The pathogen protein EspFU hijacks actin polymerization using mimicry and multivalency

Abstract

Enterohemorrhagic E. coli (EHEC) attaches to the intestine through actin pedestals that are formed when the bacterium injects the protein EspF U into host cells 1

Enterohemorrhagic Escherichia coli raises the I-BAR

“…This array, combined with the proximity of bound IRTKS and N-WASP may promote the clustering of multiple copies of each molecule into a scaffold that promotes highly-efficient actin assembly. At a minimum, the presence of multiple N-WASP binding sites on each EspF U molecule provides the capacity for significant amplification of the actin assembly potential at this level, which is supported by experimental data showing that each additional repeat in EspF U effectively increases and amplifies N-WASP activity, with maximal activity reaching the same level as unregulated N-WASP (6).…”

“…EspF U directly activates the nucleation promoting factor N-WASP (neuronal Wiskott-Aldrich syndrome protein), which in turn activates Arp2/3-mediated actin polymerization. A repeated domain of EspF U is a structural mimic of the autoinhibitory element in N-WASP, such that EspF U binding relieves N-WASP autoinhibition (5,6). Although Tir does not interact directly with EspF U , translocation of Tir and EspF U into cells is sufficient for the induction of pedestal formation, indicating that the two are indirectly linked via a cellular factor.…”

“…Although not addressed by this analysis, because the EspF U derivative used in these experiments contained 5 repeats, it will be interesting to determine whether the proximity of the binding sites on a single repeat of EspF U permits simultaneous occupation by IRTKS and N-WASP. Each 47-residue repeat in a single EspF U molecule has the capacity to bind an N-WASP GBD, and EspF U derivatives containing increasing numbers of repeats display proportional increases in N-WASPmediated actin assembly in a variety of in vitro and in vivo assays (5,6). If each repeat of a single EspF U molecule is also able to bind an SH3 domain of IRTKS, which has not yet been tested, then the net effect might be to generate a reticular array of Tir molecules bound to IRTKS, in turn linked to one of the repeats of an EspF U molecule.…”

Allosteric N-WASP activation by an inter-SH3 domain linker in Nck

“…The activation motif in EspF U and EspF has the same pattern of hydrophobic residues as the Nck motif (Fig. 4B) (Φ-Φ-x-x-Φ-x-x-x-Φ; Φ is any hydrophobic residue and x is any residue) (39)(40)(41). Structural studies by multidimensional NMR have shown that the EspF U peptide adopts a helical conformation when bound to the WASP GBD (40).…”

An efficient method to produce recombinant bacterial effector Tir coupled cytoskeleton protein (TCCP) complexed with host protein insulin receptor tyrosine kinase substrate (IRTKS)

“…Previous studies have shown that the number of PRR regions in TCCP of EHEC varies from three to six and is related to the efficiency of actin polymerization (Garmendia et al, 2005). However, the underlying mechanism remains poorly understood (Sallee et al, 2008). Although the structure of the SH3 complex with one peptide from the PRR has been determined (Aitio et al, 2010), it is inadequate to fully explain the linkage between the number of PRRs and the level of actin polymerization.…”