Molecular mechanisms of intracellular transport
Keywords :
Golgi complex, Rab GTPases, intracellular transport, model membranes, cell cycle, cytokinesis
Group leader : Bruno Goud
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Team « Molecular mechanisms of intracellular transport »
Our team focuses on the mechanisms that govern vesicular transport and membrane trafficking in eukaryotic cells, with emphasis on the Golgi complex. Our goal is to better understand these complex events, both from a cell biology point of view (identification and functional characterization of proteins involved) and from a physics point of view (understanding of the physical parameters underlying transport events).
1) Function of Rab GTPases in membrane trafficking and organelle biogenesis
The Rab GTPases are key regulators of intracellular transport. We are particularly interested in Rab6 and Rab11, respectively associated with the Golgi and recycling endosomes. The latest significant results were the identification and characterization of several new effectors of Rab6, particularly myosin II (Miserey-Lenkei et al. 2010. Nat Cell Biol 12:645-54) and myosin Vb. A two-hybrid screen with Rab6 effectors identified other Rab proteins whose function could be coupled with that of Rab6, especially Rab2 and Rab39. We have characterized the function of a Rab6 isoform (Rab6C), a retrogene encoding a protein associated with the centrosome and involved in cell cycle (Young et al. 2010. J Mol Biol 397:69-88).
With regard to Rab11, we have clarified the function of this protein and two of its partners (Rab11FIP2 and MyosinVb) in the recycling of langerin (lectin present in the Birberck granules of Langerhans cells) (Gidon et al, submitted for publication). In collaboration with the team of F. Radvanyi (UMR 144), we have studied the expression of 284 genes encoding Rab proteins and their effectors in bladder cancer. The functional validation of the deregulated genes is underway.
2) Towards the understanding of global cell architecture
This project that we recently initiated in the laboratory, aims at understanding the principles that govern the organization of cellular compartments in the cell space. For this purpose, we have combined micropatterning technology with a novel computational imaging approach in order to quantify the 2D or 3D steady-state organization of diverse endomembranes with probabilistic density maps (Schauer et al. 2010. Nature Methods 7:560-566) (Fig. 1). This approach is now used to identify cellular and extracellular factors underlying the organization of the cell compartments.
Figure1: Analysis of CD63-, Rab5-, Sec13- and Rab6-positive endomembranes on crossbow-shaped patterns. Maximum intensity projection of the corresponding deconvolved immuno-fluorescence images (first row), the 2D density estimation maps (second row) and the 3D dens
3) Understanding physical parameters underlying transport events
For several years, we study the physical parameters underlying intracellular transport using model membranes (Giant Unilamellar Vesicles or GUVs). We focus in particular on sorting and fission processes and we have highlighted the important role played by membrane curvature and membrane tension in these processes. Experiments are underway to try to measure membrane tension of cellular compartments in vivo. We also recently studied the binding and activity of ArfGAP1 (“GTPase activating protein” for the Arf1 protein involved in the formation of COPI coat) on membranes with different curvatures and showed the existence of a gradient of Arf1 concentration on the curved regions (Fig. 2). These results suggest that in the cell, the dissociation of the COPI coat follows the step of fission (Ambroggio et al. 2010. EMBO J 29:292-303).
Figure2: Generation of an Arf1 gradient on a membrane nanotube pulled from a GUV by optical tweezers.
4) Relationships between the Golgi complex and centrosome
We are interested in the function of proteins such as GMAP-210 or AKAP-450, capable of binding to both the centrosome and Golgi membranes (Rivero et al. 2009, EMBO J 28:1016:28; Cardenas and al. 2009, BMC Biol 7:56). This study is done in collaboration with the group of Rosa Rios (University of Seville, Spain).