The vasodilator-stimulated phosphoprotein (VASP) functions as a cellular regulator of actin dynamics. VASP may initialise actin polymerisation, suggesting a direct interaction with monomeric actin. The present study demonstrates that VASP directly binds to actin monomers and that complex formation depends on a conserved four amino acid motif in the EVH2 domain. Point mutations within this motif drastically weaken VASP/G-actin interactions, thereby abolishing any actin-nucleating activity of VASP. Additionally, actin nucleation was found to depend on VASP oligomerisation since VASP monomers fail to induce the formation of actin filaments. Phosphorylation negatively affects VASP/G-actin interactions preventing VASP-induced actin filament formation.