F-actin binding and bundling are crucial to a plethora of cell processes, including morphogenesis, migration, adhesion and many others. SWAP-70 was recently described as an in vitro F-actin-binding and -bundling protein. Fluorescence cross-correlation spectroscopy measurements with purified recombinant SWAP-70 confirmed that it forms stable oligomers that facilitate F-actin bundling. However, it remained unclear how SWAP-70 oligomerization and F-actin binding are controlled in living cells. We addressed this by biophysical approaches, including seFRET, FACS-FRET and FLIM-FRET. PIP3-mediated association with the cytoplasmic membrane and non-phosphorylated Y426 are required for SWAP-70 to dimerize and to bind F-actin. The dimerization region was identified near the C terminus where R546 is required for dimerization and, thus, F-actin bundling. The in vitro and in vivo data presented here reveal the functional relationship between the cytoplasm-to-membrane translocation and dimerization of SWAP-70, and F-actin binding and bundling, and demonstrate that SWAP-70 is a finely controlled modulator of membrane-proximal F-actin dynamics.This article has an associated First Person interview with the first author of the paper.