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Myosin light chain kinase plays an essential role in S. flexneri dissemination
M Rathman1, P de Lanerolle, H Ohayon
1Unité de Pathogénie Microbienne Moléculaire, INSERM U389 and Station Centrale de Microscopie Electronique, Institut Pasteur, 75724 Paris Cedex 15, France.
Abstract:
Shigella flexneri, the causitive agent of bacillary dysentery, has been shown to disseminate in colonic epithelial cells via protrusions that extend from infected cells and are endocytosed by adjacent cells. This phenomenon occurs in the region of the eukaryotic cell's adherens junctions and is inhibited by pharmacological reagents or host cell mutations that completely disrupt the junctional complex. In this study, inhibitors of the myosin light chain kinase (MLCK) were shown to dramatically decrease intercellular spread of S. flexneri but to have no inhibitory effect on bacterial entry, multiplication or actin-based motility within the host cell. Furthermore, cell-to-cell spread of Listeria monocytogenes, another bacterial pathogen that uses an actin-based mechanism to move within the eukaryotic cytoplasm and to spread from cell to cell, was not affected by the MLCK inhibitors, indicating that (1) the inhibition of S. flexneri cell-to-cell spread in treated cells is not due to a complete break down of cell-cell contacts, which was subsequently confirmed by confocal microscopy, and (2) MLCK plays a role in a S. flexneri-specific mechanism of dissemination. Myosin has been shown to play a role in a variety of membrane-based phenomena. The work presented here suggests that activation of this molecule via phosphorylation by MLCK, at the very least participates in the formation of the bacteria-containing protrusion, and could also contribute to the endocytosis of this structure by neighboring cells.
Insights
Shigella flexneri spreads between cells using protrusions, a process inhibited by myosin light chain kinase (MLCK) inhibitors. This specific mechanism highlights MLCK
Area of Science:
- Microbiology
- Cell Biology
- Pathogen-Host Interactions
Background:
- Shigella flexneri causes bacillary dysentery and spreads between colonic epithelial cells via protrusions.
- This intercellular dissemination occurs near adherens junctions and is sensitive to junctional complex disruption.
Purpose of the Study:
- To investigate the role of myosin light chain kinase (MLCK) in the intercellular spread of Shigella flexneri.
- To determine if MLCK is involved in bacterial entry, intracellular motility, or cell-to-cell dissemination.
Main Methods:
- Treatment of infected cells with myosin light chain kinase (MLCK) inhibitors.
- Assessment of bacterial entry, intracellular multiplication, and actin-based motility.
- Evaluation of intercellular spread of S. flexneri and Listeria monocytogenes.
- Confocal microscopy to confirm the integrity of cell-cell contacts.
Main Results:
- MLCK inhibitors significantly reduced intercellular spread of S. flexneri.
- Inhibitors did not affect bacterial entry, multiplication, or actin-based motility within host cells.
- MLCK inhibitors did not inhibit the cell-to-cell spread of Listeria monocytogenes.
- Confocal microscopy confirmed that cell-cell contacts remained intact in treated cells.
Conclusions:
- MLCK plays a crucial role in the Shigella flexneri-specific mechanism of cell-to-cell dissemination.
- MLCK activity is involved in the formation of bacteria-containing protrusions and potentially their subsequent endocytosis by adjacent cells.
- The findings differentiate the spread mechanism of S. flexneri from that of Listeria monocytogenes, which is not affected by MLCK inhibition.