Li, Q.; Geng, X.; Yan, H.; Li, Z.; Shi, M.; Zhu, Z.; Niu, T.; Zhao, C.; Shu, K.; Gao, Y.; Feng, H.; Liu, S.; Jiang, Q.; Bu, P.; Li, D.; Gao, P.

The innate immune sensor AIM2 detects cytosolic DNA and initiates inflammatory responses, yet its activation mechanism remains incompletely understood. Here, we show that AIM2 undergoes liquid-liquid phase separation upon DNA binding, forming dynamic condensates both in vitro and in cells. These condensates serve as platforms for inflammasome and PANoptosome assembly, promoting immune activation across multiple pathways. Direct structural determination from condensates reveals the assembly of active-form ASC filaments. Mechanistically, liquid-phase condensation is governed by multivalent interactions involving different AIM2 domains, including previously uncharacterized regions and species-specific elements. In vitro and in vivo assays show that mutants specifically disrupting condensation impair immune complex assembly, cell death initiation, antimicrobial defense, and intestinal homeostasis. Moreover, AIM2-DNA condensates function as regulatory hubs targeted by host- and pathogen-derived factors to balance immune homeostasis or facilitate immune evasion. These findings establish liquid-phase condensation as a fundamental mechanism of AIM2 activation and a potential therapeutic target.