Mechanistic of Rac and Cdc42 synchronization at the cell edge by ARHGAP39-dependent signaling nodules and the impact on protrusion dynamics

Compartmentalization of GTPase regulators into signaling nodules dictates the GTPase pathways selected. Rac and Cdc42 are synchronized at the cell edge for effective protrusion in motile cells but how their activity is coordinated remains elusive. Here, we discovered that ARHGAP39, a Rac and Cdc42 GTPase-activating protein, sequentially interacts with WAVE and mDia2 to control Rac/lamellipodia and Cdc42/filopodia protrusions, respectively. Mechanistically, ARHGAP39 binds to WAVE and, upon phosphorylation by Src kinase, inactivates Rac to promote Cdc42-induced filopodia formation. With our optimized FRET biosensor, we detected active Cdc42 at the filopodia tips that controls filopodia extension. ARHGAP39 is transported to filopodia tips by Myosin-X where it binds mDia2 and inactivates Cdc42 leading to filopodia retraction. Failure in lamellipodia to filopodia switch by defective ARHGAP39 impairs cell invasion and metastasis. Our study reveals that compartmentalization of ARHGAP39 within Rac/Cdc42 signaling nodules orchestrates the synchronization of lamellipodia/filopodia crosstalk and highlights the intricate regulation of leading edge dynamics in migrating cells.