Recruitment and membrane interactions of host cell proteins during attachment of enteropathogenic and enterohemorrhagic Escherichia coli.
Résumé
Enteropathogenic (EPEC) and enterohemorrhagic Escherichia coli (EHEC) are attaching and effacing pathogens frequently associated with infectious diarrhoea. EPEC and EHEC use a type III secretion system (T3SS) to translocate effectors that subvert different cellular processes to sustain colonization and multiplication. The eukaryotic proteins Na+/H+ exchanger regulatory factor 2 (NHERF2) and Annexin A2 (AnxA2), which are involved in regulation of intestinal ion channels are recruited to the bacterial attachment sites. Using a stable HeLa-HA-NHERF2 cell line we found partial colocalization of AnxA2 and NHERF2; in EPEC infected cells AnxA2 and NHERF2 were extensively recruited to the site of bacterial attachment. We confirmed that NHERF2 dimerises and found that NHERF2 interacts with AnxA2. Moreover, we found that AnxA2 also binds both the N- and C-terminal domains of the bacterial efector Tir through its C-terminal domain. Immunofluorescence of HeLa cells infected with EPEC showed that AnxA2 is recruited to the site of bacterial attachment in a Tir-dependent manner, but independently of Tir-induced actin polymerization. Our results suggest that AnxA2 and NHERF2 form a scaffold complex that links adjacent Tir molecules at the plasma membrane forming a lattice that could be involved in retention and dissemination of other effectors at the bacterial attachment site.