Cytoskeleton: Filopodia self-assemble

Filopodia are finger-like projections made up of bundled actin filaments that have many biological roles, including cell migration. How filopodia are formed has been difficult to determine, as current experimental systems do not recapitulate aspects of filopodium biology. Kirschner and colleagues now address this question using an in vitro model system in which filopodia can self-assemble in the absence of a pre-existing cytoskeletal network.

To examine how filopodia are formed, the authors incubated frog egg extracts with lipid bilayers (and not liposomes, as had been done previously) to recapitulate actin nucleation at membranes in vitro. Dense, long structures grew from the bilayer surface that were made up of bundled actin filaments. Actin polymerization occurred at the tip of the structures, where, similarly to filopodia, proteins such as vasodilator-stimulated phosphoprotein (VASP), neural Wiskott–Aldrich syndrome protein (NWASP), CDC42 and the formin diaphanous 2 (DIA2), localized. Moreover, the kinetics of actin monomer addition at the tip were similar to those of filopodium growth in vivo. Together, these findings indicate that these structures (which the authors term filopodium-like structures (FLSs)) resemble filopodia and mimic filopodium formation in vivo.

So how do filopodia assemble? Using FLSs as an in vitro model system, the authors could not detect preformed domains on the membranes, indicating that filopodium assembly is not template driven. Instead, filopodia self-assemble on permissive membrane surfaces enriched in phosphatidylinositol (4,5)-bisphosphate. Specifically, transducer of CDC42-dependent actin assembly 1 (TOCA1; also known as FNBP1L), which interacts with membrane lipids through its F-BAR domain, is recruited early to sites where FLSs later form. This in turn recruits NWASP, followed by the actin-related protein 2/3 (ARP2/3) complex, actin, VASP, DIA2 and the bundling protein fascin. Interestingly, ARP2/3 complex-driven actin polymerization is necessary for the initiation of FLS formation but is not strictly required for elongation. Instead, a significant reduction of FLS elongation was seen only when both ARP2/3 and diaphanous-related formins were inhibited, suggesting that they both have roles in this process.

On the basis of their findings, the authors propose that filopodia self-assemble in a step-wise manner: negatively charged membranes signal the recruitment of F-BAR domain superfamily proteins, such as TOCA1, which in turn recruit nucleation-promoting factors, such as NWASP, leading to the recruitment of the ARP2/3 complex and actin, thereby initiating FLS formation.

Rachel David

Nature Reviews Molecular Cell Biology 11, 756-757, (November 2010) [doi:10.1038/nrm2991]