Lamellipodin, an Ena/VASP Ligand, Is Implicated in the Regulation of Lamellipodial Dynamics

Abstract

Lamellipodial protrusion is regulated by Ena/VASP proteins. We identified Lamellipodin (Lpd) as an Ena/VASP binding protein. Both proteins colocalize at the tips of lamellipodia and filopodia. Lpd is recruited to EPEC and Vaccinia, pathogens that exploit the actin cytoskeleton for their own motility. Lpd contains a PH domain that binds specifically to PI(3,4)P2, an asymmetrically localized signal in chemotactic cells. Lpd's PH domain can localize to ruffles in PDGF-treated fibroblasts. Lpd overexpression increases lamellipodial protrusion velocity, an effect observed when Ena/VASP proteins are overexpressed or artificially targeted to the plasma membrane. Conversely, knockdown of Lpd expression impairs lamellipodia formation, reduces velocity of residual lamellipodial protrusion, and decreases F-actin content. These phenotypes are more severe than loss of Ena/VASP, suggesting that Lpd regulates other effectors of the actin cytoskeleton in addition to Ena/VASP.

Profilin1 regulates PI(3,4)P 2 and lamellipodin accumulation at the leading edge thus influencing motility of MDA-MB-231 cells

“…Activation of receptor tyrosine kinases (RTKs) such as EGFR (EGF receptor) and PDGFR (PDGF receptor) lead to PI3 kinase (PI3K)-mediated generation of D3-phosphoinositides, including PI(3,4,5)P 3 and PI(3,4)P 2 [the latter mostly through PI(3,4,5)P 3 dephosphorylation by 5′-phosphatases]. Consistent with this, PDGF stimulation has been shown to recruit lamellipodin or even the isolated PH domain of lamellipodin to the leading edge and the tips of dorsal ruffles (21,22). We found that PI3K inhibition by LY294002 dramatically inhibits lamellipodin accumulation at the leading edge in profilin1 knockdown MDA-231 cells (with or without rescue by R88L-profilin1) under PDGF-stimulated condition (Fig.…”

“…Lamellipodin, a phosphoinositide binding protein, was previously shown to play an important role in recruiting Ena/VASP to the leading edge in fibroblasts and melanoma cells (note that lamellipodin targets to the leading edge independently of Ena/ VASP) (21,22). Lamellipodin depletion by siRNA markedly reduced VASP accumulation at the leading edge in profilin1 knockdown MDA-231 cells (Fig.…”

“…Lamellipodin contains a PH (pleckstrin homology) domain that binds PI(3,4)P 2 with much higher affinity than any other tested 3′-phosphorylated phosphoinositides (D3-phosphoinositides) such as PI3P or PI(3,4,5)P 3 (21). According to a recent report (24), PI(3,4)P 2 is generated at the sites of lamellipodin recruitment in cells.…”

Cell biology of embryonic migration

“…Additional factors that are important in the network formation and maintenance are regulators of actin polymerization, such as profilin that binds and targets actin monomers to the barbed ends, therefore preventing self-nucleation and ensuring actin incorporation into the proper network, and ADF/cofilin that severs the filaments at the branch points (debranching) and promotes depolymerization from the pointed ends. In filopodia, anti-capping proteins, such as Ena-VASP and their partners (Bailly et al, 1999;Krause et al, 2004;Lafuente et al, 2004) and bundling proteins, such as fascin (Adams, 2004a;Adams, 2004b), are especially important. A large number of other actin-binding proteins participate in the leading edge actin dynamics, facilitating different stages of actin assembly, disassembly, sequestering, and crosslinking.…”

Vasodilator-stimulated phosphoprotein restricts cell-to-cell spread of Shigella flexneri at the cell periphery

“…S1, available in the online Supplementary Material). The EVH1 domains target Ena/VASP proteins to sites of actin rearrangements via interactions with proteins containing the proline-rich motif (D/E)(F/L/W/Y)PPPPX(D/E)(D/E) (designated FPPPP), including zyxin and vinculin at focal adhesions and lamellipodin at the leading edge (Hoffman et al, 2006;Krause et al, 2004;Niebuhr et al, 1997). EVH1 domains also bind to diaphanous formin (Dia) and mammalian diaphanous formin 2 (mDia2), inhibiting actin nucleation and elongation (Barzik et al, 2014;Bilancia et al, 2014), and recent data suggest that VASP activates WAVE (Wiskott-Aldrich syndrome verprolin homology protein) via EVH1-mediated interactions (Chen et al, 2014;Havrylenko et al, 2015).…”