Sandrine Etienne-Manneville (website)
The small G protein Cdc42 plays an evolutionary conserved role in cell polarity. It contributes to the polarization and directed migration of epithelial cells and astrocytes. The goal of this project is to better determine the regulatory mechanisms of Cdc42 localisation at the cell cortex. SCRIB is a large PDZ domain containing protein capable of organizing sub-membranous protein complexes controlling cell polarity. We have previously identified the Scrib PDZ domains as key mediators of the polarity pathway leading to Cdc42 activation and PAR proteins recruitment at the leading edge cell cortex. Despite being central for its function, the molecular basis of Scrib recruitment and stabilization at the cell cortex is poorly understood. In addition to the amino-terminal LRR domains which link Scrib to the cell cortex by yet unclear mechanisms, we have recently observed that the carboxyterminal domain may also play a role. Part of this project will be devoted to the functional characterization of newly identified Scrib binding partners in collaboration with the Boxem group, who specialize in identification of binding domains by two hybrid. We will then determine if the key cortical partners of Scrib are involved in the perturbation of Scrib and Cdc42 localization and cell polarity in cancer cells of glial and epithelial origin.
We have previously demonstrated that Cdc42 and its GEF bPIX were recruited to Scrib cortical domains via a Arf6 dependent vesicular traffic. One hypothesis is that Scrib binding to bPIX leads to the activation of Cdc42. These events will be followed by video microscopy and immunofluorescence to determine where Cdc42 associates with cytoplasmic membrane compartments and where it is activated. Raichu probes will be used to localize Cdc42 activity during polarization of migrating cells and during polarization of epithelial cells. The role of these molecular interactions in cell polarity will be determined by the organisation of the microtubule network in migrating astrocytes and in polarized epithelial cells (in collaboration with the Akhmanova group). Finally, the speed, the direction and the persistence of migration will be analysed in appropriate in vitro assays.
We expect to provide a clearer view of the fundamental polarity signals and in particular in the molecular mechanisms responsible for Cdc42 recruitment at the cell cortex and the exact role of Scrib in these events.
- Osmani N, Vitale N, Borg J-P, Etienne-Manneville S. Scrib controls Cdc42 localization and activity to promote cell polarization during astrocyte migration. Curr Biol CB. 2006 Dec 19;16(24):2395–405.
- Nola S, Sebbagh M, Marchetto S, Osmani N, Nourry C, Audebert S, et al. Scrib regulates PAK activity during the cell migration process. Hum Mol Genet. 2008 Nov 15;17(22):3552–65.
- Osmani N, Peglion F, Chavrier P, Etienne-Manneville S. Cdc42 localization and cell polarity depend on membrane traffic. J Cell Biol. 2010 Dec 27;191(7):1261–9.
- Boëda B, Etienne-Manneville S. Spectrin binding motifs regulate Scribble cortical dynamics and polarity function. eLife. 2015;4.