Figure S2.
Quantification of RelA nuclear translocation, MAPK p38 induction, and cytokine release in gene-edited EL4 cells. (A) RelA translocation to the nucleus in WT and gene-edited EL4 cells. EL4 cell lines (30 min after addition to IL1β-labeled SLBs) were fixed and stained for RelA (magenta); DAPI stained nuclei (blue). Scale bar, 50 µm. (B) Quantification of RelA nucleus-to-cytoplasm staining ratio. Violin plots show the single-cell distribution RelA nucleus-to-cytoplasm staining ratio. Colored dots superimposed on violin plots correspond to the average value in the independent experiments (n = 3 or 4 experimental replicates per cell line; each replicate encompasses measurements from >2,000 cells. Bars represent mean ± SEM. P values were calculated using a two-tailed unpaired Student's t test. (C) Phospho-p38 staining intensity in WT and gene-edited EL4 cells. EL4 cell lines (30 min after addition to IL1β-labeled SLBs) were fixed and stained for phospho-p38 (magenta); DAPI stained nuclei (blue). Scale bar, 50 µm. (D) Quantification of phospho-p38 staining intensity. Violin plots show the single cell distribution phospho-p38 staining intensity. Colored dots superimposed on violin plots correspond to the average value in the independent experiments (n = 3 or 4 experimental replicates per cell line; each replicate encompasses measurements from >6,000 cells). Bars represent mean ± SEM. P values were calculated using a two-tailed unpaired Student's t test. (E) IL2 release in WT and gene-edited EL4 cells. IL2 release was measured by ELISA 24 h after IL1β stimulation. Values for gene-edited cells shown relative to EL4 WT. Average values calculated from three independent experiments. Bars represent mean ± SEM. (F and G) MyD88-GFP puncta can fuse and split. (F) Example of two MyD88-GFP puncta undergoing fusion. (G) Example of MyD88-GFP puncta undergoing fission. Scale bar, 2 µm. Fluo., fluorescence; p-p38, phospho-p38.

Quantification of RelA nuclear translocation, MAPK p38 induction, and cytokine release in gene-edited EL4 cells. (A) RelA translocation to the nucleus in WT and gene-edited EL4 cells. EL4 cell lines (30 min after addition to IL1β-labeled SLBs) were fixed and stained for RelA (magenta); DAPI stained nuclei (blue). Scale bar, 50 µm. (B) Quantification of RelA nucleus-to-cytoplasm staining ratio. Violin plots show the single-cell distribution RelA nucleus-to-cytoplasm staining ratio. Colored dots superimposed on violin plots correspond to the average value in the independent experiments (n = 3 or 4 experimental replicates per cell line; each replicate encompasses measurements from >2,000 cells. Bars represent mean ± SEM. P values were calculated using a two-tailed unpaired Student's t test. (C) Phospho-p38 staining intensity in WT and gene-edited EL4 cells. EL4 cell lines (30 min after addition to IL1β-labeled SLBs) were fixed and stained for phospho-p38 (magenta); DAPI stained nuclei (blue). Scale bar, 50 µm. (D) Quantification of phospho-p38 staining intensity. Violin plots show the single cell distribution phospho-p38 staining intensity. Colored dots superimposed on violin plots correspond to the average value in the independent experiments (n = 3 or 4 experimental replicates per cell line; each replicate encompasses measurements from >6,000 cells). Bars represent mean ± SEM. P values were calculated using a two-tailed unpaired Student's t test. (E) IL2 release in WT and gene-edited EL4 cells. IL2 release was measured by ELISA 24 h after IL1β stimulation. Values for gene-edited cells shown relative to EL4 WT. Average values calculated from three independent experiments. Bars represent mean ± SEM. (F and G) MyD88-GFP puncta can fuse and split. (F) Example of two MyD88-GFP puncta undergoing fusion. (G) Example of MyD88-GFP puncta undergoing fission. Scale bar, 2 µm. Fluo., fluorescence; p-p38, phospho-p38.

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