Figure 5. Cytoplasmic DNA-induced AIM2 oligomerization and pyroptosis.

a, b, Confocal live cell images of 293-EGFP cells (a) or 293-AIM2-EGFP-N1 (b) following transfection with vehicle (control, upper panels) or Cy™3-labeled DNA (red, lower panels). c, d, Confocal live cell images of NALP3^−/− -AIM2-EGFP-N1 bone marrow macrophages following transfection with vehicle (c) or Cy™3-labeled DNA (d). Notice the oligomerization of AIM2-GFP by the red DNA in the 293-AIM2-EGFP-N1 cells (b, bottom panels) and NALP3^−/− -AIM2-EGFP-N1 macrophages (d). The green (AIM2-EGFP), red (Cy™3-labeled DNA), gray (DIC), and blue (Hoechst stain, nucleus) channels, and the merged channels are indicated. The two white arrows (lower-second panel from left in (d)) indicate staining of the cytoplasmic DNA with the blue Hoechst stain, which specifically stains DNA. Notice the pyroptotic cell death features induced by the Cy™3-labeled DNA in NALP3^−/− -AIM2-EGFP-N1 macrophages (d), but not in the 293-AIM2-EGFP-N1 (b). e, Immunoblot for ASC showing the presence of the oligomeric ASC pyroptosomes only in the pellets of poly (dA:dT)-transfected (6^th lane), but not in the pellets of untransfected (5^th lane), NALP3^−/− macrophages. A shorter exposure and a more detailed legend of this gel is shown in Supplementary Fig. 12. f, Percentages of LDH release into the culture medium of NALP3^−/− macrophages transfected with vehicle (1^st and 2^nd columns), non-specific control siRNA (siCon, 3^rd and 4^th columns) or mouse AIM2-specific siRNA (siAIM2, 5^th and 6^th columns) followed by transfection with or without poly (dA:dT) (1 µg/1 × 10⁶ cells) as indicated. Values represent mean ± S.D. (n = 3); *, P < 0.001.