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. 2021 Aug 26;12:741839. doi: 10.3389/fimmu.2021.741839

Figure 4.

Figure 4

NLRP3 agonists promote TRIM65 reduction to favor NEK7-NLRP3 interaction and NLRP3 inflammasome assembly. (A) PMA-differentiated and LPS-primed THP-1 cells were stimulated with MSU for different durations, and the expression of TRIM65 was analyzed by western blotting. (B) PMA-differentiated and LPS-primed THP-1 cells were stimulated with MSU, R837 or nigericin, and the expression of TRIM65 was analyzed by western blotting. (C) PMA-differentiated and LPS-primed THP-1 cells were treated with different doses of proteasome inhibitors (MG132, 1 h) or autophagy inhibitors (3-MA, 3 h), and then stimulated with R837 for 4 h, and the expression of TRIM65 was analyzed by western blotting. (D) PMA-differentiated and LPS-primed THP-1 cells were stimulated with MSU, and the endogenous interaction between NLRP3 and TRIM65 was analyzed by immunoprecipitation and western blotting. (E) LPS-primed BMDM cells were stimulated with ATP and Nigericin, and the interaction between NLRP3 and NEK7 was analyzed by immunoprecipitation and western blotting. (F) Mcherry-NLRP3 and Flag-NEK7 were cotransfected with or without GFP- TRIM65 in HEK-293T cells. The interaction between NLRP3 and NEK7 was analyzed by immunoprecipitation and western blotting. Data are representative of three independent experiments.