Multiple actin binding domains of Ena/VASP proteins determine actin network stiffening

摘要: Vasodilator-stimulated phosphoprotein (Ena/VASP) is an actin binding protein, important for dynamics in motile cells and developing organisms. Though VASP’s main activity the promotion of barbed end growth, it has F-actin site can form tetramers, so could additionally play a role crosslinking bundling cell. To test this activity, we performed rheology reconstituted networks presence wild-type VASP or mutants lacking ability to tetramerize bind G-actin and/or F-actin. We show that increasing amounts increase network stiffness up certain point, beyond which actually decreases with concentration. The maximum 10-fold higher than pure networks. Confocal microscopy shows forms clustered filament bundles, explaining reduction elasticity at high Removal tetramerization results significantly reduced bundle clustering, indicating flexible tetrameric structure causes clustering. Removing either diminishes effect on elasticity, but does not eliminate it. Mutating together, mutating saturating monomeric actin, eliminates stiffness. propose that, cell, confers only moderate increases linear unlike other crosslinkers, stiffening may be tuned by local concentration actin.