SummaryIn wild-type Salmonella, the length of the¯agellar hook, a structure consisting of subunits of the hook protein FlgE, is fairly tightly controlled at < 55 nm. Because¯iK mutants produce abnormally elongated hook structures that lack the ®lament structure, FliK appears to be involved in both the termination of hook elongation and the initiation of ®lament formation. FliK, a soluble protein, is believed to function together with a membrane protein, FlhB, of the export apparatus to mediate the switching of export substrate speci®city (from hook protein to¯agellin) upon completion of hook assembly. We have examined the location of FliK during¯agellar morphogenesis. FliK was found in the culture supernatants from the wild-type strain and from¯gD (hook capping protein),¯gE (hook protein) and¯gK (hook-®lament junction protein) mutants, but not in that from a¯gB (rod protein) mutant. The amount of FliK in the culture supernatant from the¯gE mutant was much higher than in that from the¯gK mutant, indicating that FliK is most ef®ciently exported prior to the completion of hook assembly. Export was impaired by deletions within the N-terminal region of FliK, but not by C-terminal truncations. A decrease in the level of exported FliK resulted in elongated hook structures, sometimes with ®laments attached. Our results suggest that the export of FliK during hook assembly is important for hook-length control and the switching of export substrate speci®city.

Enteropathogenic Escherichia coli and enterohemorrhagic E. coli are diarrheagenic bacterial human pathogens that cause severe gastroenteritis. These enteric pathotypes, together with the mouse pathogen Citrobacter rodentium, belong to the family of attaching and effacing pathogens that form a distinctive histological lesion in the intestinal epithelium. The virulence of these bacteria depends on a type III secretion system (T3SS), which mediates the translocation of effector proteins from the bacterial cytosol into the infected cells. The core architecture of the T3SS consists of a multi-ring basal body embedded in the bacterial membranes, a periplasmic inner rod, a transmembrane export apparatus in the inner membrane, and cytosolic components including an ATPase complex and the C-ring. In addition, two distinct hollow appendages are assembled on the extracellular face of the basal body creating a channel for protein secretion: an approximately 23 nm needle, and a filament that extends up to 600 nm. This filamentous structure allows these pathogens to get through the host cells mucus barrier. Upon contact with the target cell, a translocation pore is assembled in the host membrane through which the effector proteins are injected. Assembly of the T3SS is strictly regulated to ensure proper timing of substrate secretion. The different type III substrates coexist in the bacterial cytoplasm, and their hierarchical secretion is determined by specialized chaperones in coordination with two molecular switches and the so-called sorting platform. In this review, we present recent advances in the understanding of the T3SS in attaching and effacing pathogens.

IntroductionAlmost all the bacterial flagellum lies beyond the cell membrane and, therefore, its protein subunits have to be exported. With only a few exceptions (FlgA, FlgH and FlgI), all of which are associated with the basal body outer-ring structure, the proteins are exported by a specialized pathway that is a member of a large family, called type III secretion systems (TTS systems) (Hueck, 1998). The components of the flagellar export apparatus have been identified (Minamino and Macnab, 1999). They consist of six integral membrane proteins (FlhA, FlhB, FliO, FliP, FliQ and FliR) and various soluble components (FliI, FliH, the general chaperone FliJ and the substratespecific chaperones FlgN, FliS and FliT (Yokoseki and Kutsukake, 1994;Fraser et al., 1999;Minamino et al., 2000;Auvray et al., 2001).With the exception of the flagellar system, TTS systems are used for the secretion of virulence factors by a wide range of pathogenic bacteria. As well as the striking similarity in visual appearance between the flagellar basal body and its homologous virulence structure, the needle complex (Kubori et al., 1998;Tamano et al., 2000;Blocker et al., 2001), there are a number of proteins that are clearly conserved.One example is the ATPase that provides the energy source for export or secretion. In the flagellar system, this is named FliI. Its virulence homologues carry various names in different species; in Salmonella, for example, it is called InvC. The enzymology of Salmonella FliI has been characterized in detail (Dreyfus et al., 1993; Fan and Molecular Microbiology (2002)

Sex steroid hormones play important physiological roles in reproductive and nonreproductive tissues, including immune cells. These hormones exert their functions by binding to either specific intracellular receptors that act as ligand-dependent transcription factors or membrane receptors that stimulate several signal transduction pathways. The elevated susceptibility of males to bacterial infections can be related to the usually lower immune responses presented in males as compared to females. This dimorphic sex difference is mainly due to the differential modulation of the immune system by sex steroid hormones through the control of proinflammatory and anti-inflammatory cytokines expression, as well as Toll-like receptors (TLRs) expression and antibody production. Besides, sex hormones can also affect the metabolism, growth, or virulence of pathogenic bacteria. In turn, pathogenic, microbiota, and environmental bacteria are able to metabolize and degrade steroid hormones and their related compounds. All these data suggest that sex steroid hormones play a key role in the modulation of bacterial-host interactions.

