The LEE1 promoters from both enteropathogenic and enterohemorrhagic Escherichia coli can be activated by PerC-like proteins from either organism.
The PerC protein of enteropathogenic Escherichia coli (EPEC), encoded by the pEAF plasmid, is an activator of the locus of enterocyte effacement (LEE) pathogenicity island via the LEE1 promoter. It has been assumed that the related LEE-containing pathogen enterohemorrhagic E. coli (EHEC) lacks PerC-dependent activation due to utilization of an alternative LEE1 promoter and lack of a perC gene. However, we show here that EPEC PerC can activate both the EPEC and EHEC LEE1 promoters and that the major transcriptional start site is similarly located in both organisms. Moreover, a PerC-like protein family identified from EHEC genome analyses, PerC1 (also termed PchABC), can also activate both promoters in a manner similar to that of EPEC PerC. The perC1 genes are carried by lambdoid prophages, which exist in multiple copies in different EHEC strains, and have a variable flanking region which may affect their expression. Although individual perC1 copies appear to be poorly expressed, the total perC1 expression level from a strain encoding multiple copies approaches that of perC in EPEC and may therefore contribute significantly to LEE1 activation. Alignment of the protein sequences of these PerC homologues allows core regions of the PerC protein to be identified, and we show by site-directed mutagenesis that these core regions are important for function. However, purified PerC protein shows no in vitro binding affinity for the LEE1 promoter, suggesting that other core E. coli proteins may be involved in its mechanism of activation. Our data indicate that the nucleoid-associated protein IHF is one such protein.
Porter
Megan E
ME
Zoonotic and Animal Pathogens Research Laboratory, Medical Microbiology, University of Edinburgh, Teviot Place, Edinburgh, EH8 9AG, United Kingdom. Megan.Porter@ed.ac.uk
Mitchell
Paul
P
Free
Andrew
A
Smith
David G E
DG
Gally
David L
DL
D024506:5' Flanking Region; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D029968:Escherichia coli Proteins / Q000378:metabolism* / Q000502:physiology; D018628:Gene Dosage; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005814:Genes, Viral; D044404:Genomic Islands / Q000235:genetics / Q000502:physiology; D034982:Integration Host Factors / Q000502:physiology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010957:Plasmids; D011401:Promoter Regions, Genetic; D039002:Prophages / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D024363:Transcription Initiation Site / Q000502:physiology; D014158:Transcription, Genetic; D037521:Virulence Factors / Q000235:genetics*
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D034982:Integration Host Factors; D037521:Virulence Factors; C440502:integration host factor, E coli