Periodic Reporting for period 1 - TIRI (Role of the kinase TAOK2 in the innate immune response to viral infection)
Période du rapport: 2023-09-01 au 2025-08-31
Viral infections constitute a major threat to public health with a significant impact on life expectancy, quality of life, and the economy. One of the first warning signs of viral infections is the presence of viral nucleic acids within the cell. Pattern recognition receptors (PRR), specialized molecules that bind to invading viral RNA, trigger the innate immune response, culminating in the production of cytokines and chemokines and the expression of protective antiviral proteins. A dysbalanced innate immune response renders patients highly susceptible to virus infections or, conversely, leads to overactive immunity associated with autoimmune disease.
We previously identified the protein kinase TAOK2 as a new player in the detection of viral RNA. In the absence of TAOK2, cells are less able to fight viral infection, and the induction of the innate immune response is compromised. In this project, I will define how TAOK2 recognizes viral RNA and how that affects the processes leading to the induction of the innate immune response. This will shed light on how the innate immune response, which needs to be kept in a delicate balance to fight off viral infection without overreacting to benign signals, is modulated and fine-tuned.
We previously identified the protein kinase TAOK2 as a new player in the detection of viral RNA. In the absence of TAOK2, cells are less able to fight viral infection, and the induction of the innate immune response is compromised. In this project, I will define how TAOK2 recognizes viral RNA and how that affects the processes leading to the induction of the innate immune response. This will shed light on how the innate immune response, which needs to be kept in a delicate balance to fight off viral infection without overreacting to benign signals, is modulated and fine-tuned.
To understand how RNA associates with TAOK2 and how it stimulates kinase activity, human TAOK2 and the highly similar homologue dTao from Drosophila melanogaster were purified from Sf9 insect cells. The binding specificity to different RNA species was determined by microscale thermophoresis and electromobility shift assays, and the region of TAOK2 required for binding to RNA was determined. The structure of full-length TAOK2 was determined by cryo-electron microscopy. To determine how TAOK2 affects cellular signaling events during viral infection by phosphorylating downstream targets, we determined the phosphorylation targets of TAOK2 in infected cells by phosphoproteomics, the interaction partners of TAOK2 by AP-MS, and differentially regulated genes in infected TAOK2+/+ and TAOK2 / cells by proteome analysis. To lay the groundwork for studying the effect of TAOK2 during viral infection in vivo, we tested the susceptibility of primary mouse embryonic fibroblasts (MEFs) from wild-type and Taok2 / - mice to various virus families and showed that depletion of TAOK2 in these cells increased the replication of various viruses.
The work in this project sheds light on the role of TAOK2 during antiviral signaling, specifically how TAOK2 modulates the signaling events in response to PRR recognition of viral RNA. It also expanded the breadth of virus families that appear to be restricted by TAOK2 substantially and therefore points towards a general importance of TAOK2 in infection with various viruses. Furthermore, the structural and biochemical characterization of RNA binding to TAOK2 will be instrumental in defining novel concepts of RNA binding, as TAOK2 does not have a previously defined or predicted RNA-binding domain.