Organization and architecture of AggR-dependent promoters from enteroaggregative Escherichia coli

Enteroaggregative Escherichia coli (EAEC), is a diarrhoeagenic human pathogen commonly isolated from patients in both developing and industrialized countries. Pathogenic EAEC strains possess many virulence determinants, which are thought to be involved in causing disease, though, the exact mechanism by which EAEC causes diarrhoea is unclear. Typical EAEC strains possess the transcriptional regulator, AggR, which controls the expression of many virulence determinants, including the attachment adherence fimbriae (AAF) that are necessary for adherence to human gut epithelial cells. Here, using RNA-sequencing, we have investigated the AggR regulon from EAEC strain 042 and show that AggR regulates the transcription of genes on both the bacterial chromosome and the large virulence plasmid, pAA2. Due to the importance of fimbriae, we focused on the two AAF/II fimbrial gene clusters in EAEC 042 (afaB-aafCB and aafDA) and identified the promoter elements and AggR-binding sites required for fimbrial expression. In addition, we examined the organization of the fimbrial operon promoters from other important EAEC strains to understand the rules of AggR-dependent activation. Finally, we generated a series of semi-synthetic promoters to define the minimal sequence required for AggR-mediated activation and show that the correct positioning of a single AggR-binding site is sufficient to confer AggR-dependence.