African swine fever virus multigene family interferon inhibitory proteins: functions and applications to vaccine development

Thesis

African swine fever (ASF) poses a significant threat to global food security, with no commercially available vaccines against this viral disease. We investigated how African swine fever virus (ASFV) multigene family (MGF) proteins modulate hostinnate immunity, focusing on their role in type-I interferon (IFN) suppression. We characterised the impact of various MGF gene deletions on viral replication and immune responses in primary macrophages, corroborating previously published data that showed deletion of MGF360-12L (12L) significantly enhancing interferonstimulated gene expression early in infection. Our assessment interferon alpha (IFN-α) levels in virus stocks revealed that an excess of interferon is carried over into subsequent experiments; we resolved this contingency by using ultracentrifuged virus in our experiments. Interestingly, we showed that the growth defect observed for the GeorgiaΔK145RΔMGF360-12L (Δ12L) deletion mutant does not seem to correlate with its inability to control the host IFN response. Through molecular in vitro assays, we demonstrated that 12L inhibits interferon regulatory factor 7 (IRF7)-dependent interferon alpha-6 (IFNα6)-promoter activity and promotes proteasomal degradation of IRF7 in a dose-dependent manner, through a mechanism dependent on its putative VHL-box motif. Additionally, 12L disrupts the nuclear translocation of this transcription factor. This represents a novel strategy for IFN induction inhibition by ASFV. Additionally, we evaluated the potential of a novel immortalized porcine macrophage cell line – porcine large tumour antigen 58 (PLTA58) – as an alternative to primary cells for ASFV research. Whilst PLTA58 cells supported enhanced viral replication, they failed to replicate key viral phenotypes observed during infection of primary macrophages, likely due to differences in cellular diversity and molecular phenotypes. Our findings advance understanding of ASFV immune evasion mechanisms and provide insights for rational vaccine design through targeted gene deletion, while also assessing new tools to reduce reliance on animal-derived cells in ASFV research.

Authors: Gonzalez Gomez, R

• DOI: 10.5287/ora-aq9k7nnp0
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: interferon; MGF360-12L; ASFV; multigene family; African swine fever virus; innate immune evasion; animal health; food security; infection; vaccine development; veterinary medicine; molecular biology; molecular virology
• Subjects: Infection; Molecular virology; Food security; Animal health; African swine fever virus; Virology; Veterinary virology