Study of the Legionella pneumophila Dot/Icm type IV transporter and its substrate SidJ

Yancheng Liu, Purdue University

Abstract

Legionella pneumophila is a facultative intracellular pathogen that can replicate within a wide range of eukaryotic cells. Bacterial invasion and multiplication in human alveolar macrophages results in an acute and severe pneumonia called Legionnaires’ disease or a lesser severe form known as Pontiac fever. Critical for the infection process, L. pneumophila employs the Dot/Icm type IVB secretion system to translocate effector proteins into the host cell to create a specialized phagosome permissive for bacterial replication. Despite its central importance for virulence, the current model suggests that a functional Dot/Icm system is required only in the very early phase of infection. Here we constructed a Cre/loxP-based genetic system that allowed inducible deletion of specific bacterial genes after phagocytosis. Using this system, we examined the temporal requirement of the Dot/Icm secretion system during infection and demonstrated that this system is required for much longer than proposed previously. To date more than 280 L. pneumophila proteins have been identified as Dot/Icm translocated effectors. However, most of them are found to be dispensable for supporting bacterial intracellular growth. In the present study we reported the identification and characterization of a Dot/Icm effector protein SidJ. Unlike most substrates of the Dot/Icm transporter, SidJ is required for efficient bacterial replication in both macrophages and protozoans. Furthermore, loss of SidJ results in a deficient recruitment of ER proteins to the LCV, which implies a potential role of SidJ in intercepting host early secretory processes. To understand the molecular basis of SidJ function, we have investigated the interactions of SidJ with host proteins and the functional consequences of such interactions. Our studies revealed that SidJ interacts with the COPII component Sec31 and this interaction is important for the efficient intracellular replication of L. pneumophila. Moreover, SidJ can stimulate the formation of ER-derived vesicles in a process dependent on its ability to bind Sec31. Interestingly, we found that SidJ also binds to the major calcium sensor protein calmodulin during infection. The binding between SidJ and calmodulin accounts for the inhibitory effect of SidJ on yeast growth, but is not required for supporting bacterial intracellular growth. Taken together, we provided evidence showing that SidJ targets two host proteins during L. pneumophila infection. Whether these interactions contribute to related or separate processes is still an open question await further study.

Degree

Ph.D.

Advisors

Luo, Purdue University.

Subject Area

Microbiology

COinS