The actin cytoskeleton is essential for many fundamental biological processes, but tools to directly manipulate actin dynamics are limited to cell-permeable drugs precluding single cell perturbations. Here we describe DeActs, genetically encoded actin-modifying polypeptides, which effectively induce actin disassembly in eukaryotic cells. We demonstrate that DeActs are a universal tool for studying the actin cytoskeleton in single cells in culture, tissues, and multicellular organisms, including various neurodevelopmental model systems.