Light‐Gated Assembly of Membrane‐Bound DNA Filaments as Synthetic Cell Modules

Precise spatiotemporal control over biomolecular assembly is a central requirement for engineering synthetic cells and programmable biomaterials. Of particular interest are inducible filaments that can confer stability or mechanical transformation to membrane compartments. Here, we introduce a UV‐gated, two‐tile DNA filament system for controlled assembly on lipid membranes. The system combines a membrane‐tethered, UV‐caged Tile A, whose sticky ends are shielded by photocleavable strands, with a freely diffusing complementary Tile B. UV illumination uncages Tile A and initiates filament growth, while UV dose and tile stoichiometry modulate assembly kinetics and network morphology without compromising final filament yield. Localized illumination enables writing of defined architectures, while sequential exchange of Tile B variants enables “molecular woodblock printing” of multiple cargos into predefined regions. On giant unilamellar vesicle (GUV) membranes, UV activation produces filament networks ranging from uniform coatings to ring and cage geometries. These assemblies support selective cargo recruitment and mechanically stabilize vesicles by suppressing membrane undulations. Finally, coupling filament assembly to protein pattern formation guides where filament networks form, providing a route to autonomous spatial regulation. Together, this platform offers on‐demand activation, spatial addressability, and modular functionalization for programmable membrane interfaces and a versatile tool for biofabrication and bottom‐up synthetic biology.