Insights into the Role of VPS39 and Its Interaction with CP204L and A137R in ASFV Infection

Abstract

The African swine fever virus (ASFV) is a large and complex DNA virus that causes a highly lethal disease in swine, for which no antiviral drugs or vaccines are currently available. Studying viral-host protein-protein interactions advances our understanding of the molecular mechanisms underlying viral replication and pathogenesis and can facilitate the discovery of antiviral therapeutics. In this study, we employed affinity tagging and purification mass spectrometry to characterize the interactome of VPS39, an important cellular factor during the early phase of ASFV replication. The interaction network of VPS39 revealed associations with mitochondrial proteins involved in membrane contact sites formation and cellular respiration. We show that the ASFV proteins CP204L and A137R target VPS39 by interacting with its clathrin heavy-chain functional domain. Furthermore, we elaborate on the potential mechanisms by which VPS39 may contribute to ASFV replication and prioritize interactions for further investigation into mitochondrial protein function in the context of ASFV infection.

Keywords: A137R; CP204L; VPS39; african swine fever virus; protein–protein interaction; virus–host interaction.

Figure 1

The interaction of ASFV proteins CP204L and A137R with the host protein VPS39. (A) Subcellular localization of GFP-tagged A137R and endogenous CP204L in WSL cells 24 h after ASFV infection. (B) Colocalization of CP204L-GFP and A137R-GFP with VPS39 in WSL cells. Scale bars, 10 µm. (C) Diagram of the different truncation FLAG-tagged fragments and the predicted domain organization of porcine VPS39 protein. The co-immunoprecipitation of FLAG-tagged VPS39 fragments with CP204L and A137R in WSL cells (D) 24 h after GFP-CP204L or A137R-GFP plasmid transfection and (E) 24 h after ASFV infection. Representative immunoblots of whole-cell lysates (WCLs) and GFP immunoprecipitates (IPs) are shown. α-tubulin was used as a loading control in WCLs.

Figure 2

Network illustrating the interactions between VPS39-associated proteins localized in mitochondria. Proteins were grouped into protein complexes according to the EBI Complex Portal and visualized with STRING to illustrate the experimentally validated PPIs among them.