The interaction between the DOCK7 protein and the E2 protein of classical swine fever virus is not involved with viral replication or pathogenicity

Authors: VUONO, ELIZABETH - Mississippi State University; Ramirez Medina, Elizabeth; Silva, Christina; BERGGREN, KEITH - Oak Ridge Institute For Science And Education (ORISE); AYUSHI, RAI - Oak Ridge Institute For Science And Education (ORISE); Espinoza, Nallely; Borca, Manuel; Gladue, Douglas

Submitted to: Viruses
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/25/2023
Publication Date: 12/30/2023
Citation: Vuono, E., Ramirez Medina, E., Silva, C.M., Berggren, K., Ayushi, R., Espinoza, N.N., Borca, M.V., Gladue, D.P. 2023. The interaction between the DOCK7 protein and the E2 protein of classical swine fever virus is not involved with viral replication or pathogenicity. Viruses. 16(1). Article 70. https://doi.org/10.3390/v16010070.
DOI: https://doi.org/10.3390/v16010070

Interpretive Summary: Classical swine fever virus (CSFV) causes a devastating disease in swine, called Classical swine fever (CSF), that is currently causing disease in Central and South America, Europe, and Asia and parts of Africa. Understanding the virus-host protein interactions is necessary to understand the pathogenesis of CSF and the differences in virulence between different CSFV strains. Understanding this information will advance the development of next generation vaccines and antivirals.

Technical Abstract: The classical swine fever virus (CSFV) particle consists of three glycoproteins, all of which have been shown to be important proteins involved in many virus functions including interaction with several host proteins. One of these proteins, E2, has been shown to be directly involved with adsorption to host cell and important for virus virulence. Using the yeast two-hybrid system, we have previously shown that CSFV E2 specifically interacts with (DOCK7) dedicator of cytokinesis, a scaffolding protein. In this report, the interaction between E2 and DOCK7 was evaluated. To confirm the yeast-two-hybrid results and to determine that DOCK7 interacts in swine cells with E2, we performed co-immunoprecipitation and proximity ligation assay (PLA). After demonstrating the protein interaction in swine cells, E2 amino acid residues M49 and P130 were determined to be critical for interaction with DOCK7. This was established by using a random mutated library of E2 and a reverse yeast two-hybrid approach. These two residues were then introduced into a recombinant CSFV, E2DOCK7v, by a reverse genomics approach using the highly virulent CSFV Brescia isolate as a backbone. E2DOCKv was shown to have similar growth kinetics in swine primary macrophages and SK6 cells cultures to the parental Brescia strain. Similarly, E2DOCK7v demonstrated a similar level of virulence to the parental Brescia when inoculated in domestic pigs. Animals intranasally inoculated with 105 TCID50 developed a lethal form of clinical disease with virological and hematological kinetics changes indistinguishable from that produced by the parental strain. Therefore, interaction between CSFV E2 and host DOCK7 is not critically involved in the process of virus replication and disease production.