CROSS-TALK BETWEEN RAL AND RAC PATHWAYS IN THE CONTROL OF CELL MIGRATION

SUMMARY (English) Very little is known about the coordination and the integration among the different regulators of the motility process. This work deals with two migration-regulatory pathways: the Rac1/WRC (Wave Regulatory Complex) pathway that drives the formation of the actin polymerization network at the front of motile cells; and RalB/exocyst pathway for which the molecular mechanisms underlying its implication in cell motility were still largely unknown at the beginning of this thesis. Rac1 and RalB are small GTPases of the Rho and Ras family, respectively. WRC and exocyst complexes are their direct effectors. In searching for connections between the exocyst and migration regulators, we found that two subunits of the exocyst, Exo70 and Sec6, interact directly in vitro with two subunits of the WRC, Abi and Cyfip, respectively. Moreover, we found that exocyst subunits can interact in vitro with the whole fully-assembled WRC complex. We also showed that these two complexes associate in vivo. Functionally, the exocyst was required for WRC complex positioning at the front of migrating cells. On the other hand, we also found that two other subunits of the exocyst, Sec8 and Exo84, interact with SH3BP1 (a RhoGAP protein) by two-hybrid assay and by co-immunoprecipitation. SH3BP1 localizes at the leading edge and this localization is dependent on the exocyst. Interestingly, in vivo, the RalB/exocyst/SH3BP1 pathway specifically targets Rac1, and not Cdc42. By a combination of approaches we concluded that SH3BP1 is required to inactivate Rac1 at the front. In our model we propose that RalB/exocyst regulates cell migration by driving to the leading edge two key signaling elements of the Rac1 pathway: its effector WRC, that stimulates actin filament nucleation, and its negative regulator SH3BP1, a GAP promoting Rac1 inactivation and GDP/GTP cycling. In conclusion, this work provides novel molecular and functional links between polarized exocytosis and actin dynamics during cell motility.