I329L: A Dual Action Viral Antagonist of TLR Activation Encoded by the African Swine Fever Virus (ASFV)

Abstract

The African Swine Fever Virus (ASFV) is an economically important, large DNA virus which causes a highly contagious and frequently fatal disease in domestic pigs. Due to the acute nature of the infection and the complexity of the protective porcine anti-ASFV response, there is no accepted vaccine in use. As resistance to ASFV is known to correlate with a robust IFN response, the virus is predicted to have evolved strategies to inhibit innate immunity by modulating the IFN response. The deletion of virus host evasion gene(s) inhibiting IFN is a logical solution to develop an attenuated virus vaccine. One such candidate, the ASFV ORF I329L gene, is highly conserved in pathogenic and non-pathogenic virus isolates and in this study we confirm and extend the conclusion that it has evolved for the inhibition of innate immunity initiated through Toll-like receptors (TLRs). Specifically, the ASFV I329L extracellular (ECD) and intracellular (ICD) domains inhibit TLR signalling by two entirely different mechanisms. Bioinformatics modelling suggests that the ECD inhibits several TLR signalling pathways through a short sequence homologous to the conserved TLR dimerization domain, here termed the putative dimerization domain (PDD). Remarkably, both full length and PDD constructs of I329L were demonstrated to inhibit activation, not only of TLR3, but also TLR4, TLR5, TLR8 and TLR9. Additionally, the demonstration of a weak association of I329L with TLR3 is consistent with the formation of a non-signalling I329L-TLR3 heterodimer, perhaps mediated through the PDD of I329L. Finally, the ICD associates with TRIF, thereby impacting on both TLR3 and TLR4 signalling. Thus, I329L offers potential as a general inhibitor of TLR responses and is a rational candidate for construction and testing of an I329L deletion mutant vaccine.

Immunization of pigs with replication-incompetent adenovirus-vectored African swine fever virus multi-antigens induced humoral immune responses but no protection following contact challenge

“…This study evaluated the safety, tolerability, and efficacy of experimental subunit vaccines that were formulated, with and without adjuvant, using a cocktail of 42 replication-deficient adenovirus-vectored multicistronic expression cassettes encoding ASFV antigens. As protective ASFV antigens are yet to be identified, immunization with a cocktail of the multicistronic expression cassettes was a strategy to deliver nearly 100% of the ASFV Georgia 2007/1 proteome to mimic antigen delivery by some ASFV mutants that have been shown to confer protection and if successful, this approach could fast track identification of the protective proteins needed for the development of rationally designed prototype subunit vaccines (8)(9)(10)(11)(12)(13)42). The multicistronic expression cassettes were stable after multiple passages of the recombinant adenoviruses as judged by expression of the FLAG tag at the C-terminal of the last gene in the cassette.…”

I329L protein-based indirect ELISA for detecting antibodies specific to African swine fever virus

“…TheI329L protein is a type I transmembrane protein with 329 amino acids, which is relatively conserved in sequence and is expressed in late-stage viral infection ( Correia et al., 2023 ). Additionally, the I329L protein is homologous to the TLR family of proteins and its intracellular amino acid sequence shows 35% similarity to the BOX1 and BOX2 sequences of TLR3.…”

Dihydromyricetin inhibits African swine fever virus replication by downregulating toll-like receptor 4-dependent pyroptosis in vitro

“…In addition, TLR3 and TLR7 were upregulated after ASFV infection, whereas TLR4 and TLR6 were downregulated [ 16 ]. Another study reported that ASFV I329L inhibits the activation of TLR3, TLR4, TLR5, TLR8, and TLR9 [ 17 ]. Moreover, Li et al showed that the ASFV-induced IL-1β production is dependent on TLR4 [ 18 ]; however, the role of TLR4 signaling in ASFV infection has not been reported.…”