Analysis of the gene that regulate production of Vibrio cholerae El Tor cytolysin

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From the Tn5-inserted mutant library of Vibrio cholerae O1, we found a mutant, NF404, which lost the production of both hemolysin and cholera toxin (CT) even though the Tn5-insertion site was out side from the structural genes for hemolysin and CT.Cloning and sequencing analysis of the homologous region from the wild-type strain, revealed that the sequence spanning the coding region of an ORF1 nominated as lypA,encoding a 39.5 kDa protein. Deduced amino acid sequence of the lypA gene had 37.6% identity to the lysophospholipase L2 of Escherichia coli. Introduction of lypA into NF404 has lost cholera toxin and hemolysin productions, restored the lysophospholipase activity and CT production but not the hemolytic activity. Inactivation of lypA of the wild-type strains by single crossover insertion decreased the lysophospholipase L2 activity and cholera toxin production but not the hemolytic activity. These defective strains decreased enterotoxicity. These results suggest that lypA is possibly required for the expression of CT and may play a role in pathogenicity of Vibrio cholerae.