Stochastic combinations of actin regulatory proteins are sufficient to drive filopodia formation

Dobramysl, Ulrich; Jarsch, Iris K.; Inoue, Yoshiko; Shimo, Hanae; Richier, Benjamin; Gadsby, Jonathan R; Mason, Julia; Szalapak, Alicja M.; Ioannou, Pantelis Savvas; Correia, Guilherme Pereira; Walrant, Astrid; Butler, Richard J.; Hannezo, Édouard; Simons, Benjamin D.; Gallop, Jennifer L.

doi:10.1083/jcb.202003052

Cited by 30 publications

(43 citation statements)

References 65 publications

(64 reference statements)

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“…In cells, filopodia formation involves multiple protein complexes ( 66 ). Our in vitro work provides direct evidence that actin filament assembly facilitates IRSp53-based protrusion generation.…”

Section: Discussionmentioning

confidence: 99%

Activated I-BAR IRSp53 clustering controls the formation of VASP-actin-based membrane protrusions

Tsai

Henderson

Jarin

et al. 2022

Preprint

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Filopodia are actin-rich membrane protrusions essential for cell morphogenesis, motility, and cancer invasion. How cells control filopodia initiation on the plasma membrane remains elusive. We performed experiments in cellulo, in vitro and in silico to unravel the mechanism of filopodia initiation driven by the membrane curvature sensor IRSp53. We showed that full-length IRSp53 self-assembles into clusters on membranes depending on PIP2. Using well-controlled in vitro reconstitution systems, we demonstrated that IRSp53 clusters recruit the actin polymerase VASP to assemble actin filaments locally on membranes, leading to the generation of actin-filled membrane protrusions reminiscent of filopodia. By pulling membrane nanotubes from live cells, we observed that IRSp53 can only be enriched and trigger actin assembly in nanotubes at highly dynamic membrane regions. Our work supports a regulation mechanism of IRSp53 in its attributes of curvature sensation and partner recruitment to ensure a precise spatial-temporal control of filopodia initiation.

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Section: Discussionmentioning

confidence: 99%

Activated I-BAR IRSp53 clustering controls the formation of VASP-actin-based membrane protrusions

Tsai

Henderson

Jarin

et al. 2022

Preprint

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“…Mass spectrometry analysis of WRC immunoprecipitation shows that both proteins are pulled down abundantly by WAVE2ΔVCA, but not by WAVE2ΔPVCA (data not shown). If any of them is the key transducer responsible for lamellipodia formation or if the WRC acts as a general hub that locally concentrates and promotes the interaction of a number of molecules needed for actin polymerization, like observed in clathrin structures 39 and filopodia 40 , is still unknown. Lastly we cannot exclude the involvement of some other processes that do not depend on binding partners, such as direct membrane deformation at the leading edge and/or phase separation 41 .…”

Section: Discussionmentioning

confidence: 99%

The Scar/WAVE complex drives normal actin protrusions without the Arp2/3 complex, but proline-rich domains are required

Buracco

Singh

Claydon

et al. 2022

Preprint

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Cell migration requires the constant modification of cellular shape by reorganization of the actin cytoskeleton. The pentameric Scar/WAVE regulatory complex (WRC) is the main catalyst of pseudopod and lamellipodium formation. Its actin nucleation activity has been attributed to its ability to combine monomeric actin and Arp2/3 complex through the VCA domain of Scar/WAVE, while other regions of the complex are typically thought to mediate spatial and temporal regulation and have no direct role in actin polymerization.Here we show that the Scar/WAVE with its VCA domain deleted can still induce the formation of morphologically normal actin protrusions. Equivalent results are seen in B16-F1 mouse melanoma cells and Dictyostelium discoideum cells. This actin polymerization occurs independently of the Arp2/3 complex, whose recruitment to the leading edge is greatly reduced by the loss of the VCA domain. We also expressed Scar/WAVE with VCA and polyproline domains both deleted. In Dictyostelium cells, these were only active if WASP (which contains its own proline-rich domain) was available. Similarly, in B16-F1 cells both Abi and WAVE proline-rich domains needed to be deleted before the function of the WRC was lost. Thus we conclude that proline-rich domains play a central role in actin nucleation.Our data demonstrate a new actin nucleation mechanism of the WRC that is independent of its VCA domain and the Arp2/3 complex. We also show that proline-rich domains are more fundamental than has been thought. Together, these findings suggest a new mechanism for WRC action.

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“…Finally, we found a tandem role of IRSp53 and BIN1, in filopodia-like structure formation. The association of IRSp53 with antagonistic BAR domain proteins was reported to participate in filopodia formation (Galic et al, 2014; Dobramysl et al, 2021) or assist endocytosis (Bisi et al, 2020) in different cell types and organisms. Why proteins with opposite curvature domains are required to enable distinct remodeling events at the plasma membrane is yet an open question but suggests that the spatio-temporal regulation of these events might be more complex than previously anticipated.…”

Section: Discussionmentioning

confidence: 99%

BIN1 regulates actin-membrane interactions during IRSp53-dependent filopodia formation

Picas

Comunale

André-Arpin

et al. 2022

Preprint

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Amphiphysin 2 (BIN1) is a membrane and actin remodeling protein mutated both in congenital and adult centronuclear myopathies. The BIN1 muscle-specific isoform finely tunes muscle regeneration in adulthood and regulates myoblast fusion. However, the underlying molecular mechanisms are unknown. Here, we report that BIN1 is required for myoblast fusion and participates in the formation of filopodia-like structures at myoblasts intercellular junctions. BIN1 bundles actin in vitro and regulates the membrane-to-cortex attachment, two key processes required for myoblast fusion. We identified ezrin as a new BIN1 partner and showed that BIN1 promotes ezrin association to PI(4,5)P2 at the membrane cortex. Our results establish BIN1 and ezrin as key players at the early stages of myoblast fusion for the formation of long-lived filopodia-like structures.

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Stochastic combinations of actin regulatory proteins are sufficient to drive filopodia formation

Cited by 30 publications

References 65 publications

Activated I-BAR IRSp53 clustering controls the formation of VASP-actin-based membrane protrusions

Activated I-BAR IRSp53 clustering controls the formation of VASP-actin-based membrane protrusions

The Scar/WAVE complex drives normal actin protrusions without the Arp2/3 complex, but proline-rich domains are required

BIN1 regulates actin-membrane interactions during IRSp53-dependent filopodia formation

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