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Fig. 1. Type 1 pilus architecture and crystal structure of the FimH adhesin. (Left) High-resolution EM reveals the composite structure of the type 1 pilus. The pilus tip containing the FimH adhesin is indicated. (Right) FimH has two domains, each with Ig-like folds. The adhesin domain has a binding pocket (arrow) that can accommodate D-mannose.

cells, also known as umbrella cells, deposit on their apical surfaces a quasi-crystalline array of hexagonal complexes comprised of four integral membrane proteins known as uroplakins (18). The uroplakins (UPIa, UPIb, UPII, and UPIII), which are highly conserved among all mammalian species, are assembled into 16-nm-wide hexagonal particles that are further organized into plaques 0.3– 0.5 µm in diameter that cover almost the entire luminal surface of the bladder. All of the uroplakins have large luminal domains and, with the exception of UPIII, relatively small cytoplasmic domains. In cross section, the luminal leaflet of the apical plasma membrane appears twice as thick as the cytoplasmic leaflet. This uroplakin-embedded membrane has been termed the AUM (asymmetric unit membrane) and it is proposed to serve as a permeability barrier and to strengthen and stabilize facet cells, preventing rupturing as the bladder distends to accommodate more urine. In vitro binding assays have shown that two of the uroplakins, UPIa and UPIb, derived from diverse mammalian species, including mice and humans, can specifically bind to E. coli expressing type 1 pili (19). Binding is inhibited by the soluble FimH receptor analogue, D-mannose, and by enzymatic deglycosylation of UPIa and UPIb.

Fig. 2. Type 1 pilus-mediated bacterial attachment to the bladder epithelium. After inoculation of C57BL/6 mice with type 1-piliated UPEC, numerous bacteria (yellow) can be found attached to the luminal surface of the bladder (blue) as detected by scanning EM (A) and high-resolution freeze-dry/deep-etch EM (B). Type 1 pili mediating bacterial attachment were resolved with the high-resolution technique. The scalloped appearance of the bladder surface is attributable to the presence of the uroplakin plaques (≈0.5 µm in diameter). [Bars = 3 µm (A) and 0.5 µm (B).]

Interactions between type 1 pili and host receptor molecules such as UPIa and UPIb allow UPEC to establish an initial foothold within the urinary tract. The ability of type 1 pili to mediate bacterial attachment to the AUM of the facet cells lining the bladder lumen has been investigated by using a mouse cystitis model and microscopic techniques (13). Shortly after inoculation of UPEC into mouse bladders, numerous bacteria can be found attached to the urothelial surface as detected by scanning electron microscopy (EM) (Fig. 2A). Bacteria adhered to the bladder surface both singly and, occasionally, in large biofilmlike colonies. Bacteria were often situated in grooves and niches formed by the AUM of the facet cells (13). Almost no bacteria were found associated with bladders taken from mice infected with mutant E. coli strains that express type 1 pili lacking the FimH adhesin. High-resolution freeze-dry and freeze-fracture/deep-etch EM revealed that the FimH-containing adhesive tips of type 1 pili could interact directly with the uroplakin-embedded AUM, seemingly tethering the bacteria to the bladder surface (Fig. 2B). Type 1 pili also appeared able to interact with other adherent bacteria on the bladder surface. This is an interesting observation considering that type 1 pili have been shown to function in the formation of biofilms in vitro (20).

Type 1 Pilus-Mediated Bacterial Invasion

Uropathogenic strains of E. coli, contrary to prevailing assumptions, are not strictly extracellular pathogens. Over 10 years ago, transmission EM studies of infected rat and mouse bladders demonstrated that bladder epithelial cells can internalize UPEC in vivo (21, 22). Bacteria were observed within membrane-bound vacuoles and free within the cytoplasm of the facet cells that line the luminal surface of the bladder. It was proposed that the bladder epithelial cells internalized bacteria as part of an innate host defense mechanism. A more recent study, however, suggested that bacterial internalization by bladder epithelial cells could benefit the pathogens (13). It was shown by using a murine



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