Assaying Rho GTPase-Dependent Processes in Dictyostelium discoideum

The model organism D. discoideum is well suited to investigate basic questions of molecular and cell biology, particularly those related to the structure, regulation, and dynamics of the cytoskeleton, signal transduction, cell–cell adhesion, and development. D. discoideum makes use of Rho-regulated signaling pathways to reorganize its cytoskeleton during chemotaxis, endocytosis, and cytokinesis. In this organism the Rho family encompasses 20 members, several belonging to the Rac subfamily, but there are no representatives of the Cdc42 and Rho subfamilies. Here we present protocols suitable for monitoring the actin polymerization response and the activation of Rac upon stimulation of aggregation competent cells with the chemoattractant cAMP.