RIPK3 sequentially recruits MLKL and RIPK1 to induce PANoptosis and chemokine production.

Receptor-interacting protein kinase 3 (RIPK3) has emerged as a central player in necroptosis and apoptosis activation in specific scenarios, concurrently modulating inflammatory responses. Here, we reveal that direct activation of RIPK3 concomitantly triggers mixed lineage kinase domain-like (MLKL) phosphorylation, caspase activation, and gasdermin cleavage within individual cells, inducing PANoptotic cell death. This process is orchestrated by the formation of RIPK3-MLKL-RIPK1-FADD-Caspase-8 complexes on progressively polymerized RIPK3 homo-aggregates, achieved through sequential recruitment dictated by the differential affinities of MLKL and Receptor-interacting protein kinase 1 (RIPK1) for distinct oligomeric states of RIPK3. In this process, MLKL- and GSDMD-mediated membrane rupture is respectively inhibited by Caspase-3-dependent cleavage of RIPK3 and GSDMD cleavage, while the pro-necrotic kinase activity of RIPK3 impedes RIPK1 recruitment and attenuates caspase activation. Cross-regulation between pathways results in unique cellular morphology, altered damage-associated molecular patterns (DAMPs) release profiles and distinct chemokine secretion paradigms that differ fundamentally from classical necroptosis, apoptosis and pyroptosis. This work highlights a common mechanism unveiling RIPK3 as a multimolecular platform to modulate and integrate different programmed cell death (PCD) pathways, thus providing a framework for targeting inflammatory cell death in disease.