YDZG enhances TRIM31 expression to promote NLRP3 protein degradation.
Previous studies showed that CY-09 (Jiang et al., 2017) and OLT1177 (Marchetti et al., 2018) directly bind to NLRP3 to attenuate NLRP3 inflammasome activation. We investigated the ability of YDZG to directly bind to NLRP3 by thermal shift assay. Results suggested that YDZG does not directly bind to NLRP3 (Figure S3). A recent study found that the TRIM31 attenuates NLRP3 inflammasome activation by promoting proteasomal degradation of NLRP3 (Song et al., 2016). TRIM31 directly bound to NLRP3, then promoted K48-linked polyubiquitination and proteasomal degradation of NLRP3 in both resting and activated macrophages (Song et al., 2016). We found that YDZG enhanced TRIM31 expression in resting J774A.1 (Figure 5A) and THP-1 (Figure 5B) cells. Since constitutively expressed TRIM31 can bind to NLRP3, promoting K48-linked ubiquitination and proteasomal degradation of NLRP3, we investigated whether YDZG could mediate the expression of TRIM31 to promote NLRP3 degradation. After the NLRP3 inflammasome was activated by LPS and ATP, YDZG treatment enhanced TRIM31 protein expression (Figures 5C- D), resulting in the inhibition of NLRP3 expression and subsequent inflammasome activation. Furthermore, as shown in Figure 5E, YDZG increased the formation of TRIM31-NLRP3 complexes. In summary, these findings indicated that YDZG enhanced TRIM31 expression to promote proteasomal degradation of NLRP3 in both resting and activated macrophages.