A novel role of circCPSF6 regulating antiviral innate immunity via miR-665 and PCBP2-IPS-1 axis

ObjectiveCircular RNAs are emerging as critical regulators of biological processes, yet their contribution to innate immune response during virus infection remains insufficiently defined. This study investigated the role of circCPSF6 in modulating host antiviral responses. MethodCircCPSF6 expression was validated in human and murine cells and tissue by RT-PCR and digital PCR. Its effect on virus replication was analysed by RT-PCR in human and murine primary cells and cytokine production at protein level was measured by ELISA. Luciferase assay, RNA immunoprecipitation, RNA pulldown assay, immunoblotting was performed to explore the circRNA-miRNA regulatory mechanism. CircCPSF6 interacting protein were identified by RNA pulldown followed by mass spectrometry and regulatory mechanism was analysed by immunoblotting and confocal microscopy. ResultCircCPSF6 identified as highly conserved circRNA, suppressed during virus infection, in vitro and in vivo. Functional analyses revealed antiviral role of circCPSF6. Mechanistically, circCPSF6 has dual cross-regulatory role by sequestering proviral miR-665 to relieve expression of key antiviral genes (MyD88, STAT2, IKK{varepsilon}) and interact with RNA binding protein PCBP2 to modulate IPS-1 degradation. ConclusionCircCPSF6 exert antiviral role and regulate host innate immune signaling by direct RNA-RNA and RNA-protein interaction, highlights circRNA mediated network as a potential therapeutic target in virus infection. HighlightsCircCPSF6 is evolutionarily conserved and suppressed during virus infection. CircCPSF6 act as antiviral factor by promoting inflammatory cytokine production. CircCPSF6 sponges proviral miR-665 and alleviate MyD88, STAT2, IKK{varepsilon} expression. CircCPSF6 bind to PCBP2, limit IPS-1 degradation and sustain antiviral signaling. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/686289v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@f59bedorg.highwire.dtl.DTLVardef@cff26eorg.highwire.dtl.DTLVardef@354ea5org.highwire.dtl.DTLVardef@1be4c63_HPS_FORMAT_FIGEXP M_FIG Mechanism of host innate immune regulation by circCPSF6 during virus infection. In cell cytoplasm circCPSF6 act as ceRNA for miR-665 and RNA binding protein PCBP2. Virus infection leads to reduced circCPSF6 and increased miR-665 expression, which inhibits expression of antiviral genes, MyD88, STAT2, IKK{varepsilon}. Additionally, reduced circCPSF6 leads to elevated free PCBP2 protein, hence, promote PCBP2 mediated IPS-1 degradation. Overall, these effects contribute to reduced innate immune response and increased viral replication. C_FIG