The expression and function of HSV ICP47 and its promoter in mice

Abstract

Herpes simplex virus (HSV) protein immediate early-infected cell protein (ICP)47 inhibits the transporter associated with antigen presentation, thereby concealing infected cells from the CD8 + T-cell response in humans. By contrast, in mouse cells, ICP47 was found to be ineffective in vitro , despite some evidence for a role of this protein in vivo , specifically in the nervous system. Recent evidence suggests that the promoter that drives ICP47 might be active in the establishment of latency and during latency, leading to the possibility that a role might be found in protecting neurons that are latently infected with HSV from T-cell attack. Here, we explored this idea by studying a set of recombinant HSVs that were null for ICP47 protein and/or express Cre recombinase from ICP47 promoters, for use in a Cre-marking mouse model. Using these viruses, we found that contrary to previous studies, the activity of ICP47 in mouse cells is efficient enough to detectably reduce antigen presentation, albeit at levels much lower than in human cells. However, in vivo , we found no evidence of ICP47 influencing HSV pathogenesis or latency in a model where the virus does not spread beyond the peripheral nervous system. Finally, we show that the ability of the ICP47 promoter to drive protein expression during HSV latency is dependent on the location that it is placed in a recombinant virus genome and that such expression is undetectable from its native locus. IMPORTANCE Immune evasion and latency are key mechanisms that underlie the success of herpesviruses. In each case, interactions between viral and host proteins are required and due to co-evolution, not all mechanisms are preserved across host species, even if infection is possible. This is highlighted by the herpes simplex virus (HSV) protein immediate early-infected cell protein (ICP)47, which inhibits the detection of infected cells by killer T cells and acts with high efficiency in humans, but poorly, if at all in mouse cells. Here, we show that ICP47 retains modest but detectable function in mouse cells, but in an in vivo model we found no role during acute infection or latency. We also explored the activity of the ICP47 promoter, finding that it could be active during latency, but this was dependent on genome location. These results are important to interpret HSV pathogenesis work done in mice.

Multi-targeted loss of the antigen presentation molecule MR1 during HSV-1 and HSV-2 infection

“…To test whether expression of ICP47 during infection impacts MR1 antigen presentation, ARPE-19 epithelial cells were infected with HSV-1 KOS strain and ICP47 null mutant ICP47del 48 and analyzed by flow cytometry. Equivalent surface expression of the late viral glycoprotein gC was detected at 6 h post infection (p.i.)…”

“… Rawls et al., 1968 114 N/A HSV-1 Strain 17 vhs mutant 17(41-) Prof Roger Everett (University of Glasgow). Fenwick and Everett, 1990 79 N/A HSV-1 Strain KOS US12 mutant ICP47del Velusamy et al., 2023 48 N/A SW102 E. coli containing the pAdZ5-C5 vector Stanton et al., 2008 67 N/A 5-alpha Competent E. coli NEB Cat#C2987H Chemicals, peptides, and recombinant proteins Dulbecco’s Modified Eagle’s Medium Lonza Cat#12-604F Hanks Balanced Salt Solution Gibco Cat#14170 Fetal Calf Serum Cytiva Cat#SH30084.02 Normal donkey serum Sigma-Aldrich Cat#D9663 Ac-6-FP Acetyl-6-formylpterin Schircks Cat#11.418 BD Cytofix™ Fixation buffer BD Biosciences Cat#554655 Folimycin (concanamycin A) Sigma-Aldrich Cat#C9705 MG132 Sigma-Aldrich Cat#M7449 Fugene HD Promega Cat#E2231 Q5 High-Fidelity polymerase NEB Cat#M0491 Endo H NEB Cat#P0703S DAB Substrate Pierce Cat#34065 BamHI-HF NEB Cat#R3136 SpeI-HF NEB Cat#R3133 BglII NEB Cat#R0144 XbaI NEB Cat#R0145 Xgal Abcam Cat#ab144388 IPTG Abcam ...…”

“…HSV-1 and HSV-2 strains used in this study were HSV-2 strain 186 (courtesy Dr Naomi Truong, The Westmead Institute for Medical Research), 114 HSV-1 strain F (courtesy Dr Russell Diefenbach, Macquarie University), HSV-1 17syn+ (Prof Roger Everett, University of Glasgow), HSV-1 17syn+ vhs mutant 17(41-) (Prof Roger Everett, University of Glasgow), 79 HSV-1 KOS (courtesy Prof P Kinchington, University of Pittsburgh) and US12 (ICP47) null mutant ICP47del. 48 …”