The type III secretion injectisome is a complex nanomachine that allows bacteria to deliver protein effectors across eukaryotic cellular membranes. In recent years, significant progress has been made in our understanding of its structure, assembly and mode of operation. The principal structural components of the injectisome, from the base located in the bacterial cytosol to the tip of the needle protruding from the cell surface, have been investigated in detail. The structures of several constituent proteins were solved at the atomic level and important insights into the assembly process have been gained. However, despite the ongoing concerted efforts of molecular and structural biologists, the role of many of the constituent components of this nanomachine remain unknown.

Type III secretion systems allow Yersinia spp., Salmonella spp., Shigella spp., Bordetella spp., and Pseudomonas aeruginosa and enteropathogenic Escherichia coli adhering at the surface of a eukaryotic cell to inject bacterial proteins across the two bacterial membranes and the eukaryotic cell membrane to destroy or subvert the target cell. These systems consist of a secretion apparatus, made of approximately 25 proteins, and an array of proteins released by this apparatus. Some of these released proteins are "effectors," which are delivered into the cytosol of the target cell, whereas the others are "translocators," which help the effectors to cross the membrane of the eukaryotic cell. Most of the effectors act on the cytoskeleton or on intracellular-signaling cascades. A protein injected by the enteropathogenic E. coli serves as a membrane receptor for the docking of the bacterium itself at the surface of the cell. Type III secretion systems also occur in plant pathogens where they are involved both in causing disease in susceptible hosts and in eliciting the so-called hypersensitive response in resistant or nonhost plants. They consist of 15-20 Hrp proteins building a secretion apparatus and two groups of effectors: harpins and avirulence proteins. Harpins are presumably secreted in the extracellular compartment, whereas avirulence proteins are thought to be targeted into plant cells. Although a coherent picture is clearly emerging, basic questions remain to be answered. In particular, little is known about how the type III apparatus fits together to deliver proteins in animal cells. It is even more mysterious for plant cells where a thick wall has to be crossed. In spite of these haunting questions, type III secretion appears as a fascinating trans-kingdom communication device.

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Pathogenic bacteria of the genus Yersinia release in vitro a set of antihost proteins called Yops. Upon infection of cultured epithelial cells, extracellular Yersinia pseudotuberculosis transfers YopE across the host cell plasma membrane. To facilitate the study of this translocation process, we constructed a recombinant Yersinia enterocolitica strain producing YopE fused to a reporter enzyme. As a reporter, we selected the calmodulin-dependent adenylate cyclase of Bordetella pertussis and we monitored the accumulation of cyclic AMP (cAMP). Since bacteria do not produce calmodulin, cyclase activity marks the presence of hybrid enzyme in the cytoplasmic compartment of the eukaryotic cell. Infection of a monolayer of HeLa cells by the recombinant Y. enterocolitica strain led to a significant increase of cAMP. This phenomenon was dependent not only on the integrity of the Yop secretion pathway but also on the presence of YopB and/or YopD. It also required the presence of the adhesin YadA at the bacterial surface. In contrast, the phenomenon was not affected by cytochalasin D, indicating that internalization of the bacteria themselves was not required for the translocation process. Our results demonstrate that Y. enterocolitica is able to transfer hybrid proteins into eukaryotic cells. This system can be used not only to study the mechanism of YopE translocation but also the fate of the other Yops or even of proteins secreted by other bacterial pathogens.

Pathogenic yersiniae secrete a set of antihost proteins, called Yops, by a type III secretion mechanism.

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Size determination represents a fundamental requirement for multicomponent biological structures. Some pathogenic bacteria possess a weapon derived from the flagellum. Like the flagellum, this type-III secretion apparatus, called the injectisome, has a transmembrane basal body, but the external component is a needle-like structure instead of a hook and a filament. Here, we provide evidence that the length of this needle is determined by the size of a protein, YscP, acting as a molecular ruler.

Many pathogenic bacteria use injectisomes to deliver effector proteins into host cells through type III secretion. Injectisomes consist of a basal body embedded in the bacterial membranes and a needle. In Yersinia, translocation of effectors requires the YopB and YopD proteins, which form a pore in the target cell membrane, and the LcrV protein, which assists the assembly of the pore. Here we report that LcrV forms a distinct structure at the tip of the needle, the tip complex. This unique localization of LcrV may explain its crucial role in the translocation process and its efficacy as the main protective antigen against plague.