Fig. 4. Src kinase Lyn phosphorylates NLRP3 at Tyr 918, which facilitates NLRP3 ubiquitination and subsequent degradation.

(A) Nlrp3^–/– BMDMs were reconstituted with the constructs expressing pCMV6, NLRP3^WT, NLRP3^Y861F, NLRP3^Y918F and NLRP3^Y861F/918F, primed with LPS, stimulated with ATP. The cell lysates were immunoprecipitated with antibody to NLRP3, and blotted with antibodies to pTyr and ubiquitin, respectively. The cell lysates were blotted with antibodies to Flag and actin, respectively. (B) Nlrp3^–/– BMDMs were reconstituted with the constructs expressing NLRP3^WT and NLRP3^Y918F, primed with LPS, and stimulated with ATP. The IL-1β in the supernatants was determined by ELISA. (C) Nlrp3^–/– BMDMs were reconstituted with the constructs expressing NLRP3^WT and NLRP3^Y918F, primed with LPS and stimulated with ATP. The cell lysates were blotted with antibodies to NLRP3 and actin. Data are representative of three independent experiments (A to C). **P< 0.01, student t test.