The mechanism of active GTPase movement and actomyosin ring organization during wound healing

Transient actomyosin arrays comprised of F-actin and Myosin-2 are essential to drive numerous cellular processes including those observed during single-cell wound healing. During this process, key regulators of cytoskeletal dynamics-Rho and Cdc42-form discrete zones of activity around the wound perimeter, where Rho activity is primarily confined to the wound edge and is circumscribed by a zone of Cdc42 activity. These concentric zones of GTPase activity move forward with remarkable precision that mirrors the movement of the actomyosin ring they are regulating. This study demonstrated that the apparent movement of the zones of active Rho and Cdc42 is not a result of direct transport of the active GTPases, but rather a product of a treadmilling-type mechanism where inactivation is focused toward the trailing edges of their respective zones. This study also characterized a novel fluorescent F-actin reporter based on the Calponin homology domain of Utrophin (UtrCH), which binds F-actin without stabilizing it in vitro. This F-actin reporter faithfully reported F-actin populations, distinguished between stable and dynamic F-actin, and showed no obvious effects on processes that depend on a balance of F-actin assembly/disassembly. Together these studies present a model where actomyosin arrays formed during wound healing are templated by the small GTPases and GTPase treadmilling, and organize into higher-ordered structures with differential biological activities and cytoskeletal dynamics.