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Updated: Oct 19, 2025
Purification of Pathogen Vacuoles from Legionella-infected Phagocytes
Published on: June 19, 2012
The Legionella pneumophila Effector RavY Contributes to a Replication-Permissive Vacuolar Environment during
1Department of Microbial Pathogenesis, Boyer Center for Molecular Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
Abstract:
Legionella pneumophila is the causative agent of Legionnaires' disease and is capable of replicating inside phagocytic cells, such as mammalian macrophages. The Dot/Icm type IV secretion system is a L. pneumophila virulence factor that is essential for successful intracellular replication. During infection, L. pneumophila builds a replication-permissive vacuole by recruiting multiple host molecules and hijacking host cellular signaling pathways, a process mediated by the coordinated functions of multiple Dot/Icm effector proteins. RavY is a predicted Dot/Icm effector protein found to be important for optimal L. pneumophila replication inside host cells. Here, we demonstrate that RavY is a Dot/Icm-translocated effector protein that is dispensable for axenic replication of L. pneumophila but critical for optimal intracellular replication of the bacteria. RavY is not required for avoidance of endosomal maturation, and RavY does not contribute to the recruitment of host molecules found on replication-permissive vacuoles, such as ubiquitin, RAB1a, and RTN4. Vacuoles containing L. pneumophila ravY mutants promote intracellular survival but limit replication. The replication defect of the L. pneumophila ravY mutant was complemented when the mutant was in the same vacuole as wild-type L. pneumophila. Thus, RavY is an effector that is essential for promoting intracellular replication of L. pneumophila once the specialized vacuole has been established.
Insights
Legionella pneumophila uses the RavY effector protein to replicate inside host cells. RavY is essential for optimal intracellular bacterial growth, but not for initial vacuole formation.
Area of Science:
- Microbiology
- Cell Biology
- Infectious Diseases
Background:
- Legionella pneumophila causes Legionnaires' disease by replicating within macrophages.
- The Dot/Icm type IV secretion system delivers effector proteins essential for intracellular growth.
- Optimal intracellular replication requires coordinated action of multiple Dot/Icm effectors.
Purpose of the Study:
- To investigate the role of the predicted Dot/Icm effector RavY in L. pneumophila intracellular replication.
- To determine if RavY is translocated by the Dot/Icm system and its specific function during infection.
Main Methods:
- Demonstrated RavY translocation via the Dot/Icm system.
- Assessed L. pneumophila replication in axenic culture and within host cells (macrophages).
- Analyzed vacuole characteristics and host molecule recruitment in the presence of wild-type and ravY mutant bacteria.
Main Results:
- RavY is translocated by Dot/Icm and is dispensable for axenic growth but critical for intracellular replication.
- RavY is not involved in preventing endosomal maturation or recruiting key host molecules (ubiquitin, RAB1a, RTN4) to the vacuole.
- L. pneumophila ravY mutants survive intracellularly but exhibit significantly limited replication, which is restored in co-infections with wild-type bacteria.
Conclusions:
- RavY is a Dot/Icm effector crucial for promoting L. pneumophila replication within established replication-permissive vacuoles.
- RavY's function is specific to enhancing bacterial proliferation after the vacuole has been formed, not in its initial establishment.
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