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bioRxiv · 10.1101/2022.05.06.490861

Nucleation and stability of branched versus linear Arp2/3-generated actin filaments

Cao, L.·Ghasemi, F.·Way, M.·Jegou, A.·Romet-Lemonne, G.
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Abstract

Activation of the Arp2/3 complex by VCA-domain-bearing NPFs results in the formation of daughter actin filaments branching off the sides of pre-existing mother filaments. Alternatively, when stimulated by SPIN90, Arp2/3 directly nucleates linear actin filaments. Uncovering the similarities and differences of these two activation mechanisms is fundamental to understanding the regulation and function of Arp2/3. Analysis of individual filaments reveals that the catalytic VCA domain of WASP, N-WASP and WASH, accelerate the Arp2/3-mediated nucleation of linear filaments by SPIN90, in addition to their known branch-promoting activity. Unexpectedly, these VCA domains also destabilize existing branches, as well as SPIN90-Arp2/3 at filament pointed ends. Furthermore, cortactin and GMF, which respectively stabilize and destabilize Arp2/3 at branch junctions, have a similar impact on SPIN90-activated Arp2/3. However, unlike branch junctions, SPIN90-Arp2/3 at the pointed end of linear filaments is not destabilized by piconewton forces, and does not become less stable with time. It thus appears that linear and branched Arp2/3-generated filaments respond similarly to regulatory proteins, albeit with quantitative differences, and that they differ greatly in their responses to aging and to mechanical stress. These results indicate that SPIN90- and VCA-activated Arp2/3 complexes adopt similar yet non-identical conformations, and that their turnover in cells may be regulated differently.

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Cao, L., Ghasemi, F., Way, M., Jegou, A., Romet-Lemonne, G.. 2022-05-06. Nucleation and stability of branched versus linear Arp2/3-generated actin filaments. https://doi.org/10.1101/2022.05.06.490861

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