Abstract: The fusion protein of VP35 and biotin ligase TurboID was generated, and the Ebola virus minigenome system (EBOV MG) was utilized to simulate the Ebola virus lifecycle and formation of virus inclusion bodies (VIB) in cells. The introduction of exogenous biotin enabled the labeling of proteins to interact with VP35. Among all proximity labeling (PL) labeled proteins, 537 potential VP35-associated host proteins were identified by differential abundance analysis after quantitative mass spectrometry detection. Gene Ontology (GO) analysis found that many enriched proteins were involved in RNA binding related functions. Subsequently, the association of EBOV VP35 with RNA binding proteins EIF4B and ZNF598 was confirmed. Interruption of EIF4B and ZNF598 expression significantly inhibited EBOV trVLP replication. The study highlighted the effectiveness of PL-quantitative mass spectrometry in identifying virus-host interaction proteins, providing a valuable tool for investigating viral pathogenesis and identifying potential antiviral targets.

Key words: proximity labeling, EBOV, viral inclusion bodies, TurboID, VP35