Spatiotemporal control of inflammatory lytic cell death through optogenetic induction of RIPK3 oligomerization

Necroptosis is a programmed lytic cell death involving active cytokine production and plasma membrane rupture through distinct signaling cascades. However, it remains challenging to delineate this inflammatory cell death pathway at specific signaling nodes with spatiotemporal accuracy. To address this challenge, we developed an optogenetic system, termed Light-activatable Receptor-Interacting Protein Kinase 3 or La-RIPK3, to enable ligand-free, optical induction of RIPK3 oligomerization. La-RIPK3 activation dissects RIPK3-centric lytic cell death through the induction of RIPK3-containing necrosome, which mediates cytokine production and plasma membrane rupture. Bulk RNA-Seq analysis reveals that RIPK3 oligomerization results in partially overlapped gene expression compared to pharmacological induction of necroptosis. However, La-RIPK3 activates a group of genes likely regulated by RIPK3 kinase-independent processes. Using patterned light stimulation delivered by a spatial light modulator, we demonstrate precise spatiotemporal control of necroptosis in La-RIPK3-transduced HT-29 cells. Optogenetic control of proinflammatory lytic cell death could lead to the development of innovative experimental strategies to finetune the immune landscape for disease intervention.