Journal Article

Force and phosphate release from Arp2/3 complex promote dissociation of actin filament branches

Cao, Wenxiang, Taylor, Edwin W., Pollard, Thomas D., Bibeau, Jeffrey, Pandit, Nandan G., Johnson-Chavarria, Eric M., De La Cruz, Enrique M.

2020

Overview

Networks of branched actin filaments formed by Arp2/3 complex generate and experience mechanical forces during essential cellular functions, including cell motility and endocytosis. External forces regulate the assembly and architecture of branched actin networks both in vitro and in cells. Considerably less is known about how mechanical forces influence the disassembly of actin filament networks, specifically, the dissociation of branches. We used microfluidics to apply force to branches formed from purified muscle actin and fission yeast Arp2/3 complex and observed debranching events in real time with total internal reflection fluorescence microscopy. Low forces in the range of 0 pN to 2 pN on branches accelerated their dissociation from mother filaments more than two orders of magnitude, from hours to < 1 min. Neither force on the mother filament nor thermal fluctuations in mother filament shape influenced debranching. Arp2/3 complex at branch junctions adopts two distinct mechanical states with different sensitivities to force, which we name “young/strong” and “old/weak.” The “young/strong” state 1 has adenosine 5′- diphosphate (ADP)−Pi bound to Arp2/3 complex. Phosphate release converts Arp2/3 complex into the “old/weak” state 2 with bound ADP, which is 20 times more sensitive to force than state 1. Branches with ADP−Arp2/3 complex are more sensitive to debranching by fission yeast GMF (glia maturation factor) than branches with ADP−Pi−Arp2/3 complex. These findings suggest that aging of branch junctions by phosphate release from Arp2/3 complex and mechanical forces contribute to disassembling “old” actin filament branches in cells.

• Publisher: National Academy of Sciences
• Subject: Actin / Actin Cytoskeleton - metabolism / Actin-Related Protein 2-3 Complex - metabolism / Actins - metabolism / Adenosine / Adenosine diphosphate / Adenosine Diphosphate - metabolism / Aging / Animals / Biological Sciences / Biophysics and Computational Biology / Dismantling / Endocytosis / Filaments / Fission / Fluorescence / Fluorescence microscopy / Glia maturation factor / Glia Maturation Factor - metabolism / Microfluidics / Microscopy, Fluorescence / Models, Biological / Muscles / Networks / Phosphates - metabolism / PNAS Plus / Protein Binding / Rabbits / Schizosaccharomyces - metabolism / Stress, Mechanical / Yeast