IPAB MEDIATES MACROPHAGE APOPTOSIS INDUCED BY SHIGELLA-FLEXNERI

Shigella flexneri kills macrophages through apoptosis, involving the i nduction of host cell DNA fragmentation and characteristic morphologic al changes. Shigella can only cause damage it it escapes from the phag olysosome into the cytoplasm. The S. flexneri cytotoxic genes have bee n localized to the ipa operon of shigella's virulence plasmid. ipaB, C and D deletion mutants are not invasive and therefore not cytotoxic. In order to distinguish genes involved in the escape from the phagolys osome as distinct from cytotoxicity, we constructed Shigella strains t hat secrete law amounts of Escherichia call haemolysin (hly(low)). The se strains can escape into the cytoplasm of the macrophage even in the absence of the invasion plasmid as verified by electron microscopy an d resistance to chloroquine. Macrophages were infected with different ipa mutants expressing hly(low). Both Delta ipaC hly(low) and Delta ip aD hly(low) were cytotoxic whilst Delta ipa hly(low) and a hly(low) st rain cured of shigella's pathogenicity plasmid were not. Furthermore, both Delta ipaC hly(low) and Delta ipaD hly(low) killed through apopto sis as shown by both changes in ultrastructural morphology and fragmen tation of the host cell DNA. These results demonstrate that ipaB is es sential for S. flexneri to induce apoptosis in macrophages.