Negative Regulation of K63 Polyubiquitination by the Immune Regulator LGP2

Lenoir, Jessica

doi:https://doi.org/10.21985/n2-fhbn-s688

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Negative Regulation of K63 Polyubiquitination by the Immune Regulator LGP2

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RNA viruses cause a wide range of diseases that range from mild respiratory illness to fatal hemorrhagic fevers. To combat these pathogens, most cells in the human body scan the cell for viruses and activate a potent cell-intrinsic immune response by producing interferon (IFN), a cytokine that amplifies the innate response and activates the adaptive immune system. One pathway that leads to IFN production is the RIG-I like receptor (RLR) pathway. Antiviral sensor proteins, including RIG-I, MDA5, and LGP2, in the cytoplasm recognize features of non-self RNAs that trigger a cascade of signal transduction pathways, leading to IFN production. Sensor-RNA interactions by RIG-I and MDA5 initiate the oligomerization of a mitochondrial signaling platform known as MAVS. This signaling platform recruits and binds to ubiquitin ligase proteins known as TRAF proteins. TRAF proteins form polyubiquitin chains which recruit kinases important for activating the downstream signaling pathway, inducing transcription of IFN and other antiviral mediators important to inhibit infection and to prevent cytotoxicity. To prevent cytotoxicity, the antiviral signaling pathway encodes for negative regulators which target fundamental steps within the pathway. One negative regulator includes the viral sensor protein LGP2, which contains a C-terminal domain and DExD/H box helicase domain like RIG-I and MDA5, but no CARDs. Because of its lack of CARDs, LGP2 has been hypothesized to be a negative regulator of the RLR pathway. Many studies have reported that overexpression of LGP2 negatively regulates IFN signaling. Conversely, knocking out LGP2 in primary mouse embryonic fibroblasts lead to prolonged induction of the IFN response and enhanced synthesis of antiviral genes after synthetic dsRNA stimulation. Many different mechanisms have been proposed for LGP2 as a negative regulator. Recent results in our lab indicate that LGP2 can bind to TRAF proteins and inhibits their downstream signaling activity. We have identified that LGP2 interferes with TRAF’s ubiquitin ligase activity by preventing the formation of polyubiquitin chains to inhibit antiviral signaling, without the need of a binding site on TRAF6. TRAF6 was unable to interact with its E2 conjugating enzyme Ubc13/UBE2N in the presence of LGP2. LGP2 prevents K63 polyubiquitination by preventing interaction between the E3 ligase and Ubc13/UBE2N. Using co-immunoprecipitation experiments, we determined that Ubc13/UBE2N interacts with LGP2, and this interaction is important in the inhibition of polyubiquitin chain formation. These findings establish a mechanism that LGP2 uses to inhibit K63 polyubiquitination of E3 ubiquitin ligases involved in the antiviral response.

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