Shigella, the etiological agent of dysentery, kills macrophages by inducing apoptosis. Deletion mutants in the invasion invasion plasmid antigen B (ipaB) of Shigella flexneri are not cytotoxic. Here, we localized IpaB to the cytoplasm of macrophages infected with S. flexneri. Purified IpaB induced apoptosis when microinjected into macrophages, indicating that IpaB is sufficient to induce apoptosis. Using a GST-IpaB fusion protein as a ligand in affinity purification, we isolated four IpaB binding proteins from macrophages which were identified as the precursor and the mature polypeptides of interleukin-1beta converting enzyme (ICE) or a highly homologous protease. We found that IpaB binds directly to ICE and this enzyme is activated during S. flexneri infection. Furthermore, specific inhibitors of ICE prevented Shigella-induced apoptosis.