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Proteomics groupProteomics, Metals and Differentiation
 Team leader |
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| Staff members |
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Proteomics group |
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| Mireille Chevallet, CEA researcher Cécile Lelong, UJF associate professor Sylvie Luche, UJF technician |
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Cell biology group |
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| Catherine Aude-Garcia, CEA researcher Serge Candéias, CEA researcher Véronique Collin-Faure, CEA technician Isabelle Testard, CEA researcher |
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The research team has two lines of action in research. One project deals with technological developments for proteomics, and the main project deals with the response of cells to metallic ions and nanoparticles.
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| General theme and rationale of the research project |
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Our project will study how metallic ions and metallic oxides impact myeloid cells. There is a clear relationship between various diseases and the phagocytosis of toxic (nano)particulate material by myeloid cells, which are highly phagocytic. For example, this holds true for asbestos and mesotheliomas, silica and silicosis, and this is also highly suspected for at least part of the toxic effects of tobacco smoke or diesel particles.
As a matter of facts, this is due to the primary scavenging function of some myeloid cells such as macrophages. When such cells phagocyte particles, they try to degrade them in their lysosomes. This can lead to the liberation of toxics adsorbed on the phagocytized particles. In the case of metal-containing particles, the toxics can be heavy metal ions or transition metal ions that will be dissolved by the acidic conditions prevailing in the lysosome. Besides such toxic effects, macrophage activation following phagocytosis of particulate material also results in the production of pro-inflammatory mediators, and this inflammation, if it becomes chronic, can be part of the etiology of some diseases such as cancer. Some of these factors are known, such as TNF, IL-6 or IL-12, but the complete scope of these proteins is not known yet. There is thus a high interest to understand how myeloid cells and especially macrophages react to particles, and especially to metal containing nanoparticles, as well as the corresponding metallic ions. Our project goes beyond the simple observation of toxic effects, and aim at providing molecular mechanisms and a better understanding of the effects of these agents. Classically, this type of research will be carried out by focused approaches on targets selected on the basis of previous knowledge, and we will not neglect such approaches. However, we also believe that proteomics can deliver significant knowledge, owing to its wide and unbiased scope. Indeed, some work exists in this direction, either on the toxicity of metallic ions or on the effects of metallic oxide particles. Our research project will therefore follow three directions, a targeted one and two based on proteomics. |
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| Core research actions |
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Proteomics study of the effects of metals on myeloid cells |
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In this subproject, we will study by proteomics the effects of metal ions and metallic particles on myeloid cells.
To investigate the effects of metallic particles and ions by proteomics, we will use the same staggered strategy that we have used to investigate the differentiation of these cells. First, we will perform a comparative proteomics analysis at the scale of whole cell extracts. In this case, the fact that proteomics mainly shows the cellular stress response proteins will not be a problem, as this is exactly what we want to investigate, i.e. the details of the stress response that is made by the cells under the various conditions tested. We will then target the secreted proteins, as this will give us access to an unbiased view of the inflammatory responses. |
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Targeted studies |
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While proteomics offer the power of large scale and unbiased studies, its requirements in terms of sample consumption and its limitations in the analysis of lower abundance proteins does not make it a suitable tool for the detailed analysis of precise phenomena, especially for in vivo studies where the sample resources are often limited. This is why we will also carry out targeted studies on the response of myeloid cells to various stimuli, including metals. As the scope of such studies is overwhelmingly high, we will use the know-how present in the team and focus our targeted studies on proteins involved in DNA repair and DNA structuration in nuclei. More specifically, using in vitro and ex vivo models developed in the ProMD team, we will investigate how DNA repair protein gene transcription is controlled by soluble factors, and try to unravel the signalling pathways involved. We will also explore post-transcriptional regulation of these proteins expression. The stimulation of DNA repair activities by soluble factors will be assessed by survival of the cells following different genotoxic stress and/or by specific tests aimed at measuring activity toward specific individual lesions. The soluble factors of interest will of course be metal ions, as these are known to alter lymphoid differentiation, but also cytokines, which are known to modulate myeloid differentiation but whose synthesis may also be altered by the presence of metals. |
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Metalloproteomics |
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The previous two projects will give us insight into the reaction of the myeloid cells to the metallic stress, in other words to the output of the cellular system. However, they will not give us ideas on the entry point of the metallic stress in the cells, in other words the input signal. To gain insights in this direction, we will also carry out a metalloproteomics project. The rationale of this project is that the primary targets of the metallic stress, at least in the case of metallic ions, will be the proteins that bind this ion. Thus, we will try to fish out in the cell proteome the proteins that bind specific metal ions. To do so, we have decided to perform a preselection of these proteins by immobilized metal affinity chromatography, followed by a classical proteomics analysis. This strategy has already been applied by some groups, but not to myeloid cells We will also investigate the effects of changing the number and geometry of coordination valencies implied in the interaction with the solid phase, thereby changing the valencies available for the metal-binding site of the proteins and thus the specificity of binding. This will require to test and compare several metal hemichelators, some of which could be designed and prepared by the coordination chemists of other teams of the laboratory. |
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| Technology for proteomics |
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We are also deeply implicated in technological projects aiming at improving the performances of the proteomics toolbox, especially in the field of protein separation. We strongly believe that proteomics must continue to deal with complete proteins with their associated post-translational modifications, and cannot be reduced only to the study of digestion peptides, with the associated loss of filiation between the proteins and the peptides. We are thus working in the field of protein solubilization prior to electrophoresis, on the improvement of the electrophoretic methods per se and also on the field of protein detection after gel electrophoresis (see the associated bibliography of the team).
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| Bibliography of the team |
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Most of the group's recent papers can be found at the following url.
The author's pdf files are freely downloadable. This distribution is restricted to author's files, and not final journal's files, to comply to copyright laws. |
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Aude-Garcia C, Collin-Faure V, Luche S and Rabilloud T |