The generation of many HLA class I peptides entails a final trimming step in the endoplasmic reticulum that, in humans, is accomplished by two 'candidate' aminopeptidases. We show here that one of these, ERAP1, was unable to remove several N-terminal amino acids that were trimmed efficiently by the second enzyme, ERAP2. This trimming of a longer peptide required the concerted action of both ERAP1 and ERAP2, both for in vitro digestion and in vivo for cellular antigen presentation. ERAP1 and ERAP2 localized together in vivo and associated physically in complexes that were most likely heterodimeric. Thus, the human endoplasmic reticulum is equipped with a pair of trimming aminopeptidases that have complementary functions in HLA class I peptide presentation.

The recognition of short viral peptides associated with human histocompatibility complex (human leukocyte antigen (HLA) 1 ) class I molecules on the cell surface allows cytotoxic T lymphocytes (CTLs) to recognize and kill virus-infected cells (1). These peptides are generated by proteolytic processing of newly synthesized viral proteins in the cytosol by the combined action of proteasomes, ERAAP (endoplasmic reticulum aminopeptidase associated with antigen processing), and in some cases other peptidases (2). This degradation of viral proteins generates peptides of 8 -11 residues that are translocated to the endoplasmic reticulum lumen by transporters associated with antigen processing. These short peptides then assemble with the HLA class I heavy chain and ␤ 2 -microglobulin. Usually, two major anchor residues in the antigenic peptide, at position 2 and the C terminus (3, 4), must be deeply accommodated into specific pockets of the antigen recognition site of the HLA class I molecule to stabilize the nascent complexes (5, 6) and allow for their subsequent transport to the cell membrane where they are exposed for CTL recognition (7).Human respiratory syncytial virus (HRSV) (8), a member of the Paramyxoviridae family, is the single most important cause of bronchiolitis and pneumonia in infants and young children (9 -11). Infections of this virus occur in people of all ages, but although usually mild infections are reported in healthy adults, HRSV poses a serious health risk in immunocompromised individuals (12, 13) and in the elderly (14, 15). The single-stranded, negative-sense RNA genome of this enveloped virus codes for 11 proteins.Although the immune mechanism involved in HRSV disease and protection is not well understood, specific CD8 ϩ T lymphocytes are required for the clearance of virus-infected cells (16). Previously, several HRSV epitopes restricted by different HLA class I molecules were identified using CTLs from seropositive individuals (17-21). However, these experiments were performed with synthetic peptides against individual proteins. In contrast, only one published study attempted to elucidate the nature and diversity of the possible array of HRSV ligands restricted by individual HLA molecules (22). In this study, virus-infected cells were cultured with stable, isotope-labeled amino acids, which were expected to act as anchor residues for the HLA allele of interest. The MHC molecules were then immunoprecipitated, and mass spectrometry analysis was performed. This study identified one HRSV ligand for each of the HLA-A2 and -B7 class I molecules (22). Therefore, is only one HRSV ligand restricted by a single HLA molecule exposed on the cell membrane surface as suggested by this study? Conversely, could a particular HLA molecule bind several ligands of this small virus simultaneously? To answer these questions, we compared HLA-B27From the ‡Unidad

The transporter associated with antigen processing (TAP) delivers the viral proteolytic products generated by the proteasome in the cytosol to the endoplasmic reticulum lumen that are subsequently recognized by cytotoxic T lymphocytes (CTLs).