Role of ExoS and ExoT in inactivating Neutrophils

We study P. aeruginosa pathogenesis in a corneal model of infection. Type III secretion is crucial for establishing an infection, and we recently demonstrated that injection of two effector proteins, ExoS and ExoT, is required for preventing ROS production by infiltrating neutrophils. ExoS blocks ROS production by ADP-ribosylating Ras, which prevents activation of PI3-kinase. We are currently studying how ExoT interferes with ROS production.

Translocon function

The translocon is the structure through which effector proteins are injected into host cells. It consists of two pore-forming proteins, PopB and PopD, as well as the needle-tip (PcrV). While this structure is crucial for T3SS function, it has been very difficult to study in vitro. We have been using a genetic approach to identify protein-protein interactions in the translocon, and assigning them to translocon functions (such as translocator insertion, pore formation, and sensing of host cell contact). We recently discovered that triggering of effector secretion relies on a conformational change in the translocation pore, which is then transmitted to the needle-tip via specific protein-protein interactions with PcrV.

Effector Secretion Control

A hallmark of all type III secretion systems is that effector proteins are not exported before cell contact. In P. aeruginosa, their export is prevented by two sets of proteins, PcrG/V and the PopN complex. We have shown previously that the needle-tip protein PcrV and its export chaperone PcrG control effector secretion independently, by constraining the apparatus in an effector secretion 'off' conformation. In the case of PcrV, this is accomplished by assembling at the tip of the apparatus. PcrG, interacts with the export apparatus protein PcrD. How the second set of proteins involved in effector secretion regulation interacts with the T3SS is currently unknown. We are in the process of mapping these interactions in order to build a comprehensive model of how effector secretion is regulated.

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T3SS Model-cropped