SummaryEnteropathogenic Escherichia coli (EPEC) elicit changes in host cell morphology and cause actin rearrangement, a phenotype that has commonly been referred to as attaching/effacing (AE) lesions. The ability of EPEC to induce AE lesions is dependent upon a type III protein secretion/translocation system that is encoded by genes clustered in a 35.6 kb DNA segment, named the locus of enterocyte effacement (LEE). We used transcriptional fusions between the green fluorescent protein (gfp) reporter gene and LEE genes rorf2, orf3, orf5, escJ, escV and eae, together with immunoblot analysis with antibodies against Tir, intimin, EspB and EspF, to analyse the genetic regulation of the LEE. The expression of all these LEE genes was strictly dependent upon the presence of a functional integration host factor (IHF). IHF binds specifically upstream from the ler (orf1) promoter and appears to activate expression of ler, orf3, orf5 and rorf2 directly. The ler-encoded Ler protein was involved in activating the expression of escJ, escV, tir, eae, espB and espF. Expression of both IHF and Ler was needed to elicit actin rearrangement associated with AE lesions. In conclusion, IHF directly activates the expression of the ler and rorf2 transcriptional units, and Ler in turn mediates the expression of the other LEE genes.

Enteropathogenic Escherichia coli (EPEC) causes severe diarrhoea in young children. The locus of enterocyte effacement (LEE) pathogenicity island comprises a cluster of operons encoding a type III secretion system and related proteins that are associated with EPEC virulence. The LEE1 operon encodes Ler that positively regulates the LEE2, LEE3, LEE4, LEE5 and espG transcriptional units. The LEE operons are repressed at 27 SC and expressed at 37 SC. This paper describes a regulatory cascade of the thermoregulation of LEE operons. LEE1 including ler is repressed by H-NS at 27 SC but not at 37 SC. In contrast, the expression of the LEE2, LEE3, LEE4, LEE5 and espG transcriptional units is repressed by H-NS at both 27 SC and 37 SC. Upon shifting the culture temperature from 27 SC to 37 SC, Ler is synthesized and in turn activates the expression of LEE2, LEE3, LEE4 and espG by releasing the H-NS mediated repression. In the case of LEE5, Ler acts both by alleviating the H-NS mediated repression and by an additional mechanism, as yet to be defined.

Enteropathogenic Escherichia coli (EPEC) uses a type III secretion system (TTSS) to inject effector proteins into the plasma membrane and cytosol of infected cells. To translocate proteins, EPEC, like Salmonella and Shigella, is believed to assemble a macromolecular complex (type III secreton) that spans both bacterial membranes and has a short needle‐like projection. However, there is a special interest in studying the EPEC TTSS owing to the fact that one of the secreted proteins, EspA, is assembled into a unique filamentous structure also required for protein translocation. In this report we present electron micrographs of EspA filaments which reveal a regular segmented substructure. Recently we have shown that deletion of the putative structural needle protein, EscF, abolished protein secretion and formation of EspA filaments. Moreover, we demonstrated that EspA can bind directly to EscF, suggesting that EspA filaments are physically linked to the EPEC needle complex. In this paper we provide direct evidence for the association between an EPEC bacterial membrane needle complex and EspA filaments, defining a new class of filamentous TTSS.

Lrp (leucine-responsive regulatory protein) is a major Escherichia coli regulatory protein which regulates expression of a number of operons, some negatively and some positively. Operons that are affected by the presence or absence of Lrp includefanABC (3), gcv (10), ginA (5), gltBD (5), ilvIH (20), livJ and livK (7), lysU (6, 11), ompC and ompF (5), oppABCDF (1), papBA (3), sdaA (12), serA (12, 21), and tdh (12, 21). Some of these operons (e.g., ilvIH) are known to be directly controlled by Lrp, whereas others (e.g., ginA) are affected only indirectly by Lrp. Other operons that are affected by Lrp were identified by a plac Mu mutational analysis and by two-dimensional gel electrophoresis (5, 10). Many operons under control of Lrp are also subject to control by leucine. A striking feature of the Lrp regulon is the variety of ways that leucine and Lrp interact to regulate gene expression (16). Of the operons activated by Lrp, in some cases the activation requires leucine, in some cases the activation is negated by leucine, and in other cases the activation is independent of leucine. Similarly, for operons which are negatively regulated by Lrp, the same three subcategories have been observed: leucine negates the effect, leucine is required for the effect, and leucine has no effect. The molecular mechanisms underlying these six different patterns of regulation involving Lrp and leucine are only partially understood.Lrp, a dimer containing two identical subunits of molecular mass 18.8 kDa, is present in E. coli at a level of about 3,000 molecules per cell (32). Its amino acid sequence is evolutionarily related to that of AsnC, a regulatory protein that controls