Yersinia controls type III effector delivery into host cells by modulating Rho activity

Area of Science:

• Microbiology
• Cell Biology
• Molecular Biology

Background:

• Yersinia pseudotuberculosis utilizes type III secretion system proteins YopB and YopD to form pores and translocate Yop effectors into host cells.
• YopE and YopT are Yersinia effectors that inhibit Rho GTPases, and their absence leads to increased pore formation.

Purpose of the Study:

• To investigate the role of Rho GTPase signaling in modulating Yersinia pseudotuberculosis pore formation and effector translocation.
• To elucidate the mechanisms by which Yersinia pseudotuberculosis controls host cell actin dynamics for successful infection.

Main Methods:

• Infection of HeLa cells with Yersinia pseudotuberculosis strains, including effector mutants.
• Treatment of infected cells with Clostridium difficile toxin B and Clostridium botulinum C3 to inactivate Rho GTPases.
• Use of specific inhibitors (NSC23766) and dominant-negative constructs to modulate Rac activity.
• Analysis of pore formation using membrane impermeable dyes and YopE translocation.

Main Results:

• Inhibition of actin polymerization and inactivation of Rho, Rac, and Cdc42 significantly reduced pore formation and YopE translocation.
• While Rac inhibition affected bacterial uptake, it did not impede pore formation or translocation.
• Rho GTPases (Rho A, B, and/or C) were implicated in controlling Yop delivery.
• Beta1 integrin signaling, including Src kinase activity, was crucial for YopE translocation.
• Yersinia pseudotuberculosis infection activated Rho through YopB and beta1 integrin engagement.

Conclusions:

• Rho GTPase activation, triggered by the YopB/YopD translocon and beta1 integrin signaling, is essential for actin polymerization and Yop translocation.
• YopE and YopT activity likely terminates Yop delivery and prevents pore formation after translocation.