An early pseudorabies virus protein down-regulates porcine MHC class I expression by inhibition of transporter associated with antigen processing (TAP)

The objectives of this study were to identify the mechanism(s) of pseudorab ies virus (PrV)-induced down-regulation of porcine class I molecules and th e viral protein(s) responsible for the effect. The ability of PrV to interf ere with the peptide transport activity of TBP tvas determined by an in vit ro transport assay. In this assay, porcine kidney (PK-15) cells were permea bilized with streptolysin-O and incubated with a library of I-125-labeled p eptides having consensus moths for glycosylation in the endoplasmic reticul um (ER). The efficiency of transport of peptides from the cytosol into the ER was determined by adsorbing the ER-glycosylated peptides onto Con A-coup led Sepharose beads. Dose-dependent inhibition of TAP activity was observed in PrV-infected PK-15 cells. This inhibition,,which occurred as early as 2 h postinfection (h.p.i.), reached the maximum level by 6 h.p.i., indicatin g that TAP inhibition is one of the mechanisms by which PrV down-regulates porcine class I molecules. Infection of cells with PrV in the presence of m etabolic inhibitors revealed that cycloheximide a protein synthesis inhibit or, but not phosphonoacetic acid a herpesvirus DNA synthesis inhibitor, cou ld restore the cell surface expression of class I molecules, indicating tha t late proteins are not responsible for the down-regulation. Infection in t he presence of cycloheximide followed by actinomycin-D, which results in ac cumulation of the immediate-early protein, failed to down-regulate class I, indicating that one or more early proteins are responsible for the down-re gulation of class I molecules.