Scrib controls Cdc42 localization and activity to promote cell polarization during astrocyte migration

Background: Mammalian Scribble (Scrib) plays a conserved role in polarization of epithelial and neuronal cells. Polarization is essential for migration of a variety of cell types; however, the function of Scrib in this context remains unclear. Scrib has been shown to interact with beta PIX, a guanine nucleotide exchange factor for the small GTPases Rac and Cdc42. Cdc42 controls cell polarity from yeast to mammals during asymmetric cell division and epithelial cell polarization, as well as during cell migration. Cdc42 is, in particular, required for polarization and orientation of astrocytes in a scratch-induced polarized migration assay. Using this assay, we characterized Scrib function during polarized cell migration. Results: Depletion of Scrib by siRNA or expression of dominant-negative constructs inhibits astrocyte polarization. Like Cdc42, Scrib controls protrusion formation, cytoskeleton polarization, and centrosome and Golgi reorientation. Scrib interacts and colocalizes with beta PIX at the front edge of polarizing astrocytes. Perturbation of Scrib localization or of Scrib-beta PIX interaction inhibits beta PIX polarized recruitment. We further show that beta PIX is required for astrocyte polarization and that both the Scrib-binding motif and the GEF activity of PPIX are essential for its function. Scrib and beta PIX control Cdc42 activation and localization during astrocyte polarization. Thereby, Scrib regulates Cdc42-dependent APC and Dlg1 recruitment to the leading edge to promote cell orientation. Conclusion: We conclude that Scrib plays a key role in the establishment of cell polarity during migration. By interacting with beta PIX, Scrib controls localization and activation of the small GTPase Cdc42 and regulates Cdc42-dependent polarization pathways.