Molecular mechanism of actin filament elongation by formins

• 成果类型: Article
• 署名作者: Oosterheert, Wout; Sanders, Micaela Boiero; Funk, Johanna; Prumbaum, Daniel; Raunser, Stefan; Bieling, Peter
• 署名单位: Max Planck Society; Max Planck Society
• 刊物名称: SCIENCE
• ISSN/ISSBN: 0036-8361
• DOI: 10.1126/science.adn9560
• 发表日期: 2024-04-12
• 关键词: beam-induced motion; fission yeast; barbed-end; structural basis; cryo-em; crystal-structures; capping protein; fh1 domain; profilin; nucleation

摘要

INTRODUCTION: The dynamic turnover of actin filaments drives the morphogenesis and movement of all eukaryotic cells. The ends of actin filaments are key to controlling filament dynamics because they are the only sites where actin subunits are added or lost. Formins are highly conserved actin end-binding proteins. By recruiting actin monomers and moving with the fast-growing ends of filaments, formins act as polymerases that control actin dynamics in many biological processes. The human genome encodes 15 distinct formins, which drive filament growth with different speeds. This variety helps to create different actin networks in cells, each with its own behavior. Mutations in formins result in various neurological, immune, and cardiovascular disorders, which highlights their significance in physiology and disease. RATIONALE: Despite decades of interest in formins, our understanding of their molecular mechanism has been limited. Earlier structural studies revealed that formins adopt a dimeric, ring-like conformation, which sparked speculative and conflicting models on how they might work as actin polymerases. However, the lack of structures of formins bound to their relevant sites of activity has hindered our understanding of how these proteins precisely target, move with, and control the growth speed of actin filaments. In this work, we present high-resolution cryo-electron microscopy (cryo-EM) structures of three distinct formins bound to actin filament ends, which enabled us to resolve their mode of action. RESULTS: The structures revealed that formins encircle the actin filament end as a dimeric ring in a common asymmetric arrangement. Although one half of the ring is stably bound, the other half is loosely associated with the filament end and is free to capture a new subunit. When this new actin subunit arrives, its incorporation onto the filament destabilizes the formin arrangement. As a result, one part of the formin dimer must disengage, move forward, and establish a new binding interface with the incorporated subunit. This undock-and-lock mechanism explains why formins processively translocate with the elongating actin filament end without frequently falling behind. Differences in elongation speeds of distinct formins arise from formin positioning and amino acid variations that affect how easily certain formin-actin interactions are made or broken. Speeds can be tuned by mutating these formin-actin interfaces, which we determined by following individual formins elongating actin filaments in vitro. This establishes a molecular framework for understanding functional differences between diverging formins. Finally, we resolved how formins synergize with the essential actin-binding protein profilin in promoting filament growth. Profilin binds most of the polymerizable actin inside cells, and we visualized how these two core actin regulators collaborate at actin filament ends. This showed that rearrangements in actin during polymerization weaken the interaction with profilin, explaining rapid profilin release from the filament end after subunit incorporation. CONCLUSION: Our study provides insights into the structural mechanisms of formin-mediated actin polymerization, which paves the way for manipulating formin activity and understanding the impact of formin mutations at the mechanistic level. These insights are important for elucidating the role of formins in regulating actin dynamics during essential cellular processes in health and disease. Molecular basis of actin filament elongation by formins. From yeast to humans, formins are a class of actin-binding proteins found in all eukaryotes. They mediate actin filament elongation in key cellular processes. High-resolution cryo-EM structures showed that different formin variants share a common conformation when bound to the barbed end of actin filaments. During elongation, an incoming actin subunit destabilizes the trailing formin, which triggers its translocation. F-actin, actin filament; G-actin, monomeric actin.