In this research, we have established a drug screening method based on the autophagy signal pathway using the bimolecular fluorescence complementation - fluorescence resonance energy transfer (BiFC-FRET) technique to develop novel anti-influenza A virus (IAV) drugs. We selected Evodia rutaecarpa Benth out of 83 examples of traditional Chinese medicine and explored the mechanisms of evodiamine, the major active component of Evodia rutaecarpa Benth, on anti-IAV activity. Our results showed that evodiamine could significantly inhibit IAV replication, as determined by a plaque inhibition assay, an IAV vRNA promoter luciferase reporter assay and the Sulforhodamine B method using cytopathic effect (CPE) reduction. Additionally, evodiamine could significantly inhibit the accumulation of LC3-II and p62, and the dot-like aggregation of EGFP-LC3. This compound also inhibited the formation of the Atg5-Atg12/Atg16 heterotrimer, the expressions of Atg5, Atg7 and Atg12, and the cytokine release of TNF-α, IL-1β, IL-6 and IL-8 after IAV infection. Evodiamine inhibited IAV-induced autophagy was also dependent on its action on the AMPK/TSC2/mTOR signal pathway. In conclusion, we have established a new drug screening method, and selected evodiamine as a promising anti-IAV compound.

The nucleolus is a stress sensor associated with cell cycle progression and a central hub for the replication of pathogenic RNA viruses. However, the role of nucleolus in influenza A virus infection has not been well studied. Here we show that the interaction between NS1 protein of influenza A/Shantou/602/06 (H3N2) and nucleolin, a ubiquitous protein of nucleolus repressed RNA Pol I-dependent transcription via establishing hyper-methylation in the UCE of rRNA gene promoter. NS1 expressed cells showed significant association of ribosomal proteins with MDM2, and p53 accumulation, suggesting induced nucleolar stress. Disruption of the interaction of NS1 with nucleolin or overexpression of nucleolin in NS1 expressed cells revived RNA Pol I-dependent transcription, indicating nucleolin could be one target for NS1 to repress rRNA synthesis of host cells. Our present study suggests that NS1 protein of H3N2 could induce nucleolar stress based on epigenetic alteration of rRNA gene promoter via interaction with nucleolin.

Background: Coxsackievirus commonly infects children and occasionally causes severe meningitis and/or encephalitis in the newborn. The underlying mechanism(s) behind the central nervous system pathology is poorly defined. Methods: It is hypothesized that astrocytes may be involved in inflammatory response induced by CVB3 infection. Here we discuss this hypothesis in the context of CVB3 infection and associated inflammatory response in primary mouse astrocytes. Results: The results showed that coxsackievirus receptor (CAR) was distributed homogeneously on the astrocytes, and that CVB3 could infect and replicate in astrocytes, with release of infectious virus particles. CVB3 induced cytopathic effect and production of proinflammatory cytokines IL-1β, TNF-α, IL-6, and chemokine CXCL10 from astrocytes. Conclusion: These data suggest that direct astrocyte damage and cytokines induction could be a mechanism of virus-induced meningitis and/or encephalitis.