Single-molecule imaging of a three-component ordered actin disassembly mechanism

Abstract

Summary The mechanisms by which cells destabilize and rapidly disassemble filamentous actin networks have remained elusive; however, Coronin, Cofilin, and AIP1 have been implicated in this process. Here, using multi-wavelength single molecule fluorescence imaging, we show that mammalian Cor1B, Cof1, and AIP1 work in concert through a temporally ordered pathway to induce highly efficient severing and disassembly of actin filaments. Cor1B binds to filaments first, and dramatically accelerates the subsequent binding of Cof1, leading to heavily decorated, stabilized filaments. Cof1 in turn recruits AIP1, which rapidly triggers severing and remains bound to the newly-generated barbed ends. New growth at barbed ends generated by severing was blocked specifically in the presence of all three proteins. This activity enabled us to reconstitute and directly visualize single actin filaments being rapidly polymerized by formins at their barbed ends while simultanteously being stochastically severed and capped along their lengths, and disassembled from their pointed ends.

Actin-interacting Protein 1 Promotes Disassembly of Actin-depolymerizing Factor/Cofilin-bound Actin Filaments in a pH-dependent Manner

“…In addition, AIP1 remains bound transiently to severed filaments (Fig. 7, top right) and contributes to capping when coronin is present (20), but the effect of pH on this process remains to be investigated.…”

TIRF microscopy analysis of human Cof1, Cof2, and ADF effects on actin filament severing and turnover

“…ADF/Cofilins bind to G-actin between subdomains I and III, and strongly inhibit nucleotide exchange on actin monomers [21]. Real time imaging using multi-wavelength Total Internal Reflection Fluorescence (TIRF) microscopy has shown that yeast Cofilin and human Cof1 bind actin filaments in a cooperative manner, and induce severing at the boundaries between bare and decorated segments [22–24]. ADF/Cofilin binding also alters the structure and mechanical properties of actin filaments [21, 22, 25–29], and induces the release of cations to ‘soften’ F-actin, creating mechanical discontinuities that induce fragmentation [27].…”

Class III myosins shape the auditory hair bundles by limiting microvilli and stereocilia growth

“…8 B, middle and right). We extended our analysis to two additional potential binding partners of myosin IIIa in cochlear hair bundles: coronin-1a (found with the PPI Finder tool; He et al, 2009), an actin-binding protein that promotes the cofilinmediated severing of actin filaments (Jansen et al, 2015), and espin-like, an espin paralogue similar in structure to espin-1 (Shin et al, 2013;Zheng et al, 2014). Both coronin-1 and espin-like had similar locations at the tips of cochlear stereocilia in the presence and in the absence of class III myosins (Fig.…”