The RNA binding protein Quaking represses host interferon response by downregulating MAVS

Abstract

Quaking (QKI) is an RNA-binding protein (RBP) involved in multiple aspects of RNA metabolism and many biological processes. Despite a known immune function in regulating monocyte differentiation and inflammatory responses, the degree to which QKI regulates the host interferon (IFN) response remains poorly characterized. Here we show that QKI ablation enhances poly(I:C) and viral infection-induced IFNβ transcription. Characterization of IFN-related signalling cascades reveals that QKI knockout results in higher levels of IRF3 phosphorylation. Interestingly, complementation with QKI-5 isoform alone is sufficient to rescue this phenotype and reduce IRF3 phosphorylation. Further analysis shows that MAVS, but not RIG-I or MDA5, is robustly upregulated in the absence of QKI, suggesting that QKI downregulates MAVS and thus represses the host IFN response. As expected, MAVS depletion reduces IFNβ activation and knockout of MAVS in the QKI knockout cells completely abolishes IFNβ induction. Consistently, ectopic expression of RIG-I activates stronger IFNβ induction via MAVS-IRF3 pathway in the absence of QKI. Collectively, these findings demonstrate a novel role for QKI in negatively regulating host IFN response by reducing MAVS levels.

The RNA binding protein Quaking represses splicing of the Fibronectin EDA exon and downregulates the interferon response

“…Among GSEA gene sets that significantly correlated with gene expression changes caused by QKI KO in the absence of poly(I:C) transfection, the second highest scoring was the MOSERLE_IFNA_RESPONSE gene set ( 43 ) (normalized enrichment score of 2.42 with FDR q -val = 0.002; Figure 1B , Supplementary Table S2 ). This analysis implied that QKI represses the expression of many genes induced by type I IFNs, which is consistent with our previous report that QKI suppresses IFN induction ( 26 ).…”

“…To investigate the mechanisms by which QKI regulates host immunity, we performed genome scale transcriptome profiling in parental (WT) and QKI knock-out (QKO#3) HuH7 cells ( 26 ) activated by poly(I:C) transfection (Figure 1A ). RNA-seq data, which had an average depth of 75M reads per sample, were analyzed to identify differential expression and processing of transcripts.…”

“…HuH7 and A549 cells were maintained in Dulbecco's modified Eagle's medium (DMEM, Gibco) supplemented with 10% fetal bovine serum (FBS), 1% penicillin–streptomycin (Pen Strep, Gibco), and 1% HEPES (Gibco) in a 37°C humidified incubator with 5% CO 2 . QKI knock-out cell lines (HuH7 QKO#3, HuH7 QKO#13, HuH7 QKO#14 and A549 QKO#1) were generated using CRISPR–Cas9 technology as previously described ( 26 ). Guide sequences and editing events are listed in Supplementary Table S3 .…”

Definition of germ layer cell lineage alternative splicing programs reveals a critical role for Quaking in specifying cardiac cell fate

“…HuH7 cells were transfected using Lipofectamine2000 (ThermoFisher), according to the manufacturer's instructions. NKX2.5 GFP hESCs were co-transfected with pX458-sgQKI as described (Liao et al, 2020) or control empty vector along with pCEP-puro which contains a puromycin-resistance gene. One day later puromycin was added at 1 μg/ml and cells were selected for 3 days, and then expanded in the absence of puromycin.…”

Characterization and functional interrogation of SARS-CoV-2 RNA interactome

“…Interestingly, ILF3 was shown to participate to antiviral type I interferon response by promoting the expression of IFNB1 and interferon-stimulated gene (ISG) during host translational shutoff (Watson et al, 2020). QKI was shown to repress host interferon response by downregulating MAVS expression at a post-transcriptional level (Liao et al, 2020). This suggests that SARS-CoV2 has evolved several mechanisms to dampen the type I interferon response by targeting specific RBPs.…”