Autoactivation of the AggR regulator of enteroaggregative Escherichia coli in vitro and in vivo

Enteroaggregative Escherichia coli (EAEC) causes diarrhea in diverse populations worldwide. The AraC-like regulator AggR is a key virulence regulator in EAEC. AggR-regulated genes include those encoding the Aggregative Adherence Fimbria, the dispersin protein, and a type VI secretion system. This study characterizes the regulation of the aggR promoter (P-aggR). Using primer extension analysis, the transcriptional start site of the aggR promoter was located 40 nucleotides upstream of the translational start. P-aggR was found to be autoregulated and DNA footprinting revealed the presence of two AggR-binding sites: one upstream of the transcriptional start site and one downstream. Additionally, P-aggR was found to be positively regulated by the DNA-binding protein FIS and negatively regulated by the global regulator H-NS. To further understand this complex regulation scheme, a bacterial luciferase reporter system was used with a mouse model of EAEC colonization. This allowed for the in vivo measurement of P-aggR, P-fis, and P-hns activity. EAEC present in the mouse intestine possessed relatively high levels of P-fis and P-aggR activity and a low level of P-hns when compared with in vitro experiments. The data provide significant insights into the regulation cascade leading to aggR expression in the mammalian intestine during EAEC infection.