BIOGENESIS AND REGULATION OF THE VIBRIO-CHOLERAE TOXIN-COREGULATED PILUS - ANALOGIES TO OTHER VIRULENCE FACTOR SECRETORY SYSTEMS

Biogenesis of the toxin-coregulated pilus (TCP) of Vibrio cholerae 01 is essential for successful bacterial colonization of the small intest ine. Pilus assembly requires the products of at least seven genes loca ted on the chromosome adjacent to the pilin-encoding gene, tcpA. Previ ously reported TnphoA insertions in the TCP-assembly-deficient V. chol erae strains, KP2.21 and KP4.2, were isolated from the chromosome for further analysis. Nucleotide sequencing of the tcpE=phoA and tcpF=phoA fusions and corresponding clones of the region containing the intact genes revealed the presence of two open reading frames (ORFs) of 340 a nd 338 amino acids, designated TcpE and TcpF, respectively. The partia l sequence of an ORF downstream from the TcpF coding sequence was dete rmined to correspond to the global virulence regulator, ToxT. Proteins corresponding to the observed ORFs were visualized with the T7 promot er/RNA polymerase expression system. Computer-generated alignment algo rithms predict that a homology exists between TcpE and the Klebsiella pneumoniae pullulanase secretion proteins PulD and PulF, the Xanthomon as campestris extracellular enzyme secretion factor XpsF, the Bacillus subtilis DNA competence protein ComG-ORF2, and the Yersinia entercoli tica Yop secretion determinant YscC. These observations provide a mode l to investigate further the relationship between the secretion mechan isms utilized by these seemingly diverse virulence determinants. Addit ionally, an extreme C-terminal segment of TcpE shows striking homology to the transmembrane segment of the eukaryotic integrin beta-1 chain, which could imply a role for TcpE in not only TCP secretion, but also host cell interaction.