Type 3 secretion systems - Escherichia coli: Pathotypes and Principles of Pathogenesis - page 412

a structured linker ( Crepin et al., 2005 ; Yip et al., 2005b ). Based on studies
of the Salmonella SPI-1 ortholog of EscD, PrgH, this protein likely forms a
24-mer ring in intimate contact with the EscJ ring in the periplasm ( Sanowar
et al., 2009 ; Schraidt et al., 2010 ; Schraidt and Marlovits, 2011 ). PrgH contains
a cytoplasmic domain and a periplasmic domain, which are separated by a
transmembrane helix. The cytoplasmic domain possesses a Forkhead-domain
associated (FHA) fold ( McDowell et al., 2011 ; Barison et al., 2012 ; Lountos
et al., 2012 ), and has been proposed to interact with elements of the export
apparatus ( Johnson et al., 2008 ). Whether this interaction is phosphorylation-
dependent, as predicted from the FHA fold, remains unknown. The periplasmic
region of PrgH is composed of three domains, each possessing a similar fold
to the two domains of EscJ ( Spreter et al., 2009 ). Collectively, the dual inner
membrane ring system of the T3SS is proposed to act as a structural foundation
for recruitment/stabilization and final assembly of the cytosolic ATPase and
several multispan membrane proteins that create an energized export appara-
tus for chronological secretion of downstream T3SS components and effectors
(see below for further details on the export apparatus components).
The outer-membrane ring is composed of a single protein, EscC, which belongs
to the secretin family of outer-membrane channels (see Korotkov et al., 2011 for
review). Its stoichiometry remains controversial, with 12-mers reported for the
Shigella T3SS secretin MxiD ( Hodgkinson et al., 2009 ), 13-mers for the Yersinia
T3SS secretin YscC ( Burghout et al., 2004b ), and 15-mers for the Salmonella
SPI-1 T3SS InvG ( Schraidt and Marlovits, 2011 ). This discrepancy may reflect
true system-to-system variation or may correspond to inherent plasticity in the
oligomerization state of this component, particularly when isolated in vitro. High-
resolution structural information on the transmembrane domain is lacking, but the
structure of the periplasmic domain from the EPEC secretin EscC has been reported
( Spreter et al., 2009 ); it is composed of two domains N0 and N1, also found in
secretins from unrelated systems ( Korotkov et al., 2009 ). The N1 domain possesses
a similar fold to the domains of the periplasmic regions of EscJ and PrgH. Intrigu-
ingly, domains possessing this fold were also found in other oligomeric assemblies
( Korotkov et al., 2009 ; Levdikov et al., 2012 ; Meisner et al., 2012 ), leading to the
suggestion that this fold is a common 'ring-building motif' ( Spreter et al., 2009 ).
In most T3SSs, a lipoprotein termed the pilotin promotes outer-membrane
targeting, insertion and oligomerization of the secretin component ( Crago and
Koronakis, 1998 ; Burghout et al., 2004a ). Pilotins are presumed to act through a
concerted binding of the C-terminal end of the secretin and interaction with lipids
in the membrane ( Okon et al., 2008 ; Ross and Plano, 2011 ); however, the pilotins
of different systems do not show any measurable sequence conservation, and even
their overall structure is not conserved ( Lario et al., 2005 ; Izore et al., 2011 ; Koo
et al., 2012 ). Of note, a pilotin has not yet been identified in the EPEC T3SS, and
the sequence of the corresponding secretin, EscC, appears to lack a C-terminal
pilotin-binding domain, suggesting that this protein utilizes an alternative mecha-
nism for its assembly in the outer membrane ( Gauthier et al., 2003 ).