Reconstituted systems for studying the architecture and dynamics of actin networks.

Actin, a ubiquitous protein essential for numerous cellular functions, is found in all eukaryotes. Despite extensive research across molecular to organismal scales, fundamental questions persist regarding the regulation of dynamic actin architectures, their interaction with membranes, and their mechanical properties. Characterizing the factors governing these processes presents significant challenges. This review emphasizes the value of simplified, reconstituted systems in addressing these unresolved questions. We particularly highlight the critical importance of macroscopic, network-level reconstitutions for tackling these issues. We first describe the available methodological toolkit for (1) controlling actin polymerization spatiotemporally and (2) confining actin networks within closed environments to examine boundary constraint effects or the impact of limited component availability on network properties. We then review studies employing these reconstituted systems to investigate how actin architecture influences various processes and how dynamic actin structures are established and maintained. Further, we discuss how network-level reconstitutions have enhanced our understanding of actin networks' mechanical properties and their interaction with the lipid membranes. Throughout the review, we discuss future perspectives for each topic and explain how macroscale reconstitutions can provide deeper mechanistic insights into actin-related processes.