Extracellular UDP and P2Y₆ function as a danger signal to protect mice from vesicular stomatitis virus infection through an increase in IFN-β production

Extracellular nucleotides that constitute a "danger signal" play an important role in the regulation of immune responses. However, the function and mechanism of extracellular UDP and P2Y₆ in antiviral immunity remain unknown. In this study, we demonstrated the in vitro and in vivo protection of UDP/P2Y₆ signaling in vesicular stomatitis virus (VSV) infection. First, we demonstrated that VSV-infected cells secrete UDP from the cytoplasm as a danger signal to arouse surrounding cells. Meanwhile, expression of the UDP-specific receptor P2Y₆ also was enhanced by VSV. Consequently, UDP protects RAW 264.7 cells, murine embryonic fibroblasts, bone marrow-derived macrophages, and L929 cells from VSV and GFP lentivirus infection. This protection can be blocked by the P2Y₆ selective antagonist MRS2578 or IFN-α/β receptor-blocking Ab. VSV-induced cell death and virus replication were both enhanced significantly by knocking down and knocking out P2Y₆ in different cells. Mechanistically, UDP facilitates IFN-β secretion through the p38/JNK- and ATF-2/c-Jun-signaling pathways, which are crucial in promoting antiviral immunity. Interestingly, UDP was released through a caspase-cleaved pannexin-1 channel in VSV-induced apoptotic cells and protected cells from infection through P2Y₆ receptor in an autocrine or paracrine manner. Furthermore, UDP also protected mice from VSV infection through P2Y₆ receptors in an acute neurotropic infection mouse model. Taken together, these results demonstrate the important role of extracellular UDP and P2Y₆ as a danger signal in antiviral immune responses and suggest a potential therapeutic role for UDP/P2Y₆ in preventing and controlling viral diseases.