Arp2/3 complex interactions and actin network turnover in lamellipodia

Abstract

Cell migration is initiated by lamellipodia-membraneenclosed sheets of cytoplasm containing densely packed actin filament networks. Although the molecular details of network turnover remain obscure, recent work points towards key roles in filament nucleation for Arp2/3 complex and its activator WAVE complex. Here, we combine fluorescence recovery after photobleaching (FRAP) of different lamellipodial components with a new method of data analysis to shed light on the dynamics of actin assembly/disassembly. We show that Arp2/3 complex is incorporated into the network exclusively at the lamellipodium tip, like actin, at sites coincident with WAVE complex accumulation. Capping protein likewise showed a turnover similar to actin and Arp2/3 complex, but was confined to the tip. In contrast, cortactin-another prominent Arp2/3 complex regulator-and ADF/cofilin-previously implicated in driving both filament nucleation and disassembly-were rapidly exchanged throughout the lamellipodium. These results suggest that Arp2/3-and WAVE complex-driven actin filament nucleation at the lamellipodium tip is uncoupled from the activities of both cortactin and cofilin. Network turnover is additionally regulated by the spatially segregated activities of capping protein at the tip and cofilin throughout the mesh.

Glia Maturation Factor (GMF) Interacts with Arp2/3 Complex in a Nucleotide State-dependent Manner

“…We have demonstrated for the first time that GMF␥ has a clear preference for ADP-bound versus ATP-bound Arp2/3 complex, which could have important physiological implications. Indeed, in vitro branches formed by Arp2/3 complex persist for hundreds of seconds (47), whereas branch turnover in cells occurs within a few seconds (48), implying the existence of cellular factors that accelerate debranching. One mechanism for debranching has been proposed to involve cofilin-mediated disassembly of the mother filament (49).…”

14-3-3σ stabilizes a complex of soluble actin and intermediate filament to enable breast tumor invasion

“…To test this, we performed fluorescence recovery after photobleaching (FRAP) experiments using an F-actin live-cell probe (30) and observed a ∼twofold increase in the rate of F-actin fluorescence recovery in sh-14-3-3σ cells (Fig. 2C), consistent with an increase in actin-polymerization kinetics rather than a suppression of filament turnover (31). By performing Western blot on whole cell lysates, we observed no difference in the total actin levels between cells (Fig.1B and Fig.…”

The β‐thymosins: Intracellular and extracellular activities of a versatile actin binding protein family

“…These regions are thin, veil-like extensions of the plasma mem- brane containing a dense branched network of actively cycling actin filaments. Their barbed-ends are directed towards the cell periphery and their pointed-ends to the cytoplasm [for review see Pollard and Borisy, 2003;Lai et al, 2008]. Here Tb4 might be involved in securing a high actin monomer concentration essential for rapid formation of new filaments (see above) by forward diffusing as actin:Tb4 complex from the rear of the lamellipodium.…”