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Listeria monocytogenes moves rapidly through the host-cell cytoplasm by inducing directional actin assembly
G A Dabiri1, J M Sanger, D A Portnoy
1Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104.
Abstract:
Listeria monocytogenes is an intracellular parasite that can readily infect the macrophage-like cell line J774 and the kidney epithelial cell PtK2. After being ingested, the organism escapes from the phagolysosome into the host-cell cytoplasm. N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)-phallacidin, a specific stain for actin filaments (F-actin), demonstrates that within 1 hr of initiation of infection, the bacteria become surrounded by host-cell cytoplasmic actin filaments. By 3 hr, long projections of F-actin begin to form at one end of the bacteria. These actin structures colocalize with the actin-bundling protein alpha-actinin as well as with tropomyosin. Microinjection of fluorescently labeled alpha-actinin in living cells demonstrates that the formation of these F-actin projections is associated with bacterial movement, actin filaments rapidly assembling behind the bacteria as they migrate through the cytoplasm. These F-actin tails attain lengths up to 40 microns. The movement of the bacteria through the cytoplasm is rapid, 0.12-1.46 microns/sec. Within 2 min of cytochalasin D (0.5 micrograms/ml) treatment, all bacterial intracellular movement stops, and additional bacteria-associated actin assembly is blocked. A nonmotile Listeria mutant induces comparable actin assembly and moves at speeds similar to the wild type, indicating that the forces required for intracellular bacterial movement are generated by the host cell. L. monocytogenes can dramatically stimulate host-cell actin assembly in a directional manner, which serves to rapidly propel the bacteria through the cytoplasm, allowing the organisms to move to peripheral membranes and spread to uninfected cells.
Insights
Listeria monocytogenes hijacks host cell actin filaments to create propulsion tails, enabling rapid intracellular movement and cell-to-cell spread. This bacterial motility relies on host cell forces, not bacterial action.
Area of Science:
- Microbiology
- Cell Biology
- Infectious Diseases
Background:
- Listeria monocytogenes is an intracellular pathogen infecting host cells.
- Upon entry, it escapes the phagolysosome into the cytoplasm.
Purpose of the Study:
- To investigate the mechanism of intracellular bacterial movement.
- To understand the role of host cell actin in Listeria pathogenesis.
Main Methods:
- Infection of J774 and PtK2 cells with L. monocytogenes.
- Staining with N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)-phallacidin to visualize actin filaments (F-actin).
- Microinjection of fluorescently labeled alpha-actinin into living cells.
- Treatment with cytochalasin D to inhibit actin polymerization.
Main Results:
- L. monocytogenes infection induces rapid assembly of host cell F-actin behind migrating bacteria.
- These actin tails, associated with alpha-actinin and tropomyosin, reach lengths up to 40 microns.
- Bacterial movement (0.12-1.46 microns/sec) is halted by cytochalasin D, which also blocks actin assembly.
- A nonmotile Listeria mutant induces similar actin assembly, indicating host cell-generated forces.
Conclusions:
- L. monocytogenes utilizes host cell actin dynamics for rapid intracellular propulsion.
- The bacteria stimulate directional actin assembly, facilitating movement and spread to new cells.
- Host cell machinery, not the bacteria, generates the forces for intracellular migration.