ADF/Cofilin Accelerates Actin Dynamics by Severing Filaments and Promoting Their Depolymerization at Both Ends

Abstract

SummaryActin-depolymerizing factor (ADF)/cofilins contribute to cytoskeletal dynamics by promoting rapid actin filament disassembly. In the classical view, ADF/cofilin sever filaments, and capping proteins block filament barbed ends whereas pointed ends depolymerize, at a rate that is still debated. Here, by monitoring the activity of the three mammalian ADF/cofilin isoforms on individual skeletal muscle and cytoplasmic actin filaments, we directly quantify the reactions underpinning filament severing and depolymerization from both ends. We find that, in the absence of monomeric actin, soluble ADF/cofilin can associate with bare filament barbed ends to accelerate their depolymerization. Compared to bare filaments, ADF/cofilin-saturated filaments depolymerize faster from their pointed ends and slower from their barbed ends, resulting in similar depolymerization rates at both ends. This effect is isoform specific because depolymerization is faster for ADF- than for cofilin-saturated filaments. We also show that, unexpectedly, ADF/cofilin-saturated filaments qualitatively differ from bare filaments: their barbed ends are very difficult to cap or elongate, and consequently undergo depolymerization even in the presence of capping protein and actin monomers. Such depolymerizing ADF/cofilin-decorated barbed ends are produced during 17% of severing events. They are also the dominant fate of filament barbed ends in the presence of capping protein, because capping allows growing ADF/cofilin domains to reach the barbed ends, thereby promoting their uncapping and subsequent depolymerization. Our experiments thus reveal how ADF/cofilin, together with capping protein, control the dynamics of actin filament barbed and pointed ends. Strikingly, our results propose that significant barbed-end depolymerization may take place in cells.

The actin filament twist changes abruptly at boundaries between bare and cofilin-decorated segments

“…For example, barbed-end boundaries may be more readily identified than pointed-end ones. Alternatively, this behavior could arise if severing occurred preferentially at the pointed-end side of the cluster, as reported (32,33). A third possible explanation would be a mechanism in which cofilin clusters grew asymmetrically and more rapidly in the pointed-end direction (8), such that clusters extended to the filament pointed end.…”

Twinfilin uncaps filament barbed ends to promote turnover of lamellipodial actin networks

“…The discrepancy of data between biochemical and cellular experiments suggests that additional factors might regulate uncapping of CP-capped barbed ends. Both CARMIL and ADF/cofilin can enhance uncapping of filament barbed ends in vitro 26,36,39 . However, to our knowledge the evidence of filament uncapping in cells is missing.Twinfilin is a conserved ADF/cofilin-like protein that is composed of two actin-binding ADF-H (actin depolymerization factor homology) domains, followed by CPI-motif containing C-terminal tail 40 , which binds CP with high affinity 41-43 and interacts with membrane phosphoinositides 42 .…”

Microfluidics‐Assisted TIRF Imaging to Study Single Actin Filament Dynamics

“…This ensures that filament pointed-ends are free to be exposed to a solution containing protein of interest. This method has recently been used to study the effect of ADF on filament pointed ends Wioland et al, 2017).…”