Extended Data Fig. 6: Activity of NLRP3 variants in response to nigericin treatment.
a-b. Biplot for the ASC50 of untreated (UT) and nigericin (2 μM) treated NLRP3 variants from the INFEVERS database (a) or the ClinVar database (b). c-d. Percentage of cells expressing GFP-tagged wild-type (WT), Y861H (c), R920Q (d) in reconstituted THP-1 cells treated with or without doxycycline (Dox, 100 ng/mL) for 6 hours. e. ASC speck percentage in the cell population with an increasing amount of NLRP3 expression for wild-type (WT), and F410S NLRP3 in ASC-BFP HEK392T cells 12 hours post-transfection with or without 1-hour nigericin (Nig., 2 μΜ) treatment. The same WT control data is used as in Extended Data Fig. 5 for R556*, as results were generated from the same experiment. f. ASC50 of NLRP3 variants treated with H2O, 50 mM,100 mM, or 1 M KCl for 12 hours simultaneously with plasmids expression. g. ASC50 of NLRP3 variants pre-treated with H2O, 50 mM,100 mM, or 1 M KCl, 30 mins prior to nigericin treatment. ASC50 is calculated in relative to the activity of each variant without nigericin treatment. h. Overlay of inactive NLRP3 in purple (PDB: 7PZC) with the NEK7-bound active NLRP3 in orange (PDB: 8EJ4). NEK7 is shown in green. Wild-type residues representing the ___location of Y861H and R920Q NLRP3 variants are highlighted as spheres in red. i. ASC50 for the NLRP3 variants overexpressed in ASC-BFP HEK293T cells with empty vector (EV) or mCherry-NEK7K64M. Each dot represents a single variant (a-b), or one independent repeat (c-d, f-g and i). Data were pooled from 3–12 independent repeats (a-b, n value for each variant is listed in Supplementary Table 1), or 3 (c-e, i), 2 (f), 4 (g) independent repeats (mean in a-b, mean and s.e.m. in c-g, i). Two-tailed t-test in c-d and i, one-way ANOVA with Dunnett’s multiple-comparisons test in g.
