Epstein-Barr virus (EBV) transformed B lymphoblastoid cell lines (LCL) efficiently process exogenous
antigens for MHC class II-restricted presentation via the
chloroquine-sensitive endosomal pathway. Using MHC class II-restricted T cell clones specific for EBV structural
proteins, however, we frequently observed significant responses to autologous LCL cells without the addition of exogenous virus. Such responses were reduced by pre-treating the LCL with
acyclovir (ACV), a
drug blocking productive
EBV infection. This suggested T cell recognition of
antigen synthesized by LCL cells spontaneously entering virus productive cycle, and led us to question by what route(s) MHC class II-restricted presentation of endogenously synthesized virion
proteins was occurring. Cell sorting experiments, using the viral envelope
glycoprotein gp340 as a surface marker of productively-infected cells, confirmed that stimulatory activity lay within the gp340-positive fraction. However, closer analysis revealed that most of these cells were not productively-infected but were EBV receptor-positive and had bound released virus. We infer that receptor-mediated delivery of released virus into the endosomal pathway is one route whereby an LCL can present endogenously synthesized EBV
proteins on
MHC class II molecules. To ask whether another, more direct, route of processing was possible, we used a recombinant
vaccinia viral vector to express gp340 de novo in ACV-treated LCLs. Significantly, these cells presented the endogenously synthesized
antigen to autologous gp340-specific T cell clones via a
chloroquine-resistant pathway. In the same experiments,
vaccinia-mediated expression of a
signal peptide-deleted form of gp340 did not lead to T cell stimulation, suggesting that this second route of processing required entry of endogenously synthesized
antigen into the endoplasmic reticulum.