Transcriptional regulation of the yghJ-pppA-yghG-gspCDEFGHIJKLM cluster encoding the type II secretion pathway in enterotoxigenic E. coli

Department of Microbiology and Immunology, University of Melbourne, Victoria 3010, and Murdoch Childrens Research Institute, Royal Children's Hospital, Parkville, Victoria 3052, Australia

^* To whom correspondence should be addressed. Email: r.browne{at}unimelb.edu.au.

	Abstract

The gene cluster gspCDEFGHIJKLM codes for various structural components of the type II secretion pathway which is responsible for the secretion of heat-labile enterotoxin by enterotoxigenic E. coli (ETEC). In this work, we used a variety of molecular approaches to elucidate the transcriptional organization of the ETEC type II secretion system and to unravel the mechanisms by which the expression of these genes is controlled. We showed that the gspCDEFGHIJKLM cluster and three other upstream genes, yghJ, pppA, and yghG are co-transcribed and that a promoter located in the upstream region of yghJ plays a major role in the expression of this 14-gene transcriptional unit. Transcription of the yghJ promoter was repressed 168-fold upon a temperature downshift from 37 to 22^oC. This temperature-induced repression was mediated by the global regulatory proteins, H-NS and StpA. Deletion mutagenesis showed that the promoter region encompassing positions -321 and +301 relative to the start site of transcription of yghJ was required for full repression. The yghJ promoter region is predicted to be highly curved and bound H-NS or StpA directly. The binding of H-NS or StpA blocked transcription initiation by inhibiting promoter open complex formation. Unraveling the mechanisms of regulation of type II secretion by ETEC enhances our understanding of the pathogenesis of ETEC and other pathogenic varieties of E. coli.