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Activation of Ran GTPase by a Legionella effector promotes microtubule polymerization, pathogen vacuole motility and infection.
- Source :
-
PLoS pathogens [PLoS Pathog] 2013 Sep; Vol. 9 (9), pp. e1003598. Date of Electronic Publication: 2013 Sep 19. - Publication Year :
- 2013
-
Abstract
- The causative agent of Legionnaires' disease, Legionella pneumophila, uses the Icm/Dot type IV secretion system (T4SS) to form in phagocytes a distinct "Legionella-containing vacuole" (LCV), which intercepts endosomal and secretory vesicle trafficking. Proteomics revealed the presence of the small GTPase Ran and its effector RanBP1 on purified LCVs. Here we validate that Ran and RanBP1 localize to LCVs and promote intracellular growth of L. pneumophila. Moreover, the L. pneumophila protein LegG1, which contains putative RCC1 Ran guanine nucleotide exchange factor (GEF) domains, accumulates on LCVs in an Icm/Dot-dependent manner. L. pneumophila wild-type bacteria, but not strains lacking LegG1 or a functional Icm/Dot T4SS, activate Ran on LCVs, while purified LegG1 produces active Ran(GTP) in cell lysates. L. pneumophila lacking legG1 is compromised for intracellular growth in macrophages and amoebae, yet is as cytotoxic as the wild-type strain. A downstream effect of LegG1 is to stabilize microtubules, as revealed by conventional and stimulated emission depletion (STED) fluorescence microscopy, subcellular fractionation and Western blot, or by microbial microinjection through the T3SS of a Yersinia strain lacking endogenous effectors. Real-time fluorescence imaging indicates that LCVs harboring wild-type L. pneumophila rapidly move along microtubules, while LCVs harboring ΔlegG1 mutant bacteria are stalled. Together, our results demonstrate that Ran activation and RanBP1 promote LCV formation, and the Icm/Dot substrate LegG1 functions as a bacterial Ran activator, which localizes to LCVs and promotes microtubule stabilization, LCV motility as well as intracellular replication of L. pneumophila.
- Subjects :
- Animals
Bacterial Proteins genetics
Cell Line
Enzyme Activation
GTPase-Activating Proteins antagonists & inhibitors
GTPase-Activating Proteins genetics
Gene Silencing
Humans
Legionella pneumophila genetics
Legionella pneumophila immunology
Legionella pneumophila ultrastructure
Legionnaires' Disease immunology
Legionnaires' Disease metabolism
Legionnaires' Disease microbiology
Legionnaires' Disease pathology
Macrophages immunology
Macrophages metabolism
Macrophages ultrastructure
Mice
Microtubule Proteins chemistry
Microtubule Proteins metabolism
Microtubules ultrastructure
Mutation
Phagocytosis
Phagosomes enzymology
Phagosomes ultrastructure
Polymerization
Protein Stability
Protein Transport
Virus Replication
ran GTP-Binding Protein antagonists & inhibitors
ran GTP-Binding Protein genetics
Bacterial Proteins metabolism
GTPase-Activating Proteins metabolism
Legionella pneumophila physiology
Macrophages microbiology
Microtubules metabolism
Phagosomes metabolism
ran GTP-Binding Protein metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1553-7374
- Volume :
- 9
- Issue :
- 9
- Database :
- MEDLINE
- Journal :
- PLoS pathogens
- Publication Type :
- Academic Journal
- Accession number :
- 24068924
- Full Text :
- https://doi.org/10.1371/journal.ppat.1003598