Advances in mechanistic understanding of integrin-mediated adhesion highlight the importance of precise control of ligand presentation in directing cell migration. Top-down nanopatterning limited the spatial presentation to sub-micron placing restrictions on both fundamental study and biomedical applications. To break the constraint, here we propose a bottom-up nanofabrication strategy to enhance the spatial resolution to the molecular level using simple formulation that is applicable as treatment agent. Via self-assembly and co-assembly, precise control of ligand presentation is succeeded by varying the proportions of assembling ligand and nonfunctional peptide. Assembled nanofilaments fulfill multi-functions exerting enhancement to suppression effect on cell migration with tunable amplitudes. Self-assembled nanofilaments possessing by far the highest ligand density prevent integrin/actin disassembly at cell rear, which expands the perspective of ligand-density-dependent-modulation, revealing valuable inputs to therapeutic innovations in tumor metastasis.