Abstract
SummaryFilopodia assemble unique integrin-adhesion complexes to sense the extracellular matrix. However, the mechanisms of integrin regulation in filopodia are poorly defined. Here, we report that active integrins accumulate at the tip of myosin-X (MYO10)-positive filopodia, while inactive integrins are uniformly distributed. We identify talin and MYO10 as the principal integrin activators in filopodia. In addition, deletion of MYO10’s FERM domain, or mutation of its β1-integrin-binding residues, reveals MYO10 as facilitating integrin activation, but not transport, in filopodia. However, MYO10’s isolated FERM domain alone cannot activate integrins, potentially because of binding to both integrin tails. Finally, because a chimera construct generated by swapping MYO10-FERM by talin-FERM enables integrin activation in filopodia, our data indicate that an integrin-binding FERM domain coupled to a myosin motor is a core requirement for integrin activation in filopodia. Therefore, we propose a two-step integrin activation model in filopodia: receptor tethering by MYO10 followed by talin-mediated integrin activation.
Highlights
Filopodia are actin-rich ‘‘antenna-like’’ protrusions that are responsible for constantly probing the cellular environment composed of neighboring cells and the extracellular matrix (ECM)
Integrin activation occurs at filopodia tips independently of cellular forces and focal adhesions We and others have previously described the formation of integrin-mediated ECM-sensing adhesions at filopodia tips (Shibue et al, 2012; Jacquemet et al, 2019; Lagarrigue et al, 2015; Alieva et al, 2019; Gallop, 2020)
To gain further insights into how integrin activity is regulated in MYO10 filopodia, we first assessed the spatial distribution of high-affinity and unoccupied b1-integrin in U2-OS cells overexpressing fluorescently tagged MYO10 using structured illumination microscopy (SIM) (Figures 1A–1C) and scanning electron microscopy (SEM) (Figure 1D)
Summary
Filopodia are actin-rich ‘‘antenna-like’’ protrusions that are responsible for constantly probing the cellular environment composed of neighboring cells and the extracellular matrix (ECM). Filopodia have a unique cytoskeleton composed of tightly packed parallel actin filaments with barbed ends oriented toward the filopodium tip (Mattila and Lappalainen, 2008). This organization allows molecular motors, such as unconventional myosin-X (MYO10), to move toward and accumulate at the tips (at approximately 600 nm/s) (Kerber et al, 2009). By doing so, these molecular motors are thought to transport various proteins, including integrins, along actin filaments to the tips of filopodia (Jacquemet et al, 2015; Arjonen et al, 2014; Berg and Cheney, 2002; Hirano et al, 2011; Zhang et al, 2004). The subsequent maturation of these filopodia adhesions into nascent and focal adhesions can promote directional cell migration (Hu et al, 2014; Jacquemet et al, 2016, 2019)
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