Dalin He, Jing Yang, Yitong Cui, Bingrong Wu, Feng Wei, Siming Zhu, Youxiang Diao, Yi Tang
Abstract Accumulating data support the role of exosomes from virus-infected cells (especially RNA viruses) in the selective encapsulation of viral proteins, genomic material, and even whole virions, thereby mediating intercellular communication events and contributing to viral transmission. Tembusu virus (TMUV), a single-stranded RNA virus, has posed a threat to global public health since its emergence in 2010. However, whether exosomes can mediate TMUV intercellular transmission remains obscure. Here, we show that exosomes derived from TMUV-infected cells (TMUV-exosomes) contained viral genomic RNA and partial viral proteins (C, prM, E, NS1, NS2B, NS4B, and NS5). TMUV-exosomes enter HEK293 cells primarily via a dynamin-dependent, cholesterol-sensitive caveolae-mediated (CavME) endocytosis pathway, thereby suppressing the subsequent expression of key antiviral immune genes to create a proviral microenvironment that supports productive TMUV infection. Remarkably, central to this exosomal hijacking mechanism, the viral NS4A protein and host Rab27a protein form a critical regulatory axis. Specifically, NS4A upregulates Rab27a expression to potentiate the release of exosomes encapsulating viral nucleic acids and proteomic components. Overall, our study provides a mechanistic model for understanding the role of exosomes in TMUV intercellular transmission and identifies the NS4A–Rab27a axis as a critical regulatory node. Disrupting this interaction may represent a potential therapeutic strategy for TMUV infection, warranting further investigation.