Cofilin-1 is an essential redox sensor for NLRP3 inflammasome activation

Introduction

NLRP3 has a pivotal role in nucleating the inflammasome, a cytoplasmic multiprotein complex that mediates the maturation of the proinflammatory cytokine interleukin-1β (IL-1β) by activating caspase-1. Mutations in the gene encoding NLRP3 cause a spectrum of autoinflammatory disease, the cryopyrin-associated periodic syndromes (CAPS). It has been reported that generation of reactive oxygen species (ROS) is a major NLRP3 inflammasome-activating factor. However, the molecular mechanism by which a change in cellular redox state leads to NLRP3 inflammasome activation has not been elucidated. Here we show that cofilin-1, a redox sensitive actin binding protein, is involved in NLRP3 inflammasome activation.

Objectives

To investigate how ROS activates the NLRP3 inflammasome.

Methods

Cell culture supernatants from bone marrow derived macrophages (BMDMs) of wild-type or NLRP3-KO mice were analyzed by mass spectrometry. Inflammasome activation was analyzed by Western blotting of secreted IL-1β or ASC oligomerization. The interaction of NLRP3 with cofilin-1 was examined by co-immunoprecipitation from BMDMs or transfected cells.

Results

Cofilin-1 is highly secreted along with IL-1β from LPS-primed BMDMs in response to the known NLRP3 activator, ATP, whereas knockdown of cofilin-1 reduces NLRP3 inflammasome activation. Cofilin-1 directly interacts to the nucleotide-binding domain (NBD) of the NLRP3 protein in LPS-primed BMDMs. However, cofilin-1 is dissociated from NLRP3 in a ROS-sensitive manner when the cells are stimulated with the NLRP3 inflammasome activators, ATP or nigericin, which induce oxidation of cofilin-1. Indeed, the interaction of cofilin-1 with NLRP3 is increased significantly when the oxidation site of cofilin-1 is substituted from cysteine to alanine. Moreover, the assembly of inflammasome components is impaired in cells expressing oxidation-resistant mutant cofilin-1.

Conclusion

Taken together, these findings suggest that cofilin-1 is a key component in regulating the NLRP3 inflammasome in response to ROS. In addition, our data suggest a potential target for the inflammatory conditions involving the NLRP3 inflammasome, including gout, type 2 diabetes mellitus, atherosclerosis, and Alzheimer's disease.