Japanese Encephalitis Virus NS1 Protein regulates the expression and distribution of TIM-1 to promote viral infection

Abstract

Japanese encephalitis virus (JEV) is a mosquito-borne Flavivirus, which may cause severe encephalitis in humans, horses, and other animals. TIM-1 has been identified to be a receptor that promotes various viruses to enter into target cells in recent years. In the present study, we found that TIM-1 protein was significantly increased in A549 cells at the late stage of JEV infection, while the transcription levels of TIM-1 remained unaltered. Interestingly, we found that NS1 protein plays a key role in increasing the expression of TIM-1 in cells infected with JEV. Further, we found that the NS1 protein also efficiently regulates TIM-1 protein to distribute in the cytoplasm in JEV-infected cells, and the amount of TIM-1 protein located on the cell membrane was reduced instead. As a consequence, NS1 protein antagonize TIM-1 mediated viral restriction for further viral infection and propagation in the late stage of infection. In molecular mechanism, the molecular weight of TIM-1 increased a bit in the present of NS1. Expression of NEU1 down-regulated TIM-1 expression and oseltamivir treatment increased the expression of TIM-1. Therefore, our data indicated that JEV NS1 protein facilitated the sialylation modification of TIM-1 to up-regulate the level of TIM-1 expression and regulate its distribution. Collectively, our study revealed that JEV NS1 protein regulates the expression and distribution of TIM-1 by facilitating its sialylation to antagonize TIM-1-mediated JEV restriction for further infection. Understanding the functional interplays between TIM-1 and NS1 proteins will offer new insights into virushost interaction. IMPORTANCET-cell immunoglobulin and mucin domain protein 1 (TIM-1) is known to promote cellular entry of various enveloped viruses. We discovered a novel phenomenon of dynamic expression and functional regulation of TIM-1 during JEV infection. Firstly, TIM-1 protein increased in the cytoplasm at the late stage of JEV infection. Furthermore, JEV NS1 protein up-regulated the intracellular expression of TIM-1 and antagonize TIM-1-mediated viral restriction for further viral infection. In molecular mechanism, JEV NS1 protein facilitated the sialylation of TIM-1 to increase the level of TIM-1 expression and cytoplasm distribution. Taken together, at the late stage of infection, JEV employs a strategy in which NS1 promotes JEV release by promoting the sialylation of TIM-1, causing it to be primarily localized in the cytoplasm. Therefore, we discovered new functions of TIM-1 and JEV NS1 during the process of JEV infection, and provide a new insight into the interactions between JEV and cell hosts.