Figure 5

N-terminal five amino acids are indispensable for GSDMD-N to form oligomers and trigger pyroptosis. (A) Sequence alignment of N-terminal 20 amino acids of human GSDMA, GSDMB, GSDMC, GSDMD and mouse GSDMD (upper panel), and schematic representation of N-terminal deletions of GSDMD-N fused with HBD^* (lower panel). (B) Viabilities of CHO cells expressing different GSDMD-N-HBD^*-HA deletion mutants after stimulation with DMSO or 4-OHT for 0.5 h. Data represented the mean ± sd of three independent experiments. (C) Immunoblot analysis of lysates of CHO cells in B resolved by 4% - 12% gradient SDS-PAGE under non-reducing condition. (D) Deconvolution microscopy of CHO cells expressing different GSDMD-N-HBD^*-HA deletion mutants treated with 4-OHT for 1 h. After treatment, cells were immunostained for HA and counterstained with Hoechst and PI. PI positive indicates dead cell. Scale bar, 5 μm.