Luteolin inhibits herpes simplex virus 1 infection by activating cyclic guanosine monophosphate-adenosine monophosphate synthase-mediated antiviral innate immunity

•Luteolin effectively inhibits the post-entry process of HSV-1 infection.•Luteolin activates the cytoplasmic DNA-sensing pathway cGAS-STING-IRF3 to promote antiviral type I interferon production in vitro and in vivo.•Luteolin binds directly to the active substrate binding site of cGAS and enhances cGAS oligomerization.•Luteolin significantly ameliorates HSV-1-induced HSE in mice, including weight loss, neurodegeneration and neuroinflammation. The successive outbreaks of large-scale infectious diseases due to virus infection have been a major threat to human health in recent decades. Herpes simplex virus I (HSV-1) is a widely-disseminated DNA virus that infects the central nervous system to cause herpes labialis, keratitis and herpes simplex virus encephalitis (HSE), resulting in recurrent lifelong clinical or subclinical episodes. Luteolin is a plant flavone that has been extensively used in the treatment of various human diseases, including carcinogenesis, inflammation and chronic degenerative diseases. The aim of this study was to investigate the antiviral molecular mechanism of luteolin against HSV-1 infection in vitro and in vivo. The antiviral effect of luteolin in cell lines was examined by viral plaque assay, RT-qPCR, Western blot and time-of-addition assay. The interaction between luteolin and cyclic guanosine monophosphate-adenosine monophosphate synthase (cGAS) was evaluated by molecular modeling and semi-denaturing detergent agarose gel electrophoresis. The efficacy of luteolin on HSE was evaluated in the HSE mouse model by analyzing weight loss, neurodegenerative symptoms and histopathological scores. Cytokine expression and protein levels were examined by RT-qPCR, Western blot and ELISA. Luteolin inhibited the early process of HSV-1 infection, without affecting the infection of acyclovir-resistant HSV-1 strains. In addition, luteolin enhanced antiviral type I interferon production and activated the cytoplasmic DNA-sensing cGAS-stimulator of interferon gene (STING) pathway. Luteolin directly bound the active substrate binding site and promoted the oligomerization of cGAS. Luteolin also inhibited HSE-related weight loss, neurodegeneration and neuroinflammation in mice caused by HSV-1 infection. Furthermore, luteolin enhanced type I interferon expression and stimulated the cGAS-STING signaling pathway in vivo. Luteolin inhibited the post-entry process of HSV-1 by activating the cGAS-STING pathway to promote antiviral interferon production. These