IpaC from Shigella and SipC from Salmonella possess similar biochemical properties but are functionally distinct

Abstract

Invasion plasmid antigen C (IpaC) is secreted via the type III secretion system (TTSS) of Shigella flexneri and serves as an essential effector molecule for epithelial cell invasion. The only homologue of IpaC identified thus far is Salmonella invasion protein C (SipC/SspC), which is essential for enterocyte invasion by Salmonella typhimurium. To explore the biochemical and functional relatedness of IpaC and SipC, recombinant derivatives of both proteins were purified so that their in vitro biochemical properties could be compared. Both proteins were found to: (i) enhance the entry of wild‐type S. flexneri and S. typhimurium into cultured cells; (ii) interact with phospholipid membranes; and (iii) oligomerize in solution; however, IpaC appeared to be more efficient in carrying out several of the biochemical properties examined. Overall, the data indicate that purified IpaC and SipC are biochemically similar, although not identical with respect to their in vitro activities. To extend these observations, complementation analyses were conducted using S. flexneri SF621 and S. typhimurium SB220, neither of which is capable of invading epithelial cells because of non‐polar null mutations in ipaC and sipC respectively. Interestingly, both ipaC and sipC restored invasiveness to SB220 whereas only ipaC restored invasiveness to SF621, suggesting that SipC lacks an activity possessed by IpaC. This functional difference is not at the level of secretion because IpaC and SipC are both secreted by SF621 and it does not appear to be because of SipC dependency on this native chaperone as coexpression of sipC and sicA in SF621 still failed to restore detectable invasiveness. Taken together, the data suggest that IpaC and SipC differ in either their ability to be translocated into host cells or in their function as effectors of host cell invasion. Because IpaB shares significant sequence homology with the YopB translocator of Yersinia species, the ability for IpaC and SipC to associate with this protein was explored as a potential indicator of translocation function. Both proteins were found to bind to purified IpaB with an apparent dissociation constant in the nanomolar range, suggesting that they may differ with respect to effector function. Interestingly, whereas SB220 expressing sipC behaved like wild‐type Salmonella, in that it remained within its membrane‐bound vacuole following entry into host cells, SB220 expressing ipaC was found in the cytoplasm of host cells. This observation indicates that IpaC and SipC are responsible for a major difference in the invasion strategies of Shigella and Salmonella, that is, they escape into the host cell cytoplasm. The implications of the role of each protein's biochemistry relative to its in vivo function is discussed.

Functional Similarities between theicm/dotPathogenesis Systems ofCoxiella burnetiiandLegionella pneumophila

“…Moreover, interspecies complementation performed with the Pseudomonas aeruginosa pcrV gene and a Y. pseudotuberculosis lcrV mutant (the proteins encoded by these two genes are homologous to one another) revealed that this component determines the size of the translocation channel (22). Additional studies in which such interspecies complementation experiments were performed revealed important information about the functions of different S. flexneri and S. enterica effectors (20,43).…”

The type III secretion system apparatus determines the intracellular niche of bacterial pathogens

“…Shigella IpaB and IpaC each restored the ability of their corresponding Salmonella deletion strains to translocate effectors and invade nonphagocytic cells, but only expression of Shigella IpaC in Salmonella substantially altered the intracellular fate of this bacterium in epithelial cells and macrophages. This indicates that Shigella IpaC and Salmonella SipC confer different inherent vacuole lysis abilities to these two bacteria (6). Furthermore, enhanced phagosomal escape is not unique to Shigella IpaC, as complementation with the translocator protein CipC from another cytosolic pathogen, Chromobacterium, similarly enhanced vacuole lysis by Salmonella.…”

IpaD of Shigella flexneri Is Independently Required for Regulation of Ipa Protein Secretion and Efficient Insertion of IpaB and IpaC into Host Membranes

“…S. flexneri invasion of Henle 407 cells was monitored with a gentamicin protection assay as previously described (27,30). Henle 407 cells were seeded into 24-well plates and grown overnight to form preconfluent monolayers.…”