Study on IRF1 Transcriptional Activation Regulation of Radiation-Induced Cell Death and Inflammatory Response under Mutagenic Stress

Background

Recent multi-omics studies have revealed the key role of structural cells in immune regulation, but the mechanism remains unclear. Based on this, this study mainly explores how the transcriptional activation of interferon regulatory factor 1 (IRF1) determines the fate of structural cells and regulates communication between structural cells and immune cells when exposed to ionizing radiation, cytotoxic chemicals, and SARS-CoV-2 virus infection.

Research Source

This paper was jointly completed by Fenghao Geng, Jianhui Chen, Bin Song, and other authors from research institutions such as Sichuan University, Soochow University, and Chengdu Medical College, and will be published in the journal “Cellular & Molecular Immunology” in 2024.

Research Objective

This study aims to reveal the transcriptional activation of IRF1 in response to ionizing radiation and explore how it leads to cell death and inflammatory response. This involves mechanisms such as posttranslational modification (PTM) in the nuclear localization signal region and mutual regulation with the single-stranded DNA sensor SSBP1.

Research Process

Experimental Procedure

1. Sample Selection and Treatment: The authors exposed biological samples such as rats, primates, and humans to ionizing radiation, and detected IRF1 protein levels at different time points through immunohistochemistry (IHC).
2. RNA-seq and Single-cell RNA-seq (scRNA-seq): High-throughput sequencing was used to determine IRF1 expression in different cell types in rat skin.
3. Experimental Observation and Analysis: Different cell lines were subjected to ionizing radiation and other stimuli to study IRF1 nuclear translocation and transcriptional activity, including Luciferase reporter assay, co-immunoprecipitation (Co-IP), and other experiments.
4. Mass Spectrometry Analysis (MS): Mass spectrometry analysis was performed on the posttranslational modifications of IRF1.
5. Gene Knockout and Mutant Modeling: IRF1 gene knockout mice and IRF1 mutants were used to study its active molecular mechanism.
6. ssbp1 and cgas/Sting Pathway: Analyzed mtDNA leakage and its impact on the IRF1 signaling pathway.

Main Experimental Results

1. Changes in IRF1 Expression: IHC results showed that IRF1 expression was upregulated after irradiation, visible in different skin samples (rats, primates, etc.), and its expression changes in different cell types were confirmed in scRNA-seq data.
2. Nuclear Translocation and Transcriptional Activity: Rapid nuclear translocation of IRF1 and increased transcriptional activity after irradiation were observed through Luciferase assay and immunofluorescence staining.
3. Role of Posttranslational Modifications: Mass spectrometry analysis found that phosphorylation and acetylation in the nuclear localization signal region of IRF1 significantly regulate its transcriptional activity.
4. Inhibitory Effect of ssbp1: The transcriptional activity of IRF1 is significantly inhibited by the single-stranded DNA binding protein SSBP1, promoting its nuclear translocation.
5. Regulation of Cell Fate: IRF1 affects the proliferation, migration, and senescence of skin cells through nuclear translocation and posttranslational modifications. Especially under high-dose radiation, IRF1 overexpression significantly increased cell mortality.
6. Radiation-Induced Inflammatory Response: The transcriptional activation of IRF1 is associated with the infiltration of inflammatory cells in skin tissue. IRF1 gene knockout mice showed significantly reduced skin radiation damage.
7. Drug Intervention Experiments: Through molecular screening and structural simulation, two small molecule inhibitors were found to significantly inhibit IRF1 transcriptional activity, and their effects on alleviating radiation-induced inflammation and cell death were validated in in vivo and in vitro experiments.

Main Conclusions

This study revealed the activation pathway of IRF1 in structural cells under genotoxic stress. The research found that IRF1 senses mtDNA leakage and initiates the cgas/Sting signaling pathway, thereby triggering inflammatory cell death and radioactive skin damage. This not only provides a new perspective for understanding the role of IRF1 in radiation-induced inflammatory responses but also suggests the possibility of IRF1 as a potential therapeutic target.

Significance and Value of the Research

- This study not only advances the in-depth understanding of the role of structural cells in immune responses but also provides a theoretical basis for developing new therapeutic strategies. IRF1 inhibitors show great application potential, especially in radioactive skin damage and infectious diseases (such as COVID-19).

Research Highlights

• Discovery of Nuclear Translocation & Posttranslational Modifications: This study first revealed the key role of phosphorylation and acetylation modifications on the nuclear localization signal of IRF1 under radiation stress.
• Role of ssbp1: Proposed a new mechanism of IRF1 inhibition in structural cells, indicating that ssbp1 as an IRF1 repressor protein can significantly affect IRF1 activity and cell fate.
• Drug Screening and Application: Through molecular simulation screening, two novel small molecule IRF1 inhibitors were discovered, which can effectively prevent radiation-induced cell death and skin inflammation.

Through these achievements, the research not only provides new targets and therapeutic approaches for radiation-induced inflammatory diseases but also opens up a new understanding of the role of structural cells in immune regulation and stress response.