Cryo-EM reveals the transition of Arp2/3 complex from inactive to nucleation-competent state

Shaaban, Mohammed; Chowdhury, Saikat; Nolen, Brad J.

doi:10.1038/s41594-020-0481-x

Cited by 52 publications

(115 citation statements)

References 56 publications

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“…S4). In this regard our structure of the activated Arp2/3 complex in branch junctions agrees with a recently published in vitro structure of the NPF Dip1-activated S. pombe Arp2/3 complex that generates unbranched actin filaments [29] (Figure S4, Figure S5a,b). Also, in both structures of the activated Arp2/3 complex, ArpC2 and ArpC4 form a stable core and define the centre of rotation and translation of two subcomplexes consisting of ArpC2, Arp3, ArpC3 and ArpC4, ArpC5, ArpC1, Arp2 that move against each other to form the short pitch conformation (Movies S4 and S5).…”

Section: Resultssupporting

confidence: 91%

“…In the inactive complex and the recent in vitro structure of the Dip1-activated S. pombe Arp2/3 complex, the ArpC5 N-terminus has been shown to bind the side of subdomain 3 of Arp2 [7,25,29]. In our structure, no density for this interaction can be observed (Figure 4b).…”

Section: Resultsmentioning

confidence: 44%

“…Arp2 and Arp3 are the only two subunits forming interfaces with the daughter filament (Fig. 3, a and e), confirming that the stability of connecting the activated Arp2/3 complex to the newly growing daughter filament is solely accomplished by actin-like interfaces [9,29].…”

Section: Resultsmentioning

confidence: 70%

“…A recent single-particle cryo-EM study of the inactive Arp2/3 complex bound to two WCA peptides showed that their C-helices interacted with structurally similar regions in both Arp2-ArpC1 and Arp3 [4]. Previously it was also suggested that for Arp2/3 complex activation, NPF binding to the Arp3 binding site requires the Arp3 C-terminal tail to be displaced from its position in the inactive conformation of the Arp2/3 complex, releasing its autoinhibitory function [4,34] and also eventually allowing the D-loop of the first daughter filament actin monomer to bind the Arp3 barbed end [29]. This agrees with our structure where no density corresponding to the C-terminal tail in its inactive conformation is present.…”

Section: Resultsmentioning

confidence: 99%

“…The differential regulatory activity of ArpC5 isoforms could be caused by their varying binding strength of their N-terminus to the active conformation of the Arp2/3 complex, playing a regulatory role in interactions with cortactin, specifically selecting for Arp2/3 complexes of ArpC1/ArpC5 subunit isoform composition [38]. Additionally, activation of the Arp2/3 complex is associated with ATP binding, the binding of the C-helix of the second NPF to Arp3, and the release of the Arp3 C-terminal tail from its binding site in the inactive complex [4,29,34]. The C-terminal tail relocates to form contacts with ArpC4 and Arp2, providing further stabilization to the active conformation in the branch junction.…”

Section: Discussionmentioning

confidence: 99%

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Novel cryo-electron tomography structure of Arp2/3 complex in cells reveals mechanisms of branch formation

Fäßler

Dimchev

Hodirnau

et al. 2020

Preprint

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The actin-related protein (Arp)2/3 complex nucleates branched actin filament networks pivotal for cell migration, endocytosis and pathogen infection. Its activation is tightly regulated and involves complex structural rearrangements and actin filament binding, which are yet to be understood. Here, we report a 9.0 Å resolution structure of the actin filament Arp2/3 complex branch junction in cells using cryo-electron tomography and subtomogram averaging. This allows us to generate an accurate model of the active Arp2/3 complex in the branch junction and its interaction with actin filaments. Our structure indicates a central role for the ArpC3 subunit in stabilizing the active conformation and suggests that in the branch junction relocation of the ArpC5 N-terminus and the C-terminal tail of Arp3 is important to fix Arp2 and Arp3 in an actin dimer-like conformation. Notably, our model of the branch junction in cells significantly differs from the previous in vitro branch junction model.

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Section: Resultssupporting

confidence: 91%

Section: Resultsmentioning

confidence: 44%

Section: Resultsmentioning

confidence: 70%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 3 more Smart Citations

Novel cryo-electron tomography structure of Arp2/3 complex in cells reveals mechanisms of branch formation

Fäßler

Dimchev

Hodirnau

et al. 2020

Preprint

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Engineering (Bio)Materials through Shrinkage and Expansion

Wang

Tang

et al. 2021

Adv Healthcare Materials

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Although various (bio)fabrication technologies have achieved revolutionary progress in the past decades, engineered constructs still fall short of expectations owing to their inability to attain precisely designable functions. Shrinkable and expandable (bio)materials feature unique characteristics leading to size-/shape-shifting and thus have exhibited a strong potential to equip current engineering technologies with promoted capacities toward applications in biomedicine. In this progress report, the advances of size-/shape-shifting (bio)materials enabled by various stimuli, are evaluated; furthermore, representative biomedical applications associated with size-/shape-shifting (bio)materials are also exemplified. Toward the future, the combination of size-/shape-shifting (bio)materials and 3D/4D fabrication technologies presents a wide range of possibilities for further development of intricate functional architectures.

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Septin and actin contributions to endothelial cell–cell junctions and monolayer integrity

et al. 2022

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Septins in endothelial cells (ECs) have important roles supporting the integrity of the endothelial monolayer. Cell–cell junctions in EC monolayers are highly dynamic, with continuous retractions and protrusions. Depletion of septins in ECs leads to disruption of cell–cell junctions, which are composed of VE‐cadherin and other junctional proteins. In EC monolayers, septins are concentrated at the plasma membrane at sites of cell–cell contact, in curved‐ and scallop‐shaped patterns. These membrane‐associated septin accumulations are located in regions of positive membrane curvature, and those regions are often associated with and immediately adjacent to actin‐rich protrusions with negative membrane curvature. EC septins associate directly with plasma membrane lipids, based on findings with site‐specific mutations of septins in ECs, which is consistent with biochemical and cell biological studies in other systems. Loss of septins leads to disruption of the EC monolayer, and gaps form between cells. The number and breadth of cell–cell contacts and junctions decreases, and the number and frequency of retractions, ruffles, and protrusions at cell edges also decreases. In addition, loss of septins leads to decreased amounts of F‐actin at the cortical membrane, along with increased amounts of F‐actin in stress fibers of the cytoplasm. Endothelial monolayer disruption from loss of septins is also associated with decreased transendothelial electric resistance (TEER) and increased levels of transendothelial migration (TEM) by immune and cancer cells, owing to the gaps in the monolayer. A current working model is that assembly of septin filaments at regions of positive membrane curvature contributes to a mechanical footing or base for actin‐based protrusive forces generated at adjoining regions of the membrane. Specific molecular interactions between the septin and actin components of the cytoskeleton may also be important contributors. Regulators of actin assembly may promote and support the assembly of septin filaments at the membrane, as part of a molecular feedback loop between the assembly of septin and actin filaments.

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Cryo-EM reveals the transition of Arp2/3 complex from inactive to nucleation-competent state

Cited by 52 publications

References 56 publications

Novel cryo-electron tomography structure of Arp2/3 complex in cells reveals mechanisms of branch formation

Novel cryo-electron tomography structure of Arp2/3 complex in cells reveals mechanisms of branch formation

Engineering (Bio)Materials through Shrinkage and Expansion

Septin and actin contributions to endothelial cell–cell junctions and monolayer integrity

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