# Cytosolic Immunostimulatory DNA Ligands and DNA Damage Activate the Integrated Stress Response, Stress Granule Formation, and Cytokine Production

## TL;DR

Cytosolic DNA activates stress responses and cytokine production through a STING-PERK-G3BP1 signaling pathway, linking DNA sensing to immune and stress reactions.

## Contribution

Identifies a novel signaling axis (STING-PERK-G3BP1) connecting cytosolic DNA sensing to stress granule formation and immune response.

## Key findings

- Cytosolic dsDNA activates the integrated stress response and induces stress granules and cytokine production.
- STING-PERK-G3BP1 signaling couples DNA sensing with stress and innate immune pathways.
- STING regulates ROS production in response to DNA damage, showing crosstalk with oxidative stress pathways.

## Abstract

What are the main findings?
The presence of cytosolic dsDNA, exogenous or endogenous, activates the integrated stress response pathway, inducing stress granules and cytokine production.The STING-PERK-G3BP1 signaling axis couples DNA sensing with stress and innate immune response pathways.

The presence of cytosolic dsDNA, exogenous or endogenous, activates the integrated stress response pathway, inducing stress granules and cytokine production.

The STING-PERK-G3BP1 signaling axis couples DNA sensing with stress and innate immune response pathways.

What are the implications of the main findings?
The cGAS-STING pathway is a valuable target in cancer immunotherapy and pathogen invasion. Combining DNA-damaging agents like radiation or chemotherapy agents with STING activators can enhance cellular response and improve the effectiveness of immunotherapy and host immune response.Understanding the modes of STING activation and its role as a central signaling hub that fine-tunes the cell’s response depending on the specific type of cellular threat or stress has implications for developing therapies for diverse pathologies.

The cGAS-STING pathway is a valuable target in cancer immunotherapy and pathogen invasion. Combining DNA-damaging agents like radiation or chemotherapy agents with STING activators can enhance cellular response and improve the effectiveness of immunotherapy and host immune response.

Understanding the modes of STING activation and its role as a central signaling hub that fine-tunes the cell’s response depending on the specific type of cellular threat or stress has implications for developing therapies for diverse pathologies.

The presence of aberrant double-stranded DNA (dsDNA) in the cytoplasm of cells is sensed by unique pattern recognition receptors (PRRs) to trigger innate immune response. The cyclic GMP–AMP synthase (cGAS)–stimulator of interferon genes (STING) signaling pathway is activated by the presence of non-self or mislocalized self-dsDNA from nucleus or mitochondria released in response to DNA damage or cellular stress in the cytoplasm. Activation of cGAS leads to the synthesis of the second messenger cyclic GMP–AMP (cGAMP), which binds and activates STING, triggering downstream signaling cascades that result in the production of type I interferons (IFNs) and proinflammatory cytokines. Here, we show that diverse immunostimulatory dsDNA ligands and chemotherapy agents like Doxorubicin and Taxol trigger the integrated stress response (ISR) by activating endoplasmic reticulum (ER) stress kinase, protein kinase RNA-like ER kinase (PERK), in addition to the canonical IFN pathways. PERK-mediated phosphorylation and inactivation of the alpha subunit of eukaryotic translation initiation factor-2 (eIF2α) result in the formation of stress granules (SGs). SG formation by dsDNA was significantly reduced in PERK knockout cells or by inhibiting PERK activity. Transcriptional induction of IFNβ and cytokines, ISR signaling, and SG formation by dsDNA was dampened in cells lacking PERK activity, STING, or key stress-granule nucleating protein, Ras-GAP SH3 domain-binding protein 1 (G3BP1), demonstrating an important role of the signal transduction pathway mediated by STING and SG assembly. Lastly, STING regulates reactive oxygen species (ROS) production in response to DNA damage, highlighting the crosstalk between DNA sensing and oxidative stress pathways. Together, our data identify STING–PERK–G3BP1 signaling axis that couples cytosolic DNA sensing to stress response pathways in maintaining cellular homeostasis.