In situ structure and organisation of the type IX secretion system

Abstract

The Bacteroidetes type IX secretion system (T9SS) consists of at least 19 components that translocate proteins with a type A or type B C-terminal domain (CTD) signal across the outer membrane. The overall organisation and architecture of this system including how the secretion pore (Sov) interacts with the other components is unknown. We used cryo-electron tomography to obtain the first images of the T9SS including PorK/N rings inside intact Porphyromonas gingivalis cells. Using proteomics, we identified a novel complex between Sov, PorV and PorA and showed that Sov interacts with the PorK/N rings via PorW and a new component PGN_1783. A separate complex comprising the outer membrane β barrel protein PorP, PorE, and the type B CTD protein PG1035 was also identified. Similarly, the Flavobacterium johnsoniae PorP-like protein, SprF was found bound to the major gliding motility adhesin, SprB. Based on these data, we propose cell surface anchorage for type B CTD proteins to PorP-like proteins and a unique model where the PorK/N rings function as an outer membrane barrier to maintain the close proximity of the translocon to the shuttle and 2 attachment complexes inside the rings, ensuring the harmonized secretion and cell surface attachment of the T9SS substrates.

The Type IX Secretion System: Advances in Structure, Function and Organisation

“…Although the structure of Sov is not solved, it is confidently modelled to have a very similar structure to SprA ( Figure 1 C). We have recently shown that Sov associates with either PorV or the plug in a similar manner to SprA [ 34 ] (non-peer-reviewed)and this interaction is discussed below in greater detail.…”

“…The existence of this large ring structure in the cell was questioned by Leone et al [ 37 ], where the authors suspected that the rings were an artefact of the purification method. Using electron cryotomography (ECT) of whole P. gingivalis cells, we recently showed that the rings are indeed present in cells [ 34 ]. The ring structures observed in-vivo were very similar in size and shape to the purified PorK/N rings [ 34 ].…”

“…Using electron cryotomography (ECT) of whole P. gingivalis cells, we recently showed that the rings are indeed present in cells [ 34 ]. The ring structures observed in-vivo were very similar in size and shape to the purified PorK/N rings [ 34 ]. Furthermore, these structures were absent in the porK mutant, confirming that this protein was also involved in the formation of the rings observed in-vivo [ 34 ].…”

“…The ring structures observed in-vivo were very similar in size and shape to the purified PorK/N rings [ 34 ]. Furthermore, these structures were absent in the porK mutant, confirming that this protein was also involved in the formation of the rings observed in-vivo [ 34 ]. In our in-vitro analysis, we had predicted that the rings were tethered to the outer membrane, and located in the periplasm [ 23 ].…”

“…However, it was unclear how these components interacted with the rest of the T9SS. Our recent comprehensive proteome interactome analysis of the T9SS components provided significant insights into the organisation of this novel system [ 34 ]. The Sov translocon was found to associate with the PorK/N rings via the PorW protein and another new component PGN_1783 ( Figure 3 ).…”