into mammalian cells has been achieved using azobenzene-modified polymers (AMPs) as permeabilizing agents. Irradiation with UV and visible light triggers polarity switches upon cis-trans isomerization of the azobenzene moieties. Photoresponsive permeability and pore opening promoted by trans-AMPs, but not cis-AMPs, in supported lipid bilayers are observed.


Langmuir | 2010

Permeabilization of lipid membranes and cells by a light-responsive copolymer.

S. Sebai; Sophie Cribier; Ali Karimi; Dominique Massotte; Christophe Tribet

Membrane permeabilization is achieved via numerous techniques involving the use of molecular agents such as peptides used in antimicrobial therapy. Although high efficiency is reached, the permeabilization mechanism remains global with a noticeable lack of control. To achieve localized control and more gradual increase in membrane perturbation, we have developed hydrophobically modified poly(acrylic acid) amphiphilic copolymers with light-responsive azobenzene hydrophobic moieties. We present evidence for light triggered membrane permeabilization in the presence azobenzene-modified polymers (AMPs). Exposure to UV or blue light reversibly switches the polarity of the azobenzene (cis-trans isomerization) in AMPs, hence controlling AMP-loaded lipid vesicles permeabilization via in situ activation. Release of encapsulated probes was studied by microscopy on isolated AMP-loaded giant unilamellar vesicles (pol-GUVs). We show that in pH and ionic strength conditions that are biologically relevant pol-GUVs are kept impermeable when they contain predominantly cis-AMPs but become leaky with no membrane breakage upon exposure to blue light due to AMPs switch to a trans-apolar state. In addition, we show that AMPs induce destabilization of plasma membranes when added to mammal cells in their trans-apolar state, with no loss of cell viability. These features make AMPs promising tools for remote control of cell membrane permeabilization in mild conditions.


Biochimie | 1998

Physical techniques for the study of exocytosis in isolated cells

Jean-Pierre Henry; François Darchen; Sophie Cribier

Membrane traffic is an important aspect of cell biology which implies shuttle vesicles and multiple binding/fusion events. In spite of rapid progress at the biochemical level, the mechanism of fusion is still not understood. A detailed physical description of the phenomenon is possible at the level of the plasma membrane where secretory vesicles fuse with the cell membrane, a process known as exocytosis. This process is specially active in neurons (release of neurotransmitter) and in endocrine cells (release of hormones), where exocytosis is tightly regulated. Among the biophysical techniques developed, cell membrane capacitance measurements by the technique of patch-clamp and amperometry of the oxidizable secretory products have resulted in interesting information. These techniques have described the initial fusion pore, its fluctuations, the efflux of material through the pore and its irreversible expansion. Optical techniques, using bioluminescent and fluorescent probes are also in progress. For instance, the dye FM 1-43 binds to but is not translocated through biological membranes and it has been used to measure membrane surface, as done by capacitance measurement. Evanescent wave fluorescence microscopy has been recently introduced to analyse the behaviour of secretory granules in the vicinity of the plasma membrane.


Biochemical Journal | 2000

Interactions between beta-enolase and creatine kinase in the cytosol of skeletal muscle cells.

Georges Foucault; Monique Vacher; Sophie Cribier; Martine Arrio-Dupont

We studied interactions in vivo between the cytosolic muscle isoform of creatine kinase (M-CK) and the muscle isoform of 2-phospho-D-glycerate hydrolyase (beta-enolase) in muscle sarcoplasm by incubating glycerol-skinned fibres with FITC-labelled beta-enolase in the presence or absence of free CK. A small amount of bound beta-enolase was observed in the presence of large concentrations of CK. The mobility of enolase was measured in cultured satellite cells by modulated-fringe-pattern photobleaching. FITC-labelled beta-enolase was totally mobile in both the presence and the absence of CK but its diffusion coefficient was slightly lower in the presence of CK. This suggests a weak interaction in vivo between enolase and CK.


Scientific Reports | 2016

Quantitative fluorescence spectroscopy and flow cytometry analyses of cell-penetrating peptides internalization pathways: optimization, pitfalls, comparison with mass spectrometry quantification

Françoise Illien; Nicolas Rodriguez; Mehdi Amoura; Alain Joliot; Manjula Pallerla; Sophie Cribier; Fabienne Burlina; Sandrine Sagan

The mechanism of cell-penetrating peptides entry into cells is unclear, preventing the development of more efficient vectors for biotechnological or therapeutic purposes. Here, we developed a protocol relying on fluorometry to distinguish endocytosis from direct membrane translocation, using Penetratin, TAT and R9. The quantities of internalized CPPs measured by fluorometry in cell lysates converge with those obtained by our previously reported mass spectrometry quantification method. By contrast, flow cytometry quantification faces several limitations due to fluorescence quenching processes that depend on the cell line and occur at peptide/cell ratio >6.108 for CF-Penetratin. The analysis of cellular internalization of a doubly labeled fluorescent and biotinylated Penetratin analogue by the two independent techniques, fluorometry and mass spectrometry, gave consistent results at the quantitative and qualitative levels. Both techniques revealed the use of two alternative translocation and endocytosis pathways, whose relative efficacy depends on cell-surface sugars and peptide concentration. We confirmed that Penetratin translocates at low concentration and uses endocytosis at high μM concentrations. We further demonstrate that the hydrophobic/hydrophilic nature of the N-terminal extremity impacts on the internalization efficiency of CPPs. We expect these results and the associated protocols to help unraveling the translocation pathway to the cytosol of cells.

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Philippe F. Devaux

Centre national de la recherche scientifique

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Sandrine Sagan

École Normale Supérieure

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Jean-Pierre Henry

Centre national de la recherche scientifique

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Georges Foucault

Centre national de la recherche scientifique

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Martine Arrio-Dupont

Centre national de la recherche scientifique

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Sébastien Huet

Centre national de la recherche scientifique

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Viet Samuel Tran

Centre national de la recherche scientifique

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Frédéric Pincet

École Normale Supérieure

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Isabelle Fanget

Centre national de la recherche scientifique

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Monique Vacher

Centre national de la recherche scientifique

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