Site-specific cation release drives actin filament severing by vertebrate cofilin

Significance Cofilin is an essential actin regulatory protein that severs filaments, which accelerates network remodeling by increasing the concentration of filament ends available for elongation and subunit exchange. The molecular basis of how cofilin binding interactions fragment filaments, which have stiffness comparable to commercial laboratory plastics, remains a central and unresolved mystery of cellular actin cytoskeleton reorganization. In this study we demonstrate that actin filament severing by vertebrate cofilin is driven by the linked dissociation of a single, site-specific cation that controls filament structure and mechanical properties, and that filament severing is an essential function of cofilin in cells. This work establishes that discrete interactions with cations serve a central regulatory function in mediating actin filament fragmentation by certain classes of severing proteins. Actin polymerization powers the directed motility of eukaryotic cells. Sustained motility requires rapid filament turnover and subunit recycling. The essential regulatory protein cofilin accelerates network remodeling by severing actin filaments and increasing the concentration of ends available for elongation and subunit exchange. Although cofilin effects on actin filament assembly dynamics have been extensively studied, the molecular mechanism of cofilin-induced filament severing is not understood. Here we demonstrate that actin filament severing by vertebrate cofilin is driven by the linked dissociation of a single cation that controls filament structure and mechanical properties. Vertebrate cofilin only weakly severs Saccharomyces cerevisiae actin filaments lacking this “stiffness cation” unless a stiffness cation-binding site is engineered into the actin molecule. Moreover, vertebrate cofilin rescues the viability of a S . cerevisiae cofilin deletion mutant only when the stiffness cation site is simultaneously introduced into actin, demonstrating that filament severing is the essential function of cofilin in cells. This work reveals that site-specific interactions with cations serve a key regulatory function in actin filament fragmentation and dynamics.