Expression control mechanisms of LEE pathogenic factors by AAA proteases in Enterohaemorrhagic and Enteropathogenic Eschericha coli

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1. Enterohemorrhagic Escherichia coli (EHEC) and Enteropathogenic E. coli (EPEC) cause histopathological lesions of the intestinal epithelial cells that are known as attaching and effacing (A/E) lesions. Both strains carry a unique region of chromosomal DNA known as the locus for enterocyte effacement (LEE) that encodes a type III secretion system (TTSS) and virulence proteins. We have recently reported that AAA proteases (CIpXP, Lon) and molecular chaperones control 2 TTSSs in Salmonella Typhimurium: (i) flagellum biogenesis and (ii) the expression of the Pathogenicity Island 1. We demonstrated that the EHEC and EPEC clpPX mutants strongly impaired the secretion of virulence proteins by TTSS and repressed transcription from all the LEE promoters. The rpoS mutation in EHEC enhanced transcription from all the LEE promoters and the secretion of virulence proteins, and it could partially suppress the defects of the clpPX mutation. These data indicate that the EHEC ClpXP protease is a posi … More tive regulator for LEE expression, and this regulation occurs via 2 pathways: the o^<S->dependent and os-independent pathways.2. Recently, 2 LEE-encoded regulators-GrlA (global regulator of LEE activator) and GrlR (Grl repressor)-were identified. Iyoda et al. reported that ClpXP protease controls the expression of LEEpromoters via the regulation of GrlR levels in EHEC (J.Bacteriol. 2005. 187: 4086-94). However, the control mechanisms underlying the expression of LEEpromoters by Gr1A, GrlR, and AAA proteases remain unclear. Therefore, we established an assay system using nonpathogenic E. coli that enables analysis of the interaction among these proteins by measuring the activities of the LEE2, LEE3, and LEE5promoters. Our data revealed that the activation of ler (LEE-encoded regulator) expression by Gr1A entailed the participation of the domain that exists for a length of 131 base pairs downstream from the ler stop codon. Presently, we are analyzing the LEE expression control mechanisms exerted by Gr1R, Gr1A, and AAA proteases in detail. Less