Role of herpes simplex virus 1 protein ICP47 in antigen presentation and pathogenesis
Creators: name: Velusamy, Thilaga
Publisher: name: The Australian National University
Year: 2020

description: The herpes simplex virus (HSV) immunomodulatory protein, ICP47, conceals infected cells from CD8+ T cells by inhibiting the presentation of peptides on MHC class I. The mechanism by which ICP47 exerts this function is by binding to the transporter associated with antigen processing (TAP) protein, blocking peptide transport and loading onto MHC I molecules in the ER. The earliest studies of ICP47 supported by biochemical and in vitro observations noted marked species specificity with human but not mouse TAP being inhibited by this protein. However, later work demonstrated that ICP47 can contribute to HSV neurovirulence in mice. The discordance between biochemical and in vivo data leaves our understanding of ICP47 and its role in evading CD8+ T cells incomplete. Data from our laboratory suggested that ICP47 is likely to be expressed during the establishment and maintenance of HSV-1 latency, however, its exact function during these stages of infection is unknown. Therefore, in this study, we sought to re-visit the discrepancies discussed above and investigate the role of ICP47 during HSV-1 infection. We utilised different strains of HSV and mice, as well as an alternate infection model and unique methods to quantify the effect of ICP47 on levels of antigen presentation. In our mouse model, where HSV is confined to the peripheral nervous system, deletion of ICP47 from HSV-1 KOS did not alter lesion development, virus load, spread or reactivation. Likewise, latency was unaffected by ICP47 deficiency as determined using a sensitive Cre-marking mouse model. Further observations from the Cre-marking mouse model revealed that unlike the ICP47 promoter inserted in an ectopic locus, native promoters did not induce additional neuronal marking by Cre beyond lytic infection. We evaluated the reasons behind the difference in marking using newly generated recombinant viruses. Subsequent flank infection of ROSA26R mice with these viruses showed that the local genomic context is also important for regulation of gene expression. By contrast to our in vivo pathogenesis data, we were able to show that ICP47 does inhibit antigen presentation significantly on HSV-infected mouse cells using in vitro antigen presentation assays. However, in mouse cells, antigen presentation was ablated by 44%, compared to an 85% reduction in human cells. As CD8+ T cells have been shown to recognize very few peptide-MHC I complexes on the surface of target cells, it is important to consider the efficiency at which ICP47 inhibits human and mouse TAP. Therefore, we used mass spectrometry to identify and quantify MHC I bound peptides derived from HSV-1 during viral infection. We found that more peptide sequences were presented on mouse cells infected with ICP47 null virus compared to those infected with wild-type virus. We quantified the presentation of 14 of these peptides and the contribution of ICP47 to this process in human and mouse cells. We found that ICP47 almost entirely blocks human TAP-mediated peptide presentation, though the degree of inhibition was somewhat peptide-specific. Conversely, the effect of ICP47 on mouse TAP was far less profound, resulting in only up to five-fold reduction in MHC-peptide abundance. In conclusion, this study shows that despite significant inhibition of antigen presentation in mouse cells, ICP47 may not be an effective immune modulator in mice and suggests a need for re-evaluation of suitable mouse models.