UC San Diego
In vitro studies of the interaction between type III secretion system effectors and potential receptors
- Author(s): Ku, Matthew
- et al.
Many Gram-negative bacterial pathogens utilize a virulence mechanism known as the type III secretion system (T3SS). The main function of the T3SS is to transport virulence factors, known as effectors, from the bacterial cytosol into the host cell. Effectors are specifically recognized by the T3SS for transport into host cells. Chaperone proteins, which bind to some effectors before transport, may aid in this specific recognition process. Although the signals that recognize effectors for transport are known, a receptor to which these signals bind has not been identified. To identify this receptor, previously observed interactions between effectors and potential receptors in bacteria were further characterized in vitro. Using in vitro co-precipitation assays, it was found that monomers of the cytoplasmic domain of YscD, an essential inner membrane ring component of the Yersinia T3SS, were insufficient to bind the effector YopH, as had previously been observed for intact YscD isolated from Yersinia. Similarly, an unassembled form of the putative C-ring YscQ was unable to bind to chaperone-effector complexes of SycE /YopE and SycH/YopH or the free chaperone, as had been suggested by data from other organisms. These results suggest binding of an effector to a receptor with an oligomeric ring structure may only occur while the oligomeric ring is assembled. To purify oligomeric rings, a method to reconstitute the intact Yersinia injectisome in vivo using E. coli is described. Furthermore, a method to identify potential receptors for the effector YopE in bacteria is also described