Nek7 is identified as a critical mediator of NLRP3 activation downstream of potassium efflux. The study demonstrates that Nek7 interacts with NLRP3, promoting its oligomerization and activation. In the absence of Nek7, caspase-1 activation and IL-1β release are abrogated in response to stimuli that activate NLRP3 but not NLRC4 or AIM2 inflammasomes. NLRP3-activating stimuli enhance the NLRP3-Nek7 interaction, which is independent of ASC, caspase-1, or caspase-11. The catalytic activity of Nek7 is dispensable for NLRP3 activation, but the N-terminal region of Nek7 is essential for the interaction with NLRP3. Nek7 is required for NLRP3 inflammasome activation in vivo, as demonstrated by mouse chimeras reconstituted with wild-type, Nek7−/−, or Nlrp3−/− hematopoietic cells. These findings suggest that Nek7 acts as a key regulator of NLRP3 inflammasome assembly and activation, providing a potential therapeutic target for inflammatory diseases linked to NLRP3 inflammasome activation.Nek7 is identified as a critical mediator of NLRP3 activation downstream of potassium efflux. The study demonstrates that Nek7 interacts with NLRP3, promoting its oligomerization and activation. In the absence of Nek7, caspase-1 activation and IL-1β release are abrogated in response to stimuli that activate NLRP3 but not NLRC4 or AIM2 inflammasomes. NLRP3-activating stimuli enhance the NLRP3-Nek7 interaction, which is independent of ASC, caspase-1, or caspase-11. The catalytic activity of Nek7 is dispensable for NLRP3 activation, but the N-terminal region of Nek7 is essential for the interaction with NLRP3. Nek7 is required for NLRP3 inflammasome activation in vivo, as demonstrated by mouse chimeras reconstituted with wild-type, Nek7−/−, or Nlrp3−/− hematopoietic cells. These findings suggest that Nek7 acts as a key regulator of NLRP3 inflammasome assembly and activation, providing a potential therapeutic target for inflammatory diseases linked to NLRP3 inflammasome activation.