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A Visual Assay to Monitor T6SS-mediated Bacterial Competition
Published on: March 20, 2013
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Vibrio cholerae type 6 secretion system effector trafficking in target bacterial cells
Brian T Ho1, Yang Fu1, Tao G Dong2
1Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115.
Summary
The type 6 secretion system (T6SS) delivers toxic proteins to kill bacteria. Researchers engineered a Vibrio cholerae T6SS to deliver a nuclease, demonstrating its versatility in targeting periplasmic or cytosolic substrates.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- The type 6 secretion system (T6SS) is a critical virulence factor in many Gram-negative bacteria, facilitating interbacterial competition by delivering toxic effector proteins.
- VgrG proteins are essential components of the T6SS, often possessing C-terminal extensions with diverse enzymatic activities that target prey cells.
- Vibrio cholerae VgrG3 normally targets peptidoglycan in the periplasm; this study investigates modifying its effector domain.
Purpose of the Study:
- To engineer a novel T6SS effector by replacing the VgrG3 hydrolase domain with a nuclease domain from Salmonella enterica.
- To assess the capability of the modified Vibrio cholerae T6SS to deliver effectors targeting different subcellular compartments (periplasm and cytosol) in prey bacteria.
- To investigate the mechanism of effector trafficking for VgrG3 and TseL when expressed in the bacterial cytosol.
Main Methods:
- Genetic engineering of Vibrio cholerae to replace the VgrG3 C-terminal hydrolase domain with a nuclease domain from Salmonella enterica.
- Assessing the interbacterial killing activity of the engineered V. cholerae strain using its T6SS.
- Analyzing the subcellular localization of T6SS effectors (VgrG3 and TseL) when expressed in the bacterial cytosol.
Main Results:
- The engineered V. cholerae strain, equipped with the T6SS delivering the nuclease domain, successfully killed its parent strain.
- This demonstrates the V. cholerae T6SS's ability to deliver effectors targeting both periplasmic and cytosolic substrates.
- Effectors VgrG3 and TseL were found to reach the periplasm even when expressed in the cytosol, independent of classical secretion signals.
Conclusions:
- The T6SS is a versatile delivery system capable of targeting diverse substrates within prey bacteria.
- Effector proteins can exhibit non-classical trafficking pathways to reach their intended targets.
- These findings suggest an evolutionary adaptation enabling T6SS effectors to function effectively regardless of their delivery compartment.
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