VIBRIO CHOLERAE-INDUCED APOPTOSIS OF MAMMALIAN CELLS MEDIATED BY EL TOR HAEMOLYSIN

SAKA HÉCTOR A., BIDINOST CARLA, ECHENIQUE JOSÉ, CHINEN ISABEL, BONACCI GUSTAVO AND BOCCO JOSÉ L.

San Carlos de Bariloche

Congreso; BARILOCHE PROTEIN SYMPOSIUM; 2003

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB) y la Sociedad Argentina de Biofísica (SAB)

El Tor Haemolysin (ETH) is a pore-forming toxin encoded by the hlyA gene of V. cholerae O1 biotype El Tor and most of V. cholerae non-O1/ non-O139 (VCN) isolates. Previous studies demonstrated that ETH is able to induce cytolytic, enterotoxic and vacuolating activity, suggesting that this toxin may contribute to the pathogenesis of gastroenteritis caused by V. cholerae strains lacking the cholera toxin (CT). In order to explore further mechanisms of cell damage, the potential involvement of apoptosis triggered by ETH was investigated. To this end, COS-7 cell monolayers were exposed to sterile culture supernatant from a clinical VCN strain, lacking CT, or from its isogenic hlyA null mutant. At different times post-incubation the apoptosis phenomenon was analyzed by DNA fragmentation, flow cytometry and TUNEL. The parental strain but not its isogenic hlyA null mutant induced internucleosomal DNA fragmentation, hypodiploidia and TdTmediated incorporation of fluorescent d-UTP, demonstrating the apoptotic state of treated cells. Apoptosis was also observed after exposure to 75 ng/mL of purified ETH. Furthermore, apoptotic activity of both parental strain and purified ETH was completely abolished after pre-incubation with an anti-ETH antibody. These results demonstrate that ETH is capable to induce apoptosis of mammalian COS-7 cells and points to previously unknown interactions between V. cholerae and its host.