Periodic Reporting for period 1 - STARPAC (STING immune activation and regulation by phosphatidylinositol 4-phosphate-associated components)
Reporting period: 2021-09-01 to 2023-08-31
In healthy cells, DNA is confined to certain intracellular spaces, including the nucleus. Upon cellular damage, for example from a virus infection, bacterial infection, or tumor formation, DNA is mislocalized to the cytosol. This mislocalized DNA is a danger signal that is recognized by innate immune sensors, including the cGAS-STING pathway. Upon activation, this pathway mounts a potent immune response aimed to clear the pathogen (in case of a viral of bacterial infection) or cancerous cell. The critical role of the STING-mediated immune response against various danger signals is well-recognized, as examplified by the observation that mice lacking cGAS or STING are more sensitive to bacterial infections, tumor formation and virus infections. In addition, therapeutic targeting of STING in individuals suffering from tumours can activate the immune system and promote tumour regression. On the other hand, overactivation of the cGAS-STING pathway can lead to age-related diseases, including Alzhemer's disease, Parkinson, and other age-related chronic inflammatory conditions. Finally, individuals with mutations in STING or associated genes suffer from auto-inflammatory conditions including Aicardi-Goutières Syndrome and STING-associated vasculopathy with onset in infancy.
STING activation is a tightly regulated multistep process that is aimed to prevent unwanted activation (which could lead to auto-inflammatory diseases) but at the same time must robustly activate the immune system in case of a real threat. How STING is regulated is not well understood. A better understanding of STING regulation will provide new targets to combat a wide variety of diseases, including auto-inflammatory conditions, cancer, and ageing.
In this project, we investigated how STING is regulated by phosphatidyl inositol-4 phosphate (PI4P), a lipid present at specific locations within the cell. In addition, we investigated how manipulating PI4P with (FDA-approved) drugs can redirect the immune response by STING. Finally, we tested whether viruses that use PI4P for replication (such as rhinoviruses) can tamper with STING activation by changing the intracellular distribution of PI4P.
Conclusion of the action
We found that targeting PI4P and PI4P-associated factors can alter immune activation by affecting the intracellular distribution of STING. Furthermore, we show (FDA-approved) compounds that target PI4P can dampen or promote immune activation by STING, and we thus provide novel therapeutic targets for clinically relevant diseases.
STING activation is a tightly regulated multistep process that is aimed to prevent unwanted activation (which could lead to auto-inflammatory diseases) but at the same time must robustly activate the immune system in case of a real threat. How STING is regulated is not well understood. A better understanding of STING regulation will provide new targets to combat a wide variety of diseases, including auto-inflammatory conditions, cancer, and ageing.
In this project, we investigated how STING is regulated by phosphatidyl inositol-4 phosphate (PI4P), a lipid present at specific locations within the cell. In addition, we investigated how manipulating PI4P with (FDA-approved) drugs can redirect the immune response by STING. Finally, we tested whether viruses that use PI4P for replication (such as rhinoviruses) can tamper with STING activation by changing the intracellular distribution of PI4P.
Conclusion of the action
We found that targeting PI4P and PI4P-associated factors can alter immune activation by affecting the intracellular distribution of STING. Furthermore, we show (FDA-approved) compounds that target PI4P can dampen or promote immune activation by STING, and we thus provide novel therapeutic targets for clinically relevant diseases.
During this project, the role of PI4P and PI4P associated proteins in STING activation and regulation was systematically addressed. Cell lines were created with knockdowns and/or knockouts of PI4P-associated proteins, including ACBD3 and PI4KB, and their effect on STING activation and regulation was investigated. We showed that STING relocalizes to PI4P rich membranes of the Golgi upon activation, and this process is dependent on ACBD3. In addition, removing PI4P from the Golgi by inhibiting the PI4P kinase PI4KB or overexpression of the PI4P phosphatase Sac1 severely hampers STING activation. Conversely, increasing PI4P levels at the Golgi by blocking the lipid shuttling protein OSBP dramatically increased STING induced immune responses. The role of PI4P was further investigated using STING mutants that cannot bind PI4P. These mutants had severe defects in STING activation. Finally, we used a nanobody system to redirect STING to PI4P-rich target membranes, and found that relocalization was sufficient for STING activation, even in the absence of a stimulus. The results have been collated into a scientific manuscript an posted on the BioRxiv preprint server for dissemination. In addition, a revised version of the manuscript has recently been accepted in a peer-reviewed scientific journal.
We also investigated the effect of viral proteins that target PI4P for virus replication, including the 3A proteins of rhinoviruses. Although we did not find an effect of these proteins on STING activation, we observed that certain rhinovirus strains depend on STING for replication. This preliminary data has instigated a new research line aimed at deciphering this surprising proviral role of STING.
We also investigated the effect of viral proteins that target PI4P for virus replication, including the 3A proteins of rhinoviruses. Although we did not find an effect of these proteins on STING activation, we observed that certain rhinovirus strains depend on STING for replication. This preliminary data has instigated a new research line aimed at deciphering this surprising proviral role of STING.
This work identifies the critical role for PI4P and PI4P associated proteins in STING activation, and is a significant step forward in understanding how STING is activated and regulated. Targeting these STING regulating proteins has therapeutic potential in redirecting the innate immune response. This may be relevant to 1) reduce auto-inflammatory diseases caused by aberrant STING activation and 2) promote the immune response against viruses and tumors.
Finally, this project revealed the unexpected finding that rhinoviruses hijack the antiviral protein STING to promote rhinovirus replication. This may ultimately lead to new antiviral compounds that reduce or prevent infections with this group of clinically relevant viruses.
Finally, this project revealed the unexpected finding that rhinoviruses hijack the antiviral protein STING to promote rhinovirus replication. This may ultimately lead to new antiviral compounds that reduce or prevent infections with this group of clinically relevant viruses.