The Sde phosphoribosyl-linked ubiquitin transferases exploit reticulons to protect the integrity of the Legionella-containing vacuole

Legionella pneumophila grows within host cells by forming a specialized membrane-bound compartment via the Icm/Dot type IV secretion system (T4SS). T4SS translocated Sde proteins promote phosphoribosyl-linked ubiquitination (pR-Ub) of several mammalian cell proteins, including Rtn4. In response to modification, Rtn4 forms tubular ER aggregates around the Legionella-containing vacuole (LCV). The loss of sde together with sdhA results in severe vacuole disruption at early infection timepoints. We tested if Rtn4 modification allowed it to serve as a physical barrier to protect its degradation from host-derived disruptive compartments. The challenge of the rtn4-/- macrophages with {Delta}sdhA partially phenocopied the challenge of rtn4+/+ with {Delta}sde{Delta}sdhA, indicating that Rtn4 plays a role in protecting LCV integrity. Depletion of rtn1 in rtn4-/- macrophages potentiated vacuole permeability when compared to rtn4-/- macrophages, consistent with Sde targeting multiple reticulon family members to support vacuole integrity. These results indicate that L. pneumophila exploits Rtn4 in cooperation with Rtn1 to establish a replication niche by promoting Sde-mediated tubular ER aggregates, arguing that these proteins function as a physical barrier during early steps of LCV biogenesis.