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Abstract
Cell migration entails networks and bundles of actin filaments termed lamellipodia and microspikes or filopodia, respectively, as well as focal adhesions, all of which recruit Ena/VASP family members hitherto thought to antagonize efficient cell motility. However, we find these proteins to act as positive regulators of migration in different murine cell lines. CRISPR/Cas9-mediated loss of Ena/VASP proteins reduced lamellipodial actin assembly and perturbed lamellipodial architecture, as evidenced by changed network geometry as well as reduction of filament length and number that was accompanied by abnormal Arp2/3 complex and heterodimeric capping protein accumulation. Loss of Ena/VASP function also abolished the formation of microspikes normally embedded in lamellipodia, but not of filopodia capable of emanating without lamellipodia. Ena/VASP-deficiency also impaired integrin-mediated adhesion accompanied by reduced traction forces exerted through these structures. Our data thus uncover novel Ena/VASP functions of these actin polymerases that are fully consistent with their promotion of cell migration.
Highlights
Adhesion and migration are invariably driven by continuous and dynamic actin cytoskeleton remodeling (Blanchoin et al, 2014)
In EVM-KO cells rescued with EGFPtagged Vasodilator-stimulated phosphoprotein (VASP), Ena-VASP-like protein (Evl) or mammalian Protein Enabled Homolog (Mena), the mean square displacement (MSD) values were again markedly increased (Figure 1—figure supplement 2B)
At variance to earlier studies (Bear et al, 2002; Bear et al, 2000; Loureiro et al, 2002), we show that Ena/VASP proteins positively regulate 2D cell migration of distinct mesenchymal cell types, since their consecutive loss in B16-F1 cells results in cumulative, reconstitutable motility phenotypes
Summary
Adhesion and migration are invariably driven by continuous and dynamic actin cytoskeleton remodeling (Blanchoin et al, 2014). Branching by Actin-related protein (Arp) 2/3 complex is activated by the WAVE regulatory complex (WRC) downstream of Rac subfamily GTPase signaling (Eden et al, 2002; Ismail et al, 2009; Molinie and Gautreau, 2018). Knockdown or knockout of essential Arp2/3 complex (Suraneni et al, 2012; Wu et al, 2012) or WRC subunits (Innocenti et al, 2004; Schaks et al, 2018; Steffen et al, 2004) or Rac GTPases (Schaks et al, 2018; Steffen et al, 2013) all abrogated lamellipodium formation.
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