Recruitment of cytoskeletal and signaling proteins to enteropathogenic andenterohemorrhagic Escherichia coli pedestals

Enteropathogenic Escherichia coli (EPEC) is a human pathogen that attaches to intestinal epithelial cells and causes chronic watery diarrhea. A close relative, enterohemorrhagic E. coli (EHEC), causes severe bloody diarrhea a nd hemolytic-uremic syndrome. Both pathogens insert a protein, Tir, into th e host cell plasma membrane where it binds intimin, the outer membrane liga nd of EPEC and EHEC. This interaction triggers a cascade of signaling event s within the host cell and ultimately leads to the formation of an actin-ri ch pedestal upon which the pathogen resides. Pedestal formation is critical in mediating EPEC- and EHEC-induced diarrhea, get very little is known abo ut its composition and organization. In EPEC, pedestal formation requires T ir tyrosine 474 phosphorylation. In EHEC Tir is not tyrosine phosphorylated , yet the pedestals appear similar. The composition of the EPEC and EHEC pe destals was analyzed by examining numerous cytoskeletal, signaling, and ada pter proteins, Of the 25 proteins examined, only two, calpactin and CD44, w ere recruited to the site of bacterial attachment independently of Tir, Sev eral others, including ezrin, talin, gelsolin, and tropomyosin, were recrui ted to the site of EPEC attachment independently of Tir tyrosine 474 phosph orylation but required Tir in the host membrane, The remaining proteins wer e recruited to the pedestal in a manner dependent on Tir tyrosine phosphory lation or were not recruited at all. Differences were also found between th e EPEC and EHEC pedestals: the adapter proteins Grb2 and CrKII were recruit ed to the EPEC pedestal but were absent in the EHEC pedestal. These results demonstrate that although EPEC and EHEC recruit similar cytoskeletal prote ins, there are also significant differences in pedestal composition.