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. 1998 Jun;72(6):5076-84.
doi: 10.1128/JVI.72.6.5076-5084.1998.

Inhibition of major histocompatibility complex class I antigen presentation in pig and primate cells by herpes simplex virus type 1 and 2 ICP47

P Jugovic  1 , A M Hill, R Tomazin, H Ploegh, D C Johnson
Affiliations

Inhibition of major histocompatibility complex class I antigen presentation in pig and primate cells by herpes simplex virus type 1 and 2 ICP47

P Jugovic et al. J Virol. 1998 Jun.

Abstract

Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) express an immediate-early protein, ICP47, that effectively inhibits the human transporter associated with antigen presentation (TAP), blocking major histocompatibility complex (MHC) class I antigen presentation to CD8+ T cells. Previous work indicated that the mouse TAP is relatively resistant to inhibition by the HSV-1 and HSV-2 ICP47 proteins (ICP47-1 and ICP47-2) and that mouse cells infected with HSV-1 are lysed by anti-HSV CD8+ cytotoxic T lymphocytes (CTL). Therefore, mice are apparently not suitable animals in which to study the in vivo effects of ICP47. In order to find an animal model, we introduced ICP47-1 and ICP47-2 into cells from various animal species-mice, rats, guinea pigs, rabbits, dogs, pigs, cows, monkeys, and humans-and measured TAP activity in the cells. Both proteins were unable to inhibit TAP in mouse, rat, guinea pig, and rabbit cells. In contrast, ICP47-1 and ICP47-2 inhibited TAP in pig, dog, cow, and monkey cells, and the TAP in pig and dog fibroblasts was often more sensitive to both proteins than TAP in human fibroblasts. These results were extended by measuring CD8+-T-cell recognition (CTL lysis) of cells from various species. Cells were infected with recombinant HSV-1 constructed to express murine MHC class I proteins so that the cells would be recognized and lysed by well-characterized murine anti-HSV CTL unless antigen presentation was blocked by ICP47. Anti-HSV CD8+ CTL effectively lysed pig and primate cells infected with a recombinant HSV-1 ICP47- mutant but were unable to lyse pig or primate cells infected with a recombinant HSV-1 that expressed ICP47. Therefore, pigs, dogs, and monkeys may be useful animal models in which to test the effects of ICP47 on HSV pathogenesis or the use of ICP47 as a selective immunosuppressive agent.

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Figures

FIG. 1
FIG. 1
Effects of ICP47-1 and ICP47-2 on TAP-mediated transport in human and monkey cells. Streptolysin-permeabilized cells were incubated with ICP47-1 (open circles) and ICP47-2 (filled squares) for 5 min at 4°C, after which a 125I-labelled peptide library was added for a further 10 min at 37°C. The transport assay was terminated, and peptides that had been glycosylated after transport into the lumen of the ER were quantified by measuring binding to concanavalin A. The level of transport measured in the absence of ICP47 was arbitrarily set at 100%.
FIG. 2
FIG. 2
Effects of ICP47-1 and ICP47-2 on TAP in rodent cells. Permeabilized skin fibroblasts were incubated with ICP47-1 (open circles) and ICP47-2 (filled squares), after which assays of TAP-mediated peptide transport were performed as described for Fig. 1.
FIG. 3
FIG. 3
Inhibition of TAP by ICP47-1 and ICP47-2 in bovine, porcine, and canine cells. Permeabilized skin fibroblasts were incubated with ICP47-1 (open circles) and ICP47-2 (filled squares), after which TAP transport assays were performed as described for Fig. 1.
FIG. 4
FIG. 4
Construction of recombinant HSV-1 able to express murine MHC class I proteins and not ICP47. (A) Diagram depicting the genome of HSV-1. An HSV-1 recombinant, F-US5MHC, able to express the murine MHC class I heavy-chain and β2-microglobulin (β2-m) proteins, was previously described (18). In the US5 gene in F-US5MHC, the murine β2-microglobulin gene is coupled to the HSV-1 thymidine kinase (tk) promoter and the simian virus 40 (SV40) polyadenylation site, and the class I (H-2Kb) heavy-chain gene is coupled to the HSV-1 ICP6 promoter. HSV-1 mutants unable to express ICP47, F-ICP47Δ (5), or ICP47 and the neighboring US11 gene, R3631 (11), have also been described. F-ICP47Δ has a deletion of the N-terminal region of the ICP47 protein, while R3631 has a larger deletion that abolishes expression of ICP47 and US11. Vero cells were coinfected with F-US5MHC and either F-ICP47Δ or R-3631, so that recombination occurred in the infected cells. Viruses able to express murine class I proteins but unable to express ICP47, F-US5MHCΔ47 (derived from F-ICP47Δ), and F-US5MHCΔUS11/12 (derived from R3631) were isolated from these coinfections. (B) Expression of ICP47 and MHC class I proteins was assessed in cells infected with the recombinant viruses by infecting R970 cells, labelling the cells with [35S]methionine, and then simultaneously immunoprecipitating ICP47 and MHC class I proteins from cell extracts. The HSV immunoglobulin G Fc receptor composed of gE and gI was precipitated by the rabbit immunoglobulin G present in the anti-ICP47 and anti-MHC class I serum.
FIG. 5
FIG. 5
CTL lysis of human and mouse fibroblasts infected with recombinant HSV-1 expressing murine class I proteins. Fibroblasts were labelled with 51Cr for 8 to 12 h and infected for 3 h either with F-US5MHC, which expresses mouse class I (H-2Kb) proteins and ICP47, or with F-US5MHCΔ47 or F-US5MHCΔUS11/12, both of which express mouse class I proteins but not ICP47, or were left uninfected. Lymphocytes taken from lymph nodes of HSV-infected C57BL/6 (H-2Kb) mice were added to wells containing the 51Cr-labelled target cells for 4 h at 37°C. Release of 51Cr was measured and specific cell lysis was calculated.
FIG. 6
FIG. 6
Lysis of pig cells infected with recombinant HSV-1 expressing murine class I proteins by mouse anti-HSV CTL. Cells were labelled with 51Cr and infected with F-US5MHC, F-US5MHCΔ47, or F-US5MHCΔUS11/12 for 3 h or left uninfected. Lymphocytes were taken from lymph nodes of HSV-infected C57BL/6 mice and were added to wells containing the target cells for 4 h at 37°C. Release of 51Cr was measured and specific cell lysis was calculated as described in Materials and Methods.
FIG. 7
FIG. 7
Lysis of monkey cells infected with recombinant HSV-1 expressing murine class I proteins by mouse anti-HSV CTL. Cells were labelled with 51Cr and infected with F-US5MHC, F-US5MHCΔ47, or F-US5MHCΔUS11/12 for 3 h or left uninfected. Lymphocytes taken from lymph nodes of HSV-infected C57BL/6 mice were added to wells containing the target cells for 4 h at 37°C. Release of 51Cr was measured and specific cell lysis was calculated as described in Materials and Methods.
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