Cytosolic sensing of viruses

Abstract

Cells are equipped with mechanisms that allow them to rapidly detect and respond to viruses. These defense mechanisms rely partly on receptors that monitor the cytosol for the presence of atypical nucleic acids associated with virus infection. RIG-I-like receptors detect RNA molecules that are absent from the uninfected host. DNA receptors alert the cell to the abnormal presence of that nucleic acid in the cytosol. Signaling by RNA and DNA receptors results in the induction of restriction factors that prevent virus replication and establish cell-intrinsic antiviral immunity. In light of these formidable obstacles, viruses have evolved mechanisms of evasion, masking nucleic acid structures recognized by the host, sequestering themselves away from the cytosol or targeting host sensors, and signaling adaptors for deactivation or degradation. Here, we detail recent advances in the molecular understanding of cytosolic nucleic acid detection and its evasion by viruses.

Figure 1

Cell-Intrinsic Restriction and Recognition of Viruses Cells express many intrinsic antiviral restriction factors capable of blocking different stages of the virus replication cycle. They are also equiped with PRRs that detect viral PAMPs and trigger the expression of cytokines, including type I IFNs (IFN-α and IFN-β). IFNs signal via the interferon receptor (IFNAR) and upregulate the expression of hundreds of ISGs, including the antiviral factors and PRRs themselves, as well as proteins important in regulating immune responses.

Figure 2

Domain Architecture of RLRs and Model of RLR Activation (A) The three mammalian RLRs (RIG-I, MDA5, and LGP2) are superfamily 2 DExD/H-box RNA helicases. RLRs have a similar helicase core, which comprises two helicase domains termed Hel1 and Hel2, as well as an insertion domain within Hel2 known as Hel2i. At the C terminal of the Hel2 is a pincer (P) domain, also known as the bridging helices. All three RLRs also share a C-terminal domain (CTD), but only RIG-I and MDA5 have two N-terminal tandem CARDs (CARD1 and CARD2). (B) ATP-dependent conformational changes and activation of RIG-I and MDA5 occur after the recognition of stimulatory RNAs, such as base-paired 5′ PPP RNA (5′ PPP is represented by the red dot) and long dsRNA, respectively. Whereas RIG-I binds to the 5′ PPP end of base-paired RNA, MDA5 binds the stem-loop and cooperatively assembles in a head-to-tail fashion along the length of dsRNA to form a filament-like structure. K63 polyubiquitylation and oligomerization of RLRs are thought to promote downstream signaling.

Figure 3

RLR Signaling and Other Receptors Involved in Cytosolic RNA Sensing RIG-I and MDA5 signaling induces MAVS activation and oligomerization into a prion-like aggregate, which activates the TBK1 and IKK kinases. This culminates in the activation of transcription factors NF-κB, IRF-3, and IRF-7, which translocate to the nucleus and participate in the induction of antiviral genes, including those that encode IFN-α and IFN-β. Other DExD/H-box helicases, including DDX60, DHX9, DDX3, and the DDX1-DDX21-DHX36 complex, are also reported sensors of cytosolic RNA. Most of these helicases are thought to trigger IFN transcription by using RLR-dependent pathways (through RLRs themselves, MAVS, or TBK1), but the DDX1-DDX21-DHX36 complex is thought to signal through TRIF. HMGB proteins and NOD2 have also been described as receptors for cytosolic RNA and inducers of IFN responses. Lastly, it has been argued that after RNA stimulation, LRRFIP1 phosphorylates β-catenin, which translocates to the nucleus and promotes IFN-β expression. Dashed lines indicate indirect or possible signaling, and “p” indicates a phosphorylated protein.

Figure 4

Putative Intracellular DNA Sensors Involved in IFN-α and IFN-β Induction DNA present in the cytosol after viral infection induces the production of type I IFNs through a central signaling cascade involving STING, which serves as a scaffold for the phosphorylation of IRF-3 by the kinase TBK1. It was recently demonstrated that the cytosolic nucleotidyltransferase cGAS binds DNA and synthesizes the formation of a cyclic-GMP and cyclic-AMP hybrid termed cGAMP, which directly binds to and activates STING. Cytosolic DNA is reported to engage a number of additional receptors, including DAI, human IFI16 (or mouse p204), and the helicases DDX41, DHX36, and DHX9. DHX36 and DHX9 appear to be specific to CpG DNA and are reported to signal via MyD88. HMGB1, HMGB2, and HMGB3 have also been shown to promote cytosolic DNA responses. Data also suggest that ABCF1 binds DNA and interacts with HMGB2 and p204 to stimulate innate immune responses. Moreover, AT-rich DNA can be transcribed by RNA pol III into 5′-PPP-containing RNA (5′ PPP RNA), which serves as a RIG-I agonist. LRRFIP1 senses cytosolic DNA and phosphorylates β-catenin, which translocates to the nucleus and promotes IFN-β transcription. The DNA-PKc-Ku70-Ku80 and MRE11-RAD50 complexes, involved in DNA-damage responses, have additionally been suggested to bind cytosolic DNA and promote STING-dependent type I IFN responses. Ku70 is further reported to trigger the expression of type III IFNs in an IRF-1- or IRF-7-dependent manner in response to cytosolic DNA (not depicted; Zhang et al., 2011a). Proteins involved in DNA-damage responses, as well as IFI16 and RNA pol III, are abundantly present in the nucleus, highlighting the possibility that DNA sensing might also occur in that organelle (only depicted here for IFI16). Dashed lines indicate indirect or possible signaling, and “p” indicates phosphorylated proteins.