Point de Contrôle de l'orientation du fuseau mitotique The group headed by Sylvie Tournier (Laboratoire de Biologie Cellulaire et Moléculaire du Contrôle de la Proliferation, CNRS UMR 5088) focuses on the segregation of chromosomes in the yeast Schizosaccharomyces pombe. The proteins of interest are fused with auto-fluorescent GFP. This team has a very strong activity in the field of imaging living cells applied to measures of force, and also carries out measures FRAP. Video-microscopy on wide-field microscope is the technique most commonly used by this team. Approaches using laser ablation, optical tweezers and 3D imaging SPIM (Single Plane Illumination Microscopy) are also used or being development.
Group Dynamics and Nuclear Organization, headed by Olivier Gadal (Laboratory of Eukaryotic Molecular Biology, CNRS UMR 5099) examines the relationship between positioning and transcriptional activity of genes within the nuclear volume in the yeast Saccharomyces cerevisiae. Using this model organism, we can work on large populations and thus access to statistical analysis. Using fast confocal microscope (spinning disk), located on the platform of Toulouse, allow the acquisition of 3D image over time. The nanometric scale are investigated in our team by Léger-Silvestre, using transmission electron microscopy (TEM). We plan to use électron tomography, newly set up on the platform of electron microscopy, to explore the 3D organization of the nucleus at the nanometer scale.
Cytosolic and nuclear calcium signaling in plants Christian Mazars (Surfaces Cellulaires et Signalisation chez les végétaux, UMR INRA 5546) is working in the group headed by Raoul Ranjeva and focuses on changes in calcium flux in the cytosol or the nucleus of tobacco cells. It uses recombinant aequorine, but wishes to develop probe-type calcium chameleon.
Functional dynamics of the nucleus and RNA transport The group led by Pierre-Emmanuel Gleizes (Laboratoire de Biologie Moléculaire Eucaryote, CNRS UMR 5099) study the assembly and export of ribosomal subunits and analysis of the consequences of this biogenesis disorders in certain human disease (anemia of Diamond and Blakfan). The study designs are the yeast Saccharomyces cerevisiae and cell lines in culture (HeLa, U2OS). In the field of microscopy, the group's activities revolve around two aspects. A wide optical, fluorescence (Gfp protein, probes) is visualized in fixed cells or living with a wide field microscope. The acquisition of stacks of images is followed by deconvolution and 3D visualization (Amira). Quantitative approaches with automatic data processing for a large number of cells are also developed (Metamorph). The electronic scale, the team works closely with the platform of Electron Microscopy. A new microscope (200 kV) will soon introduce electronic tomography analysis.
The group of Kerstin Bystricky (Laboratory of Eukaryotic Molecular Biology, CNRS UMR 5099) studies the structure of chromatin, the architecture of the nucleus and epigenetic mechanisms regulating gene expression. The models are the yeast Saccharomyces cerevisiae and breast cancer lines. We use techniques of immunofluorescence, in situ hybridization (preserving the 3D structure of nuclei, in particular for the study of the location of genes relative to their chromosome territory in human cells) and microscopy on living cells . The main objective is to achieve in living cells in 3D analysis of two (or three) proteins coupled with protein GFP to monitor simultaneously several chromosomal loci and relative to nuclear structures (envelope, SPB, nucleolus ).
Nano Ingénierie et Intégration des Systèmes A collaboration has also been established with Aurélien Bancaud (Laboratoire d'Architecture et d'Analyze des Systèmes, CNRS UPR 8001) to develop a technique using a path consisting of triangular reflectors. From a single image of a field acquired in large, it is then possible to reconstruct the biological objects in 3D.
C/D RNAs, microRNAs and genomic imprinting The team “microRNAs, RNA CD and parental genomic imprint”, headed by Jérôme Cavaillé (Laboratory of Eukaryotic Molecular Biology, CNRS UMR 5099), currently study two families of small non-coding RNAs: C/D RNAs (or RNA methylation guides) and microRNAs (or miRNAs) the genes of which – mainly expressed within the brain - are subjected to genomic imprinting at two different chromosomal loci: 14q32 (human)/distal 12 (mouse) and 15q11q13(human)/7C(mouse). Our aim is to understand the molecular pathways in which these small RNAs are involved (i.e. search for RNA targets) as well as their biological functions (i.e. studies of transgenic or knock-out mice models). These projects are motivated by 3 main hypotheses: (1) brain-specific mRNAs can be 2’-O-methylated by C/D RNAs (2) miRNAs can regulate the expression of imprinted gene(s) (3) lack of expression of C/D RNA genes might contribute to a human disease, the Prader-Willi syndrome.
Laboratoire de Génétique Cellulaire Martine Yerle (Laboratoire de Génétique Cellulaire, INRA UMR 444) develops within the group led by Pierre Mulsant, in close collaboration with the veterinary school, genomic (DNA) and cytogenetic pigs (3D confocal microscopy). In particular, the group analyzes the change in the position of nuclear target genes during cell activation and examines how the position affects the level of gene expression. These tests are developed in collaboration with Thomas Boudier (Paris) and Alain Jauneau (Toulouse). They have been important developments in the field of segmentation in order to position these genes vis-à-vis chromosome territories.
Structure and segregation of bacterial chromosomes François Cornet (Laboratoire de Microbiologie et Génétique Moléculaires, UMR 5100) investigate the mechanism of chromosome dynamics in bacteria, with functional approaches in the model bacteria Escherichia coli. In particular we are studying the specific events occurring inside the ter region. This region and the different events it specifically hosts are of crucial importance for chromosome dynamics. It is the region of the last operations of sister chromosome separation: termination of replication, removal of the intercatenation links, and resolution of dimeric chromosomes. It is also a highly plastic region submitted to frequent gene acquisition. We are studying the Xer site-specific recombination system that resolves chromosome dimers during cell division, in particular how it is controlled and integrated into the cell cycle and how it catalyses the integration of foreign DNA encoding adaptation and pathogenicity determinants in ter.
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