A NEW CYTOLETHAL DISTENDING TOXIN (CDT) FROM ESCHERICHIA-COLI PRODUCING CNF2 BLOCKS HELA-CELL DIVISION IN G2 M PHASE/

Escherichia coil strain 1404, isolated from a septicaemic calf, carrie s a transferable plasmid called pVir which codes for the cytotoxic nec rotizing factor type 2 (CNF2). A 4 h interaction between strain 1404 a nd HeLa cells induced the formation of giant mononucleated cells block ed in G2/M phase. Mating experiments between strain 1404 and a non-pat hogenic recipient strain demonstrated that the factor(s) encoded by pV ir mediated the cell-cycle arrest. A 3.3 kb DNA fragment isolated from a DNA bank of pVir was shown to code for the factor(s) causing the ce ll-cycle arrest. Nucleotide sequence analysis revealed the presence of three genes encoding proteins sharing significant amino acid homology with the cytolethal distending toxins (CDTs) previously isolated from E. coli, Campylobacter jejuni and Shigella dysenteriae. Southern hybr idization experiments demonstrated that the pVir of other CNF2-produci ng E. coil strains contained sequences related to cdt. Although the am ino acid sequences amongst CDT diverged significantly, the two other C DTs previously isolated from E. coil were also able to block the HeLa cell cycle. In conclusion, this study demonstrates the mode of action of CDT and will help us to elucidate the role of this emerging toxin f amily in microbial pathogenesis.