Self-organized Rho and F-actin patterning in an artificial cell cortex

The cell cortex, comprised of the plasma membrane and an underlying meshwork of filamentous actin (F-actin), is remodeled during a variety of essential biological processes. Previous work has shown that prior to large-scale remodeling the cell cortex is dynamically patterned with subcellular waves of F-actin assembly and disassembly, a phenomenon termed “cortical excitability”. In developing embryos, cortical excitability is generated through coupled positive and negative feedback, with rapid activation of F-actin assembly driven by the small GTPase Rho, followed by inhibition of Rho activity. These feedback loops are proposed to serve as a mechanism for amplification of active Rho signaling at the cell equator to support furrowing during cytokinesis, while also maintaining flexibility for rapid error correction. Investigating the mechanisms that support and regulate cortical patterning is currently limited by a lack of technical approaches that can bridge our understanding of biochemical feedback signaling and cortical pattern formation, including the molecular regulation of signaling molecules, membrane dynamics, and cytoskeletal remodeling. A breakthrough in this gap in knowledge has been the development of an “artificial cortex”, made from supported lipid bilayers (SLBs) and Xenopus egg extract, which successfully reconstitutes active Rho and F-actin dynamics in a cell-free system. This reconstituted system spontaneously develops two distinct types of self-organized cortical dynamics: singular excitable Rho and F-actin waves, and non-traveling oscillatory Rho and F-actin patches. Like in vivo cortical excitability, patterning in the artificial cortex depends on Rho activity and F-actin polymerization. We also find that SLB fluidity directly influences the propensity for pattern formation in the artificial cortex, which suggests dynamic membranes support cortical patterning. These findings reveal that the cell cortex is a self-organizing structure and present a novel approach for investigating mechanisms of Rho-GTPase-mediated cortical dynamics.