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High-Throughput CRISPR Screens to Dissect Macrophage- Shigella Interactions

61 Pages Posted: 7 Dec 2020 Publication Status: Review Complete

See all articles by Yong Lai

Yong Lai

Singapore-MIT Alliance for Research and Technology - Antimicrobial Resistance Interdisciplinary Research Group

Liang Cui

Singapore-MIT Alliance for Research and Technology - Antimicrobial Resistance Interdisciplinary Research Group

Gregory H. Babunovic

Harvard T.H. Chan School of Public Health - Department of Immunology and Infectious Diseases

Sarah M. Fortune

Harvard University - Ragon Institute of MGH, MIT and Harvard

John G. Doench

Broad Institute

Timothy K. Lu

Singapore-MIT Alliance for Research and Technology - Antimicrobial Resistance Interdisciplinary Research Group

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Abstract

Shigellosis causes most diarrheal deaths worldwide, particularly affecting children. Shigella invades and replicates in the epithelium of the large intestine, eliciting inflammation and tissue destruction. To understand how Shigella rewires macrophages prior to epithelium invasion, we performed genome-wide and focused secondary CRISPR knockout and CRISPR interference (CRISPRi) screens in Shigella flexneri-infected human monocytic THP-1 cells. Knockdown of the Toll-like receptor 1/2 signaling pathway significantly reduced pro-inflammatory cytokine and chemokine production, enhanced host cell survival, and controlled intracellular pathogen growth. Knockdown of the enzymatic component of the mitochondrial pyruvate dehydrogenase complex enhanced THP-1 cell survival. Small molecule inhibitors blocking key components of these pathways had similar effects; these were validated with human monocyte-derived macrophages, which closely mimic the in vivo physiological state of cells post-infection. High-throughput CRISPR screens can elucidate how S. flexneri triggers inflammation and redirects host pyruvate catabolism for energy acquisition before killing macrophages, pointing to new shigellosis therapies.

Keywords: Host-pathogen interactions, shigellosis, host-directed therapy, CRISPR-Cas9 knockout screen, CRISPRi screen, macrophage, TLR1/2 signaling, pyruvate catabolism

Suggested Citation

Lai, Yong and Cui, Liang and Babunovic, Gregory H. and Fortune, Sarah M. and Doench, John G. and Lu, Timothy K., High-Throughput CRISPR Screens to Dissect Macrophage- Shigella Interactions. Available at SSRN: https://ssrn.com/abstract=3744429 or http://dx.doi.org/10.2139/ssrn.3744429
This version of the paper has not been formally peer reviewed.

Yong Lai

Singapore-MIT Alliance for Research and Technology - Antimicrobial Resistance Interdisciplinary Research Group ( email )

Singapore

Liang Cui

Singapore-MIT Alliance for Research and Technology - Antimicrobial Resistance Interdisciplinary Research Group

Singapore

Gregory H. Babunovic

Harvard T.H. Chan School of Public Health - Department of Immunology and Infectious Diseases ( email )

Cambridge, MA
United States

Sarah M. Fortune

Harvard University - Ragon Institute of MGH, MIT and Harvard ( email )

Boston, MA
United States

John G. Doench

Broad Institute ( email )

415 Main Street
Cambridge, MA 02142
United States

Timothy K. Lu (Contact Author)

Singapore-MIT Alliance for Research and Technology - Antimicrobial Resistance Interdisciplinary Research Group ( email )

Singapore

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