MAJOR histocompatibility complex (MHC) class I molecules present peptides derived from the endogenous protein pool to cytotoxic T lymphocytes, which can thus recognize intracellular antigen1-3. This pathway may depend on a transporter (PSF1) (refs 4-6) to mediate entry of the cytosolic peptides into a pre-Golgi compartment where they hind to class I heavy chains and promote their stable assembly with β2-microglobulin7-12. There is, however, only indirect support for this function of PSF1 (ref. 6). Here we show that PSF1 is necessary for the efficient assembly of class I molecules and enables them to present a peptide epitope derived from endogenously synthesized viral antigen. Immunochemical and genetic data demonstrate that the PSF1 poly peptide is associated with a complementary transporter chain, which is polymorphic and is encoded by the PSF2 gene13, which is closely linked to PSF1.