Arp2/3 Complex and Cofilin Modulate Binding of Tropomyosin to Branched Actin Networks

Tropomyosins are coiled-coil proteins that bind actin filaments and regulate multiple cytoskeletal functions, including actin network dynamics near the leading edge of motile cells. Previous work demonstrated that tropomyosins inhibit actin nucleation by the Arp2/3 complex and prevent filament disassembly by cofilin. We find that the Arp2/3 complex and cofilin, in turn, regulate the binding of tropomyosin to actin filaments. Using fluorescence microscopy, we show that tropomyosin (non-muscle Drosophila Tm1A) polymerizes along actin filaments, starting from “nuclei” that appear preferentially on ADP-bound regions of the filament, near the pointed end. Tropomyosin fails to bind dendritic actin networks created in vitro by the Arp2/3 complex, in part because the Arp2/3 complex blocks pointed ends. Cofilin promotes phosphate dissociation and severs filaments, generating new pointed ends and rendering Arp2/3-generated networks competent to bind tropomyosin. Tropomyosin’s attraction to pointed ends reflects a strong preference for conformations localized to that region of the filament and reveals a basic molecular mechanism by which lamellipodial actin networks are insulated from the effects of tropomyosin. •Tropomyosin (TM) binding is biased toward ADP-bound actin, near the pointed end•Arp2/3 blocks TM binding to branched actin networks by blocking the pointed end•Cofilin severs actin filaments, generating free pointed ends competent to bind TM Tropomyosins regulate multiple cytoskeletal functions, including actin network dynamics near the leading edge of motile cells. Hsiao et al. show that Arp2/3 complex and cofilin regulate tropomyosin binding to branched actin filaments, revealing a mechanism by which lamellipodial actin networks are insulated from the effects of tropomyosin.