Genetically-stable engineered optogenetic gene switches modulate spatial cell morphogenesis in two- and three-dimensional tissue cultures.

Recent advances in tissue engineering have been remarkable, yet the precise control of cellular behavior in 2D and 3D cultures remains challenging. One approach to address this limitation is to genomically engineer optogenetic control of cellular processes into tissues using gene switches that can operate with only a few genomic copies. Here, we implement blue and red light-responsive gene switches to engineer genomically stable two- and three-dimensional mammalian tissue models. Notably, we achieve precise control of cell death and morphogen-directed patterning in 2D and 3D tissues by optogenetically regulating cell necroptosis and synthetic WNT3A signaling at high spatiotemporal resolution. This is accomplished using custom-built patterned LED systems, including digital mirrors and photomasks, as well as laser techniques. These advancements demonstrate the capability of precise spatiotemporal modulation in tissue engineering and open up new avenues for developing programmable 3D tissue and organ models, with significant implications for biomedical research and therapeutic applications.