Intimin types α, β, and γ bind to nucleolin with equivalent affinity but lower avidity than to the translocated intimin receptor

Abstract

The outer membrane adhesins of enteropathogenic Escherichia coli, Citrobacter rodentium, and enterohemorrhagic E. coli (EHEC) O157:H7 that mediate attach and efface intestinal lesions are classified as intimin α, β, and γ, respectively. Each of these intimin types binds to its cognate, bacterially encoded receptor (called Tir for translocated intimin receptor) to promote tight adherence of the organism to the host-cell plasma membrane. We previously reported that γ intimin of EHEC O157:H7 also bound to a eucaryotic receptor that we determined was nucleolin. The objective of this study was to investigate in vitro and in vivo the interactions of intimins α, β, and γ with nucleolin in the presence of Tir from EHEC O157:H7. Protein binding experiments demonstrated that intimin of types α, β, and γ bound nucleolin with similar affinity. Moreover, all three intimin types colocalized with regions of nucleolin expressed on the surface of HEp-2 cells. When intimin α, β, or γ bound to Tir in vitro, the intimin interaction with nucleolin was blocked. Both Tir and nucleolin accumulated beneath intimin-presenting bacteria that had attached to the surface of HEp-2 cells. Taken together, these findings suggest that nucleolin is involved in bacterial adherence promoted by all intimin types and that Tir and nucleolin compete for intimin during adherence.