Extended Data Fig. 4 |. Lack of DDX3X leads to defects in both stress granule assembly and NLRP3 inflammasome activation.

a, Confocal microscopy of Ddx3x^fl/fl and Ddx3x^fl/flLysM^cre BMDMs stimulated with LPS and arsenite to visualize localization of the stress granule marker G3BP1 and DDX3X. Scale bars, 10 μm. b, Quantification of the number of stress granules per cell. Each data point represents the average number of stress granules per cell in each field of view. ***P = 0.0003 (unpaired two-sided t-test). Data are mean ± s.e.m. (n = 10). c, Immunoblot analysis of CASP1 cleavage in peritoneal macrophages stimulated with LPS with or without nigericin. Representative blots (n = 2 biologically independent experiments). d, Pyroptotic cell death as measured by the number of SYTOX Green+ cells. Cells were stimulated with LPS and treated with nigericin or with nigericin and arsenite. ****P ≤ 0.0001 (two-way ANOVA) for LPS + N-treated Ddx3x^fl/fl versus LPS + N-treated Ddx3x^fl/fl LysM^cre BMDMs; LPS + N-treated versus LPS + Ar + N-treated Ddx3x^fl/fl BMDMs; and LPS + N-treated versus LPS + Ar + N-treated Ddx3x^fl/fl LysM^cre BMDMs. Data are mean ± s.e.m. (n = 4). e–g, Immunoblot analysis of CASP1 cleavage in BMDMs treated with LPS or LPS and ATP (e), Pam3CSK4 or Pam3CSK4 and nigericin (f), or poly(I:C) or poly(I:C) and nigericin (g). Representative blots (n = 2 biologically independent experiments each).