BVDV NS5A Binds to CKAP2 and Activates the PI3K/AKT/mTOR Pathway to Facilitate Virus Transmission Through Tunneling Nanotubes

Bovine viral diarrhea virus (BVDV), a significant global pathogen threatening cattle industries worldwide, presents substantial challenges for disease control. Its ability to infect cattle across all age groups, coupled with incompletely understood transmission mechanisms, complicates prevention and treatment strategies. We previously reported that BVDV induced tunneling nanotubes (TNTs)—F-actin-rich cytoplasmic connections between adjacent cells—and utilizes these structures for intercellular transmission. In this study, we used lentiviral transfection to express various structural and non-structural proteins of BVDV and identified NS5A as a critical viral protein that induces the formation of TNTs. RNA-seq analysis revealed that CKAP2, a host protein, plays a key role in TNT generation, with the PI3K/AKT/mTOR signaling pathway being essential for this process. Further investigation demonstrated that CKAP2 interacts with BVDV NS5A, triggering the activation of the PI3K/AKT/mTOR pathway, thereby promoting TNT formation and enhancing viral dissemination. Our data highlight a previously unknown mechanism of BVDV spreading and replication, which could have significant implications for within-host spread and immune evasion.