High-throughput screening of E3 ubiquitin ligases identifies TRIM48 as a novel negative regulator of RIG-I signaling
The retinoic acid-inducible gene-I (RIG-I) signaling is crucial for cell-intrinsic innate antiviral immunity. Upon cytosolic detection of virus-associated RNA, it triggers a cascade inducing production of potent cytokines, mainly type I and III interferons (IFNs). While effective, dysregulated respo...
Saved in:
| Main Authors: | , , , , , , , , , , , , , |
|---|---|
| Format: | Article (Journal) |
| Language: | English |
| Published: |
October 2025
|
| In: |
Cellular signalling
Year: 2025, Volume: 134, Pages: 1-18 |
| ISSN: | 1873-3913 |
| DOI: | 10.1016/j.cellsig.2025.111973 |
| Online Access: | Verlag, kostenfrei, Volltext: https://doi.org/10.1016/j.cellsig.2025.111973 Verlag, kostenfrei, Volltext: https://www.sciencedirect.com/science/article/pii/S0898656825003882 
|
| Author Notes: | Guandi Wu, Jamie Frankish, Joschka Willemsen, Dominik Ricken, Jonas Becker, Darius Schweinoch, Jürgen Beneke, Sandra Wüst, Nina Beil, Petr Matula, Karl Rohr, Holger Erfle, Lars Kaderali, Marco Binder |
| Summary: | The retinoic acid-inducible gene-I (RIG-I) signaling is crucial for cell-intrinsic innate antiviral immunity. Upon cytosolic detection of virus-associated RNA, it triggers a cascade inducing production of potent cytokines, mainly type I and III interferons (IFNs). While effective, dysregulated responses can harm the host, requiring tight pathway control. Here, we performed a comprehensive, systematic siRNA-based high-throughput screen across 616 established and putative E3 ubiquitin ligases for their impact on RIG-I signaling. We employed a fluorescence-based live-cell imaging assay in A549 cells to monitor nuclear translocation of IRF3 and NF-κB, two key transcription factors downstream of RIG-I. Candidate genes were validated in an orthogonal secondary screen, assessing their impact on the functional antiviral response to a Rift Valley Fever reporter virus. Fourteen hits showed consistent effects on RIG-I signaling across both screens. These genes were further validated and characterized by assessing IFN-β promoter reporter activity and IFNB1 mRNA levels upon dsRNA transfection. TRIM48 emerged as a highly robust negative regulator. Overexpression of TRIM48 suppressed RIG-I-mediated activation of IRF3 and NF-κB, reduced IFN and IFN-stimulated gene expression, and enhanced viral replication. Conversely, TRIM48 deficiency enhanced RIG-I signaling and inhibited viral replication. Notably, TRIM48 acts as an induced feedback regulator upon infection, and its effect depended on its enzymatic ubiquitin ligase activity. Our high-throughput screen provides an unbiased assessment of close to all E3 ubiquitin ligases for their regulatory effect in RIG-I signaling, and identified several interesting candidates for further investigation. TRIM48 was established as a negative feedback regulator of the RIG-I pathway. |
|---|---|
| Item Description: | Online verfügbar 1 July 2025, Version des Artikels 3 July 2025 Gesehen am 21.11.2025 |
| Physical Description: | Online Resource |
| ISSN: | 1873-3913 |
| DOI: | 10.1016/j.cellsig.2025.111973 |