Section: Application Domains
Biological imagery
Recent progresses in molecular biology and light microscopy make henceforth possible the acquisition of multi-dimensional data (3d + time) at one or several wavelengths (multispectral imaging) and the observation of intra-cellular molecular dynamics at sub-micron resolutions. Automatic image processing methods to study molecular dynamics from image sequences are therefore of major interest, for instance, for membrane transport involving the movement of small particles from donor to acceptor compartments within the living cell.
The challenge is then to track gfp tags (fluorescent proteins for labeling) with high precision in movies representing several gigabytes of image data. The data are collected and processed automatically to generate information on partial or complete trajectories. In our research work, we are developing methods to perform the computational analysis of these complex 3d image sequences since the capabilities of most commercial image analysis tools for automatically extracting information are rather limited and/or require a large amount of manual interactions with the user.
Quantitative analysis of data obtained by fast 4d wide-field microscopy with deconvolution, confocal spinning-disk microscopy, Total Internal Reflectance Microscopy (TIRF), Fluorescence Recovery After Photobleaching (FRAP) combined with Green Fluorescence Protein (gfp )-tagging allows one to enlighten the role of specific proteins on HeLa human cell lines. Among these proteins, some are member of the family of Rab-gtp ases that bind reversibly to specific membranes within the cells. In our study, we aim at designing computational and statistical models to understand membrane trafficking and, more precisely to better elucidate the role of Rab family proteins inside their multiprotein complexes. We mainly focus on the analysis of transport intermediates (vesicles) that deliver cellular components to appropriate places within cells. Methods have been developed for interaction estimation between Rab11 and Langerin proteins, and dynamic estimation of Rab6a and Rab6a' proteins - involved in the regulation of transport from the Golgi apparatus to the endoplasmic reticulum.
Moreover, microscopic imaging at both the light and electron microscopic level provides multiscale unique information on protein localization and interactions, and extends and enriches that obtained from molecular and biochemical techniques. The 3D reconstruction of macromolecular structures form 2D EM (Electronic Microscopy) images of vitrified biological samples (Cryo-EM) has some advantages over other imaging techniques since it has proved to be an effective technique to investigate the structure of native cells with macromolecular resolution and preserve the whole integrity of the cell. Nevertheless, the high magnification available with EM comes with a limited field of view. Also, the very low contrast of unstained and vitrified biological specimens and the need to minimize the exposure to electron radiation make the identification of specific structures a difficult and time consuming task. Therefore one needs a gentle and time efficient way to locate structures of interest, improve image contrast and remove noise for a better interpretation of the image contents. We currently investigated image segmentation methods to analyze microtubule dynamics observed in Cryo-EM in collaboration with University of Rennes 1 - UMR 6026 (D. Chrétien).
Microtubules are long tubes of 25 nanometers in diameter formed with the 
-tubulin dimers and present in all eukaryotic cells. They play fundamental roles in cells life, in particular during their division since they are involved in chromosome segregation. The understanding of their structure and assembly mechanisms is of major importance not only in fundamental biological research, but also in medicine since microtubules are a major target of anti-mitotic drugs used in cancer therapy. The study of
their molecular architecture is possible by cryo-electron microscopy, which enables observation of biological specimens frozen at liquid nitrogen temperature in their native state. The images obtained are projection views of the specimens perpendicular to the electron beam that have a resolution on the order of a few (tenth of) Angströms.
In the coming 3 years, we will study interactions between neighboring protofilaments at the extremity of microtubules and the biological function of open sheet.
