Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.10.03.322354v1?rss=1 Authors: Bashirzadeh, Y., Redford, S. A., Lorpaiboon, C., Groaz, A., Litschel, T., Schwille, P., Hocky, G. M., Dinner, A., Liu, A. P. Abstract: Robust spatiotemporal organization of cytoskeletal networks is crucial, enabling cellular processes such as cell migration and division. -Actinin and fascin are two actin crosslinking proteins localized to distinct regions of eukaryotes to form actin bundles with optimized spacing for cell contractile machinery and sensory projections, respectively. In vitro reconstitution assays and coarse-grained simulations have shown that these actin bundling proteins segregate into distinct domains with a bundler size-dependent competition-based mechanism, driven by the minimization of F-actin bending energy. However, it is not known how physical confinement imposed by the cell membrane contributes to sorting of actin bundling proteins and the concomitant reorganization of actin networks in intracellular environment. Here, by encapsulating actin, -actinin, and fascin in giant unilamellar vesicles (GUVs), we show that the size of such a spherical boundary determines equilibrated structure of actin networks among three typical structures: single rings, astral structures, and star-like structures. We show that -actinin bundling activity and its tendency for clustering actin is central to the formation of these structures. By analyzing physical features of crosslinked actin networks, we show that spontaneous sorting and domain formation of -actinin and fascin are intimately linked to the resulting structures. We propose that the observed boundary-imposed effect on sorting and structure formation is a general mechanism by which cells can select between different structural dynamical steady states. Copy rights belong to original authors. Visit the link for more info