Flavivirus infection of mammalian cells increases the ceil SUrfaCe expression of major histocompatibility complex (MHC) class I molecules, the recognition elements for cytotoxic T cells. Here, we show that the mechanism for flavivirudnduced upregulation of class I MHC involves an increase in peptide supply to the endoplasmic reticulum. Flavivirus-mediated peptide supply for MHC class I assembly is independent of the peptlde transporters for class I antigen presentation, since infection of class I YHC peptide transport-deficient cell lines with flaviviruses results in the cell surface expression of biologically functional class I MHC peptlde complexes. The flavivirus-induced supply of antigenic peptides to the endoplasmic reticulum is not restricted to flavlvirus-encoded peptldes and independent of interferon. The data imply that peptide availability regulates surface expression of class I MHC restriction elements and suggests a mechanism for flavivirus-induced immunopathology.