Salmonella FliI is the ATPase that drives flagellar protein export. It normally exists as a complex together with the regulatory protein FliH. A fliH null mutant was slightly motile, with overproduction of FliI resulting in substantial improvement of its motility. Mutations in the cytoplasmic domains of FlhA and FlhB, which are integral membrane components of the type III flagellar export apparatus, also resulted in substantially improved motility, even at normal FliI levels. Thus, FliH, though undoubtedly important, is not essential.Bacterial flagellar assembly begins with the basal body, followed by the hook and finally the filament (11). Almost all of the external components are translocated into a central channel in the nascent structure by an export apparatus that is believed to be located within an annular pore in the basal body MS ring (5,8,21). The export apparatus consists of six integral membrane components (FlhA, FlhB, FliO, FliP, FliQ, and FliR) and three cytoplasmic components (FliH, FliI, and FliJ) (15,16). Export of bacterial flagellar proteins has characteristics in common with type III secretion of virulence factors by pathogenic bacteria (1,7,10,11).FliI is a flagellum-specific ATPase, which converts the energy of ATP hydrolysis into the energy for flagellar protein export (4, 16-18, 20, 23). FliH is a regulatory protein that is thought to prevent FliI from hydrolyzing ATP until the energy can be used for export (2,6,13,16). FliJ functions as a general chaperone that prevents export substrates from premature aggregation in the cytoplasm (12).A tentative model for the flagellar export process has been proposed (16,17,26) in which a (FliH) 2 /FliI heterotrimeric complex (16), FliJ, and substrate diffuse to the cytoplasmic domains of FlhA and FlhB and form a complex. ATP hydrolysis by FliI drives export substrate translocation through the export apparatus, placing the substrate in the channel from whence it diffuses to its final assembly destination.In the present study, we have found that overproduction of FliI and also certain second-site mutations in flhA and flhB are capable of conferring considerable export capability and motility to a fliH null mutant.Bacterial strains and plasmids used in this study are listed in Table 1 and shown graphically in Fig. 1. Luria-Bertani broth, soft tryptone-agar plates, and M9-Casamino Acids medium plus 1% glycerol were prepared as previously described (15). Effects of overproduction of FliI and FliJ on motility and flagellar protein export of a fliH null mutant.A Salmonella fliH null mutant MKM11 (6) has the sequence MSNEL-⌬226-PGVL, i.e., it contains only nine amino acid residues of FliH (Fig. 1A). It displays a leaky motile phenotype, indicating that substrate export still occurs with low probability in the absence of FliH. We transformed the null mutant with plasmids encoding the soluble export components FliH, FliI, and FliJ and examined the transformants for swarming ability ( Fig. 2A). FliH complemented the mutant. FliI overproduced from a pTrc99A-ba...

Quorum sensing (QS) is a communication mechanism between bacteria that allows specific processes to be controlled, such as biofilm formation, virulence factor expression, production of secondary metabolites and stress adaptation mechanisms such as bacterial competition systems including secretion systems (SS). These SS have an important role in bacterial communication. SS are ubiquitous; they are present in both Gram-negative and Gram-positive bacteria and in Mycobacterium sp. To date, 8 types of SS have been described (T1SS, T2SS, T3SS, T4SS, T5SS, T6SS, T7SS, and T9SS). They have global functions such as the transport of proteases, lipases, adhesins, heme-binding proteins, and amidases, and specific functions such as the synthesis of proteins in host cells, adaptation to the environment, the secretion of effectors to establish an infectious niche, transfer, absorption and release of DNA, translocation of effector proteins or DNA and autotransporter secretion. All of these functions can contribute to virulence and pathogenesis. In this review, we describe the known types of SS and discuss the ones that have been shown to be regulated by QS. Due to the large amount of information about this topic in some pathogens, we focus mainly on Pseudomonas aeruginosa and Vibrio spp.

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