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Formation of attaching and effacing lesions
The best-characterized interactions between EHEC and gut epithelia involve
the formation of AE lesions, described above, which are characteristic of EHEC
and the other AE pathogens EPEC and
C. rodentium
. AE lesion formation
requires Tir, which is translocated to the host cell by a T3SS, a multi-protein
export apparatus that spans the inner and outer membranes and facilitates direct
injection of bacterial effectors from the bacterial cytoplasm into the host cell
(see Chapter 14;
Garmendia et al., 2005
). Tir, intimin, and the entire T3SS are
encoded on a ∼35 kb chromosomal pathogenicity island called the Locus of
Enterocyte Effacement (LEE) (
McDaniel et al., 1995
).
Tir, like intimin, is absolutely required for AE lesion formation and col-
onization in many animal models (
Marches et al., 2000
;
Deng et al., 2003
;
Ritchie et al., 2003
). Once injected into the host cell, Tir integrates into the
host cell membrane in a hairpin loop conformation, with its central extracellular
domain serving as the receptor for the C-terminal extracellular domain of inti-
min (
Batchelor et al., 2000
;
Luo et al., 2000
;
Liu et al., 2002
). Intimin-binding
by Tir induces Tir clustering (
Touze et al., 2004
) and permits the cytoplasmic
domains of Tir to coordinate a downstream signaling cascade resulting in the
formation of F-actin pedestals (
Campellone et al., 2006
;
Hayward et al., 2006
;
Brady et al., 2007
).
A cytoplasmic activity of Tir critical for F-actin pedestal formation is rec-
ognition by mammalian adaptor proteins IRTKS (Insulin Receptor Tyrosine
Kinase Substrate) and/or IRSp53 (Insulin Receptor Substrate Protein 53 kDa)
(
Vingadassalom et al., 2009
;
Weiss et al., 2009
;
de Groot et al., 2011
). These
homologous proteins encode an I-BAR (membrane-binding) and an SH3
(polyproline-binding) domain, and can stimulate low-level actin assembly.
However, in EHEC strain O157:H7, robust actin pedestal formation requires a
second-type III-secreted effector, EspF
U
/TccP, which is encoded on the cryptic
prophage CP-933U/Sp14 (
Campellone et al., 2004
;
Garmendia et al., 2004
),
and which is recruited to sites of bacterial binding by the IRTKS/IRSp53 SH3
domain. In turn, EspF
U
/TccP binds directly to the GTPase binding (GBD) region
of N-WASP, resulting in its activation (
Cheng et al., 2008
;
Sallee et al., 2008
;
Campellone, 2010
) and the subsequent activation of the actin nucleator Arp
2/3 (reviewed in
Goley and Welch (2006)
and
Stradal and Scita (2006)
), finally
resulting in actin pedestal formation.
Mutants of EHEC or other AE pathogens that entirely lack intimin or Tir are
incapable of generating AE lesions and unable to colonize a range of mammalian
hosts (
Donnenberg et al., 1993a
;
Tzipori et al., 1995
;
Marches et al., 2000
;
Deng
et al., 2003
;
Ritchie et al., 2003
). Given that intimin- or Tir-deficient mutants
are almost entirely defective for mammalian cell attachment, their inability to
colonize animals may reflect a binding defect rather than a pedestal formation
defect per se. Instead, the potential role of actin assembly in the pathogenesis
of disease requires the analysis of mutants of AE pathogens that express inti-
min and Tir and are competent for attachment but are specifically defective for
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