GENERAL SECRETION PATHWAY (EPS) GENES REQUIRED FOR TOXIN SECRETION AND OUTER-MEMBRANE BIOGENESIS IN VIBRIO-CHOLERAE

The general secretion pathway (GSP) of Vibrio cholerae is required for secretion of proteins including chitinase, enterotoxin, and protease through the outer membrane. In this study, we report the cloning and s equencing of a DNA fragment from V. cholerae, containing 12 open readi ng frames, epsC to -N, which are similar to GSP genes of Aeromonas, Er winia, Klebsiella, Pseudomonas, and Xanthomonas spp. In addition to th e two previously described genes, epsE and epsM (M. Sandkvist, V. Mora les, and hi. Bagdasarian, Gene 123: 81-86, 1993; L. J. Overbye, M. San dkvist, and M. Bagdasarian, Gene 132:101-106, 1993), it is shown here that epsC, epsF, epsG, and epsL also encode proteins essential for GSP function, Mutations in the Eps genes result in aberrant outer membran e protein profiles, which indicates that the GSP, or at least some of its components, is required not only for secretion of soluble proteins but also for proper outer membrane assembly. Several of the Eps prote ins have been identified by use of the T7 polymerase-promoter system i n Escherichia coli. One of them, a pilin-like protein, EpsG, was analy zed also in V. cholerae and found to migrate as two bands on polyacryl amide gels, suggesting that in this organism it might be processed or otherwise modified by a prepilin peptidase. We believe that TcpJ prepi lin peptidase, which processes the subunit of the toxin coregulated pi lus, TcpA, is not involved in this event. This is supported by the obs ervations that apparent processing of EpsG occurs in a tcpJ mutant of V. cholerae and that, when coexpressed in E. coli, TcpJ cannot process EpsG although the PilD peptidase from Neisseria gonorrhoeae can.