Cellular hnRNP D promotes influenza A virus replication by inhibiting TBK1-IRF3-mediated innate immune response

Heterogeneous nuclear ribonucleoproteins (hnRNPs) play important roles in the life cycle of influenza A virus (IAV). Our previous mass spectrometry analysis identified cellular hnRNP D as a novel interaction partner of the IAV polymerase basic 2 (PB2) protein. However, the functional implications of hnRNP D in IAV replication and the underlying mechanisms remained unknown. In this study, we confirmed that hnRNP D directly interacts with the PB2 protein of the A/Puerto Rico/8/1934 (PR8, H1N1) strain, while also binding to other proteins within viral ribonucleoprotein complexes (vRNPs). These interactions collectively inhibit vRNPs assembly and viral polymerase activity. However, we have found that hnRNP D enhances the viral titer of IAV in A549 cells. Mechanistically, hnRNP D suppresses the activation of the interferon (IFN)-β promoter, and the mRNA levels of downstream factors in the type I IFN signaling pathway. In detail, hnRNP D inhibits IFN-β promoter activation induced by crucial antiviral proteins and interacts strongly with the interferon regulatory factor 3 (IRF3). More importantly, hnRNP D blocks the binding of TANK-binding kinase 1 (TBK1) to IRF3, thereby impeding the phosphorylation and activation of IRF3. Collectively, we unveil a novel viral immune evasion strategy by which IAV hijacks a host RNA-binding protein, hnRNP D, to facilitate self-replication. This work not only elucidates the intricate trade-off mechanisms in virus-host interaction but also identifies hnRNP D as a potential therapeutic target aimed at bolstering antiviral immunity.

Influenza viruses are serious zoonotic pathogens, causing millions of severe infections and hundreds of thousands of deaths annually. The hnRNP family is known to influence IAV replication and pathogenesis. Here, we demonstrate for the first time that hnRNP D functions as a positive factor for IAV replication. We show that hnRNP D exerts a net promoting effect on viral replication primarily by suppressing the type I interferon response. The mechanism of action involves viral infection upregulating hnRNP D, which interacts with the key transcription factor IRF3, disrupting the TBK1-IRF3 signaling axis. Our findings provide novel insights into how a host RNA-binding protein can be co-opted by IAV to dampen antiviral innate immunity, presenting a potential target for developing new therapeutic strategies against influenza.