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Evaluation of Caspase Activation to Assess Innate Immune Cell Death
Published on: January 20, 2023
Inflammatory cell death, PANoptosis, screen identifies host factors in coronavirus innate immune response as
R K Subbarao Malireddi1, Ratnakar R Bynigeri1, Raghvendra Mall1,2
1Department of Immunology, St. Jude Children's Research Hospital, Memphis, TN, 38105, USA.
Abstract:
The COVID-19 pandemic, caused by the β-coronavirus (β-CoV) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), continues to cause significant global morbidity and mortality. While vaccines have reduced the overall number of severe infections, there remains an incomplete understanding of viral entry and innate immune activation, which can drive pathology. Innate immune responses characterized by positive feedback between cell death and cytokine release can amplify the inflammatory cytokine storm during β-CoV-mediated infection to drive pathology. Therefore, there remains an unmet need to understand innate immune processes in response to β-CoV infections to identify therapeutic strategies. To address this gap, here we used an MHV model and developed a whole genome CRISPR-Cas9 screening approach to elucidate host molecules required for β-CoV infection and inflammatory cell death, PANoptosis, in macrophages, a sentinel innate immune cell. Our screen was validated through the identification of the known MHV receptor Ceacam1 as the top hit, and its deletion significantly reduced viral replication due to loss of viral entry, resulting in a downstream reduction in MHV-induced cell death. Moreover, this screen identified several other host factors required for MHV infection-induced macrophage cell death. Overall, these findings demonstrate the feasibility and power of using genome-wide PANoptosis screens in macrophage cell lines to accelerate the discovery of key host factors in innate immune processes and suggest new targets for therapeutic development to prevent β-CoV-induced pathology.
Insights
This study used CRISPR screening in macrophages to identify host factors involved in coronavirus infection and PANoptosis, a form of inflammatory cell death. Discovering these factors can lead to new therapies against severe coronavirus diseases.
Area of Science:
- Immunology
- Virology
- Genetics
Background:
- Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes significant global health issues, with incomplete understanding of viral entry and innate immune responses.
- Innate immune responses, including cytokine storms and cell death, amplify pathology during beta-coronavirus (β-CoV) infections.
- Therapeutic strategies require a deeper understanding of host-pathogen interactions and innate immune processes in β-CoV infections.
Purpose of the Study:
- To elucidate host molecules critical for β-CoV infection and PANoptosis (inflammatory cell death) in macrophages.
- To develop and validate a whole-genome CRISPR-Cas9 screening approach for identifying host factors in innate immunity.
Main Methods:
- Utilized a mouse hepatitis virus (MHV) model and a whole-genome CRISPR-Cas9 screen in macrophage cell lines.
- Performed genome-wide screening to identify host genes influencing viral entry and PANoptosis.
- Validated findings by assessing the role of identified host factors, including the known MHV receptor Ceacam1.
Main Results:
- The screen successfully identified Ceacam1, the MHV receptor, as a key factor for viral entry and subsequent cell death.
- Deletion of Ceacam1 significantly reduced viral replication and MHV-induced macrophage cell death.
- Identified several novel host factors essential for MHV infection-induced macrophage cell death.
Conclusions:
- Genome-wide PANoptosis screens in macrophages are effective for discovering host factors in innate immune responses.
- The findings highlight the feasibility of using such screens to accelerate the identification of therapeutic targets for β-CoV-induced pathology.
- Suggests new avenues for developing therapeutics to mitigate severe outcomes of coronavirus infections.
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