HDAC6, A Novel Cargo for Autophagic Clearance of Stress Granules, Mediates the Repression of the Type I Interferon Response During Coxsackievirus A16 Infection.

Autophagic cargoes ensure selective autophagy for the recognition and removal of various cytosolic aggregated proteins, damaged organelles, or pathogens. Stress granules (SGs), as antiviral immune complexes, serve a positive role in the type I interferon (IFN) response and can be targeted by autophagy (termed granulophagy). However, the cargo of granulophagy remains elusive, and it is still unknown whether granulophagy plays a role in viral infection. Here, we found that histone deacetylase 6 (HDAC6), a component of viral RNA-induced SGs, is a novel granulophagic cargo that is recognized by p62/Sequestosome 1 (SQSTM1) and mediates the degradation of SGs in coxsackievirus A16 (CA16)-infected cells. CA16 viral RNA activated the protein kinase RNA-activated (PKR)/eukaryotic translation initiation factor 2-alpha (eIF2α) pathway to promote SG assembly. The SGs were degraded by CA16-triggered autophagy via the interaction between the ubiquitin-associated (UBA) domain of p62 and the ubiquitin-binding domain (UBD) of HDAC6, which was bridged by a poly-ubiquitin chain. We also found that granulophagy repressed the type I interferon response and facilitated viral replication. These results suggest that HDAC6 might be the first identified granulophagic cargo and granulophagy could be a strategy that viruses apply to repress the antiviral immune response.

The Late Domain of Prototype Foamy Virus Gag Facilitates Autophagic Clearance of Stress Granules by Promoting Amphisome Formation.

Prototype foamy virus (PFV), a complex retrovirus belonging to Spumaretrovirinae, maintains lifelong latent infection. The maintenance of lifelong latent infection by viruses relies on the repression of the type I interferon (IFN) response. However, the mechanism involving PFV latency, especially regarding the suppression of the IFN response, is poorly understood. Our previous study showed that PFV promotes autophagic flux. However, the underlying mechanism and the role of PFV-induced autophagy in latent infection have not been clarified. Here, we report that the PFV viral structural protein Gag induced amphisome formation and triggered autophagic clearance of stress granules (SGs) to attenuate type I IFN production. Moreover, the late domain (L-domain) of Gag played a central role in Alix recruitment, which promoted endosomal sorting complex required for transport I (ESCRT-I) formation and amphisome accumulation by facilitating late endosome formation. Our data suggest that PFV Gag represses the host IFN response through autophagic clearance of SGs by activating the endosome-autophagy pathway. More importantly, we found a novel mechanism by which a retrovirus inhibits the SG response to repress the type I IFN response.IMPORTANCE Maintenance of lifelong latent infection for viruses relies on repression of the type I IFN response. Autophagy plays a double-edged sword in antiviral immunity. However, the role of autophagy in the regulation of the type I IFN response and the mechanism involving virus-promoted autophagy have not been fully elucidated. SGs are an immune complex associated with the antiviral immune response and are critical for type I IFN production. Autophagic clearance of SGs is one means of degradation of SGs and is associated with regulation of immunity, but the detailed mechanism remains unclear. In this article, we demonstrate that PFV Gag recruits ESCRT-I to facilitate amphisome formation. Our data also suggest that amphisome formation is a critical event for autophagic clearance of SGs and repression of the type I IFN response. More importantly, we found a novel mechanism by which a retrovirus inhibits the SG response to repress the type I IFN response.

Lnc-RP5 Regulates the miR-129-5p/Notch1/PFV Internal Promoter Axis to Promote the Expression of the Prototype Foamy Virus Transactivator Tas.

Prototype foamy virus (PFV) is a unique retrovirus that infects animals and humans and does not cause clinical symptoms. Long noncoding RNAs (lncRNAs) are believed to exert multiple regulatory functions during viral infections. Previously, we utilized RNA sequencing (RNA-seq) to characterize and identify the lncRNA lnc-RP5-1086D14.3.1-1:1 (lnc-RP5), which is markedly decreased in PFV-infected cells. However, little is known about the function of lnc-RP5 during PFV infection. In this study, we identified lnc-RP5 as a regulator of the PFV transcriptional transactivator (Tas). Lnc-RP5 enhanced the activity of the PFV internal promoter (IP). The expression of PFV Tas was found to be promoted by lnc-RP5. Moreover, miR-129-5p was found to be involved in the lnc-RP5-mediated promotion of PFV IP activity, while the Notch1 protein suppressed the activity of PFV IP and the expression of Tas. Our results demonstrate that lnc-RP5 promotes the expression of PFV Tas through the miR-129-5p/Notch1/PFV IP axis. This work provides evidence that host lncRNAs can manipulate PFV replication by employing miRNAs and proteins during an early viral infection.

The Long-Noncoding RNA lnc-NONH Enhances the Early Transcription of Prototype Foamy Virus Via Upregulating Expression of miR-34c-5p and Tas Protein.

BACKGROUND: Prototype foamy virus (PFV) is a complex and unique retrovirus with the longest genome among the retroviruses and is used as a vector for gene therapies. The viral Tas protein transactivates the viral long terminal repeat promoter and is required for viral replication. We have utilized RNA sequencing to identify and characterize the long-noncoding RNA NONHSAG000101 (lnc-NONH), which markedly increases in PFV-infected cells. However, little is known about the function of lnc-NONH. OBJECTIVES: We aim to explore the role of lnc-NONH during PFV infection. METHODS: To assess the lnc-NONH role during PFV infection, the siRNAs were used to silence the lnc-NONH expression. The microRNA (miRNA) mimic and inhibitor were employed to explore the function of lnc-NONH-related miRNA miR-34c-5p. Quantitative real-time polymerase chain reaction assay and Western blotting were applied to measure the mRNA and protein levels of PFV transactivator Tas. Luciferase assay was used to determine the transcriptional activity of the PFV unique internal promoter (IP). RESULTS: lnc-NONH promotes the expression of PFV Tas and miR-34c-5p. The interaction between lnc-NONH and miR-34c-5p enhances the transcriptional activity of the PFV IP. CONCLUSIONS: In the current study, we report a novel mechanism for the lnc-NONH-mediated upregulation of Tas expression. Our findings contribute to the understanding of regulatory network of Tas expression and PFV replication.