Optogenetic STING clustering system through nanobody-fused photoreceptor for innate immune regulation

Stimulator of interferon gene (STING) serves as a key mediator for regulating innate immune response. Despite the dynamic process of STING activation, the role of STING clustering in the STING-mediated immune response remains unclear due to the lack of a suitable tool. We developed an innovative optogenetic STING clustering system, OptoSTING, that employs a nanobody-fused photoreceptor-driven technique to achieve light-responsive STING clustering. By optimizing the protein configuration, we identified an optimal OptoSTING system that induced efficient, robust, and reversible clustering of STING upon blue-light illumination. We confirmed that light-induced STING clustering required ER exit to trigger the stimulation of type I interferon response because only cytosolic fragment of OptoSTING (cyt-OptoSTING) enabled to initiate immune response, not full-length OptoSTING. The precise and temporally controlled clustering by cyt-OptoSTING revealed that STING clustering facilitated the STING signaling pathway through puncta formation of IRF3 as downstream effector protein.