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High-Throughput CRISPR Screens To Dissect Macrophage-Shigella Interactions
Yong Lai1,2,3, Liang Cui1, Gregory H Babunovic4
1Antimicrobial Resistance Interdisciplinary Research Group, Singapore-MIT Alliance for Research and Technology, Singapore.
Mbio
|December 22, 2021
Summary
Shigella infection rewires host macrophages by manipulating the Toll-like receptor 1/2 pathway and pyruvate metabolism. Targeting these pathways offers new therapeutic strategies for shigellosis, a growing public health concern.
Area of Science:
- Infectious Diseases
- Immunology
- Genetics
Background:
- Shigellosis, a leading cause of diarrheal deaths globally, particularly impacts children.
- Shigella infection involves invasion and replication within the intestinal epithelium, causing inflammation and tissue damage.
- Understanding host-pathogen interactions in macrophages is crucial for developing novel shigellosis interventions, especially with rising antibiotic resistance.
Purpose of the Study:
- To investigate how Shigella flexneri manipulates host macrophages before invading the intestinal epithelium.
- To identify host pathways that can be modulated to enhance the immune response against Shigella infection.
- To discover potential therapeutic targets for shigellosis by deciphering macrophage-Shigella interactions.
Main Methods:
- Genome-wide and focused secondary CRISPR knockout and CRISPR interference (CRISPRi) screens were performed in Shigella flexneri-infected human monocytic THP-1 cells.
- The Toll-like receptor 1/2 (TLR1/2) signaling pathway and the mitochondrial pyruvate dehydrogenase complex were investigated.
- Small-molecule inhibitors were used to block key pathway components, and findings were validated in human monocyte-derived macrophages.
Main Results:
- Knockdown of the TLR1/2 signaling pathway significantly reduced pro-inflammatory cytokine and chemokine production, improved host cell survival, and controlled intracellular pathogen growth.
- Disrupting the mitochondrial pyruvate dehydrogenase complex enhanced host cell survival.
- Inhibiting these pathways demonstrated similar protective effects, validated in primary human macrophages.
Conclusions:
- Shigella flexneri manipulates host pyruvate catabolism for energy acquisition and triggers inflammation by hijacking the TLR1/2 pathway.
- Targeting TLR1/2 signaling and pyruvate metabolism presents promising host-directed therapeutic strategies for shigellosis.
- High-throughput CRISPR screening is effective in uncovering pathogen manipulation mechanisms and identifying new treatment avenues for infectious diseases.
