The Needle Component of the Type III Secreton of Shigella Regulates the Activity of the Secretion Apparatus

Abstract

Gram-negative bacteria commonly interact with eukaryotic host cells by using type III secretion systems (TTSSs or secretons). TTSSs serve to transfer bacterial proteins into host cells. Two translocators, IpaB and IpaC, are first inserted with the aid of IpaD by Shigella into the host cell membrane. Then at least two supplementary effectors of cell invasion, IpaA and IpgD, are transferred into the host cytoplasm. How TTSSs are induced to secrete is unknown, but their activation appears to require direct contact of the external distal tip of the apparatus with the host cell. The extracellular domain of the TTSS is a hollow needle protruding 60 nm beyond the bacterial surface. The monomeric unit of the Shigella flexneri needle, MxiH, forms a superhelical assembly. To probe the role of the needle in the activation of the TTSS for secretion, we examined the structurefunction relationship of MxiH by mutagenesis. Most point mutations led to normal needle assembly, but some led to polymerization or possible length control defects. In other mutants, secretion was constitutively turned "on." In a further set, it was "constitutively on" but experimentally "uninducible." Finally, upon induction of secretion, some mutants released only the translocators and not the effectors. Most types of mutants were defective in interactions with host cells. Together, these data indicate that the needle directly controls the activity of the TTSS and suggest that it may be used to "sense" host cells.Shigella flexneri causes bacillary dysentery in humans, a disease characterized by invasion of, massive inflammation in, and destruction of the colonic mucosa. The genes required for S. flexneri invasion are clustered on a 31-kb fragment of a large virulence plasmid (1). Within this region, the mxi/spa operons encode a type III secretion system (TTSS 5 or secreton), and the ipa/ipg operons encode five effector proteins abundantly secreted early in invasion, IpaA to -D and IpgD.Type III secretons are essential determinants of the interaction of many Gram-negative bacteria with animal or plant hosts, and they translocate bacterial proteins into eukaryotic host cells to manipulate them during infection. TTS apparatuses are encoded by ϳ25 genes (2), nearly all being essential for function. These devices perform regulated post-translational and co-translational protein translocation across three biological membranes (two from the bacterium and one from the host cell). In Shigella, IpaB to -D are essential for invasion and give rise to the TTSS translocon (3, 4) through which at least two other proteins, IpaA and IpgD, are thought to be translocated. Sequence similarities exist between components of TTSSs and those of the prokaryotic flagellar assembly machinery (5). TTSSs and flagella share all but host cell contact-mediated TTSS induction and the ability to translocate proteins into eukaryotic cells. 10 TTSS proteins are similar in sequence and/or membrane topology to the cytoplasmic or inner membrane proteins of flagellar hook-basal bodies (6, 7)...

Bioinformatic and Biochemical Evidence for the Identification of the Type III Secretion System Needle Protein ofChlamydia trachomatis

“…1C). CdsF does lack the P-(S/D)-(D/N)-P turn, a four-residue ordered-loop region conserved among many other characterized needle proteins (12,30,61). However, this conservation is not complete, and the motif is only partially present, for example, in EscF of E. coli, PscF of Pseudomonas spp., and SsaH from Salmonella SPI-2 (63).…”

“…The precise mechanism by which this occurs has not yet been elucidated; however, it has been postulated that a conformational change in the helical arrangement of the needle may be responsible (30). Hence, the detection of CdsF trimers or dimers could reflect an alteration in helical assembly.…”

“…CHLAMYDIA T3SS NEEDLE PROTEIN 1687 on April 7, 2019 by guest http://jb.asm.org/ "activation signal" upon host cell contact (30,60). The precise mechanism by which this occurs has not yet been elucidated; however, it has been postulated that a conformational change in the helical arrangement of the needle may be responsible (30).…”

“…Experiments using a subcutaneous murine infection model have shown that the Yersinia pestis needle protein YscF can serve as a protective antigen, indicating that the needle may have potential in vaccine development (8,35,45,54,55). In addition to its role as an injectisome component, evidence suggests that the needle is also important in host cell detection and thus regulation of T3S (12,30,60). Electron micrographs of the needles from Yersinia, Salmonella Spi-1, and Shigella, comprising the small homologous proteins YscF, PrgI, and MxiH, respectively, reveal straight rigid structures that are very short (approximately 40 to 80 nm) compared to flagella (7,25,34,55).…”

Molecular model of a type III secretion system needle: Implications for host-cell sensing

“…However, T3SSs (and not flagellar systems) undergo host-contact-mediated activation and translocation of effector proteins into host cells, apparently through direct contact of the external distal tip of the apparatus with the host cell (7)(8)(9). The role of the needle has been probed by using mutagenic studies of the needle-subunit proteins from Yersinia and Shigella (10,11). In these systems, point mutations were identified that led to constitutive secretion of effectors and experimentally uninducible needles.…”